Chloroplast

Approximately twoTemplate:Nbspbillion years ago,<ref name="Milo" /><ref name="Sánchez-Baracaldo-2017"> Template:Cite journal</ref><ref>Template:Cite journal</ref> a free-living cyanobacterium entered an early eukaryotic cell, either as food or as an internal parasite,<ref name="Campbell-2009c" /> but managed to escape the phagocytic vacuole it was contained in and persist inside the cell.<ref name="Kim-2009" /> This event is called endosymbiosis, or "cell living inside another cell with a mutual benefit for both". The external cell is commonly referred to as the host while the internal cell is called the endosymbiont.<ref name="Campbell-2009c" /> The engulfed cyanobacteria provided an advantage to the host by providing sugar from photosynthesis.<ref name="Campbell-2009c" /> Over time, the cyanobacterium was assimilated, and many of its genes were lost or transferred to the nucleus of the host.<ref name="Nakayama-2012">Template:Cite journal</ref> Some of the cyanobacterial proteins were then synthesized by host cell and imported back into the chloroplast (formerly the cyanobacterium), allowing the host to control the chloroplast.<ref name="Nakayama-2012" /><ref name="McFadden-2001" />

Chloroplasts which can be traced back directly to a cyanobacterial ancestor (i.e. without a subsequent endosymbiotic event) are known as primary plastids ("plastid" in this context means almost the same thing as chloroplast<ref name="Campbell-2009c" />).<ref name="Wise-2006b" /> Chloroplasts that can be traced back to another photosynthetic eukaryotic endosymbiont are called secondary plastids or tertiary plastids (discussed below).

Whether primary chloroplasts came from a single endosymbiotic event or multiple independent engulfments across various eukaryotic lineages was long debated. It is now generally held that with one exception (the amoeboid Paulinella chromatophora), chloroplasts arose from a single endosymbiotic event around twoTemplate:Nbspbillion years ago and these chloroplasts all share a single ancestor.<ref name="Sánchez-Baracaldo-2017" /> It has been proposed this the closest living relative of the ancestral engulfed cyanobacterium is Gloeomargarita lithophora.<ref>Template:Cite journal</ref><ref>Template:Cite journal</ref><ref name="López-García-2017">Template:Cite journal</ref> Separately, somewhere about 90–140 million years ago, this process happened again in the amoeboid Paulinella with a cyanobacterium in the genus Prochlorococcus. This independently evolved chloroplast is often called a chromatophore instead of a chloroplast.<ref name="Macorano-2021">Template:Cite journal</ref><ref group="Note" name=":0">Not to be confused with chromatophore—the pigmented cells in some animals—or chromatophore—the membrane associated vesicle in some bacteria.</ref>

Chloroplasts are believed to have arisen after mitochondria, since all eukaryotes contain mitochondria, but not all have chloroplasts.<ref name="Campbell-2009c" /><ref>Template:Cite journal</ref> This is called serial endosymbiosis—where an early eukaryote engulfed the mitochondrion ancestor, and then descendants of it then engulfed the chloroplast ancestor, creating a cell with both chloroplasts and mitochondria.<ref name="Campbell-2009c" />

Secondary and tertiary endosymbiosis

Cladogram of chloroplast evolution

Many other organisms obtained chloroplasts from the primary chloroplast lineages through secondary endosymbiosis—engulfing a red or green alga with a primary chloroplast. These chloroplasts are known as secondary plastids.<ref name="Wise-2006b" />

As a result of the secondary endosymbiotic event, secondary chloroplasts have additional membranes outside of the original two in primary chloroplasts.<ref name="Keeling-2004">Template:Cite journal</ref> In secondary plastids, typically only the chloroplast, and sometimes its cell membrane and nucleus remain, forming a chloroplast with three or four membranes<ref name="Chaal-2005">Template:Cite journal</ref>—the two cyanobacterial membranes, sometimes the eaten alga's cell membrane, and the phagosomal vacuole from the host's cell membrane.<ref name="Keeling-2004" />

The genes in the phagocytosed eukaryote's nucleus are often transferred to the secondary host's nucleus.<ref name="Keeling-2004" /> Cryptomonads and chlorarachniophytes retain the phagocytosed eukaryote's nucleus, an object called a nucleomorph,<ref name="Keeling-2004" /> located between the second and third membranes of the chloroplast.<ref name="Kim-2009" /><ref name="McFadden-2001" />

All secondary chloroplasts come from green and red algae. No secondary chloroplasts from glaucophytes have been observed, probably because glaucophytes are relatively rare in nature, making them less likely to have been taken up by another eukaryote.<ref name="Keeling-2004" />

Still other organisms, including the dinoflagellates Karlodinium and Karenia, obtained chloroplasts by engulfing an organism with a secondary plastid. These are called tertiary plastids.<ref name="Wise-2006b" />

Possible cladogram of chloroplast evolution<ref name="Keeling-2004" /><ref name="McFadden-2004">Template:Cite journal</ref><ref name="Keeling-2010" /> Circles represent endosymbiotic events. For clarity, dinophyte tertiary endosymbioses and many nonphotosynthetic lineages have been omitted.

^a It is now established that Chromalveolata is paraphyletic to Rhizaria.<ref name="Keeling-2010" />

Primary chloroplast lineages

All primary chloroplasts belong to one of four chloroplast lineages—the glaucophyte chloroplast lineage, the rhodophyte ("red") chloroplast lineage, and the chloroplastidan ("green") chloroplast lineage, the amoeboid Paulinella chromatophora lineage.<ref name="Ball-2011" /> The glaucophyte, rhodophyte, and chloroplastidian lineages are all descended from the same ancestral endosymbiotic event and are all within the group Archaeplastida.<ref name="Keeling-2004" />

Glaucophyte chloroplasts

Template:See also Template:Plain image with caption The glaucophyte chloroplast group is the smallest of the three primary chloroplast lineages as there are only 25 described glaucophyte species.<ref>Template:Cite journal</ref> Glaucophytes diverged first before the red and green chloroplast lineages diverged.<ref>Template:Cite journal</ref> Because of this, they are sometimes considered intermediates between cyanobacteria and the red and green chloroplasts.<ref name="Miyagishima-2011">Template:Cite journal</ref> This early divergence is supported by both phylogenetic studies and physical features present in glaucophyte chloroplasts and cyanobacteria, but not the red and green chloroplasts. First, glaucophyte chloroplasts have a peptidoglycan wall, a type of cell wall otherwise only in bacteria (including cyanobacteria).<ref group="Note">For this reason, glaucophyte chloroplasts are also known as 'muroplasts' from the Latin muro meaning wall.</ref> Second, glaucophyte chloroplasts contain concentric unstacked thylakoids which surround a carboxysome – an icosahedral structure that contains the enzyme RuBisCO responsible for carbon fixation. Third, starch created by the chloroplast is collected outside the chloroplast.<ref name="Wise-2006">Template:Cite book</ref> Additionally, like cyanobacteria, both glaucophyte and rhodophyte thylakoids are studded with light collecting structures called phycobilisomes.

Rhodophyta (red chloroplasts)

The rhodophyte, or red algae, group is a large and diverse lineage.<ref name="Keeling-2004" /> Rhodophyte chloroplasts are also called rhodoplasts,<ref name="Wise-2006b" /> literally "red chloroplasts".<ref>Template:Cite web</ref> Rhodoplasts have a double membrane with an intermembrane space and phycobilin pigments organized into phycobilisomes on the thylakoid membranes, preventing their thylakoids from stacking.<ref name="Kim-2009" /> Some contain pyrenoids.<ref name="Wise-2006b" /> Rhodoplasts have chlorophyll a and phycobilins<ref name="Keeling-2010" /> for photosynthetic pigments; the phycobilin phycoerythrin is responsible for giving many red algae their distinctive red color.<ref name="Campbell-2009f">Template:Cite book</ref> However, since they also contain the blue-green chlorophyll a and other pigments, many are reddish to purple from the combination.<ref name="Wise-2006b" />Template:Dubious The red phycoerytherin pigment is an adaptation to help red algae catch more sunlight in deep water<ref name="Wise-2006b" />—as such, some red algae that live in shallow water have less phycoerythrin in their rhodoplasts, and can appear more greenish.<ref name="Campbell-2009f" /> Rhodoplasts synthesize a form of starch called floridean starch,<ref name="Wise-2006b" /> which collects into granules outside the rhodoplast, in the cytoplasm of the red alga.<ref name="Kim-2009" />

Chloroplastida (green chloroplasts)

The chloroplastida group is another large, highly diverse lineage that includes both green algae and land plants.<ref name="Lewis-2004">Template:Cite journal</ref> This group is also called Viridiplantae, which includes two core clades—Chlorophyta and Streptophyta.

Most green chloroplasts are green in color, though some aren't due to accessory pigments that override the green from chlorophylls, such as in the resting cells of Haematococcus pluvialis. Green chloroplasts differ from glaucophyte and red algal chloroplasts in that they have lost their phycobilisomes, and contain chlorophyll b.<ref name="Kim-2009" /> They have also lost the peptidoglycan wall between their double membrane, leaving an intermembrane space.<ref name="Kim-2009" /> Some plants have kept some genes required the synthesis of peptidoglycan, but have repurposed them for use in chloroplast division instead.<ref name="Machida-2006">Template:Cite journal</ref> Chloroplastida lineages also keep their starch inside their chloroplasts.<ref name="Kim-2009" /><ref name="Keeling-2010" /><ref name="Lewis-2004" /> In plants and some algae, the chloroplast thylakoids are arranged in grana stacks. Some green algal chloroplasts, as well as those of hornworts, contain a structure called a pyrenoid,<ref name="Kim-2009" /> that concentrate RuBisCO and COTemplate:Sub in the chloroplast, functionally similar to the glaucophyte carboxysome.<ref name="Moroney-1999">Template:Cite journal</ref><ref>Template:Cite journal </ref>

There are some lineages of non-photosynthetic parasitic green algae that have lost their chloroplasts entirely, such as Prototheca,<ref name="Keeling-2010" /> or have no chloroplast while retaining the separate chloroplast genome, as in Helicosporidium.<ref name="Tartar-2004">Template:Cite journal</ref> Morphological and physiological similarities, as well as phylogenetics, confirm that these are lineages that ancestrally had chloroplasts but have since lost them.<ref name="Tartar-2004" /><ref>Template:Cite journal</ref>

Paulinella chromatophora

The photosynthetic amoeboids in the genus Paulinella—P. chromatophora, P. micropora, and marine P. longichromatophora—have the only known independently evolved chloroplast, often called a chromatophore.<ref name=":0" group="Note" /> While all other chloroplasts originate from a single ancient endosymbiotic event, Paulinella independently acquired an endosymbiotic cyanobacterium from the genus Synechococcus around 90 – 140 million years ago.<ref name="Macorano-2021" /><ref name="Keeling-2004" /> Each Paulinella cell contains one or two sausage-shaped chloroplasts;<ref name="Nakayama-2012" /><ref name="Nowack-2011">Template:Cite journal</ref> they were first described in 1894 by German biologist Robert Lauterborn.<ref>Template:Cite journal</ref>

The chromatophore is highly reduced compared to its free-living cyanobacterial relatives and has limited functions. For example, it has a genome of about 1 million base pairs, one third the size of Synechococcus genomes, and only encodes around 850 proteins.<ref name="Nakayama-2012" /> However, this is still much larger than other chloroplast genomes, which are typically around 150,000 base pairs. Chromatophores have also transferred much less of their DNA to the nucleus of their hosts. About 0.3–0.8% of the nuclear DNA in Paulinella is from the chromatophore, compared with 11–14% from the chloroplast in plants.<ref name="Nowack-2011" /> Similar to other chloroplasts, Paulinella provides specific proteins to the chromatophore using a specific targeting sequence.<ref>Template:Cite journal</ref> Because chromatophores are much younger compared to the canoncial chloroplasts, Paulinella chromatophora is studied to understand how early chloroplasts evolved.<ref name="Nakayama-2012" />

Secondary and tertiary chloroplast lineages

Green algal derived chloroplasts

Green algae have been taken up by many groups in three or four separate events.<ref name="Rogers-2007">Template:Cite journal</ref> Primarily, secondary chloroplasts derived from green algae are in the euglenids and chlorarachniophytes. They are also found in one lineage of dinoflagellates<ref name="Keeling-2010">Template:Cite journal</ref> and possibly the ancestor of the CASH lineage (cryptomonads, alveolates, stramenopiles and haptophytes)<ref name="Moustafa-2009">Template:Cite journal</ref> Many green algal derived chloroplasts contain pyrenoids, but unlike chloroplasts in their green algal ancestors, storage product collects in granules outside the chloroplast.<ref name="Kim-2009" />

Euglenophytes

File:Two Euglena.jpg

Euglena, a euglenophyte, contains secondary chloroplasts from green algae.

The euglenophytes are a group of common flagellated protists that contain chloroplasts derived from a green alga.<ref name="Keeling-2004" /> Euglenophytes are the only group outside Diaphoretickes that have chloroplasts without performing kleptoplasty.<ref>Template:Cite journal</ref><ref>Template:Cite journal</ref> Euglenophyte chloroplasts have three membranes. It is thought that the membrane of the primary endosymbiont host was lost (e.g. the green algal membrane), leaving the two cyanobacterial membranes and the secondary host's phagosomal membrane.<ref name="Keeling-2004" /> Euglenophyte chloroplasts have a pyrenoid and thylakoids stacked in groups of three. The carbon fixed through photosynthesis is stored in the form of paramylon, which is contained in membrane-bound granules in the cytoplasm of the euglenophyte.<ref name="Kim-2009" /><ref name="Keeling-2010" />

Chlorarachniophytes

File:Chlorarachnion reptans.jpg

Chlorarachnion reptans is a chlorarachniophyte. Chlorarachniophytes replaced their original red algal endosymbiont with a green alga.

Chlorarachniophytes are a rare group of organisms that also contain chloroplasts derived from green algae,<ref name="Keeling-2004" /> though their story is more complicated than that of the euglenophytes. The ancestor of chlorarachniophytes is thought to have been a eukaryote with a red algal derived chloroplast. It is then thought to have lost its first red algal chloroplast, and later engulfed a green alga, giving it its second, green algal derived chloroplast.<ref name="Keeling-2010" />

Chlorarachniophyte chloroplasts are bounded by four membranes, except near the cell membrane, where the chloroplast membranes fuse into a double membrane.<ref name="Kim-2009" /> Their thylakoids are arranged in loose stacks of three.<ref name="Kim-2009" /> Chlorarachniophytes have a form of polysaccharide called chrysolaminarin, which they store in the cytoplasm,<ref name="Keeling-2010" /> often collected around the chloroplast pyrenoid, which bulges into the cytoplasm.<ref name="Kim-2009" />

Chlorarachniophyte chloroplasts are notable because the green alga they are derived from has not been completely broken down—its nucleus still persists as a nucleomorph<ref name="Keeling-2004" /> found between the second and third chloroplast membranes<ref name="Kim-2009" />—the periplastid space, which corresponds to the green alga's cytoplasm.<ref name="Keeling-2010" />

Prasinophyte-derived chloroplast

File:Lepidodinium chlorophorum 68163.jpg

Lepidodinium chlorophorum's green colour is caused by a plastid derived from Pedinophyceae.

Dinoflagellates in the genus Lepidodinium have lost their original peridinin chloroplast and replaced it with a green algal derived chloroplast (more specifically, a prasinophyte).<ref name="Kim-2009" /><ref name="Hackett-2004" /> Lepidodinium is the only dinoflagellate that has a chloroplast that's not from the rhodoplast lineage. The chloroplast is surrounded by two membranes and has no nucleomorph—all the nucleomorph genes have been transferred to the dinophyte nucleus.<ref name="Hackett-2004" /> The endosymbiotic event that led to this chloroplast was serial secondary endosymbiosis rather than tertiary endosymbiosis—the endosymbiont was a green alga containing a primary chloroplast (making a secondary chloroplast).<ref name="Keeling-2010" />

Tripartite symbiosis

File:Abg4102.F1.large.jpg

Pseudoblepharisma tenue with its two photosynthetic symbionts.

The ciliate Pseudoblepharisma tenue has two bacterial symbionts, one pink, one green. In 2021, both symbionts were confirmed to be photosynthetic: Ca. Thiodictyon intracellulare (Chromatiaceae), a purple sulfur bacterium with a genome just half the size of their closest known relatives; and Chlorella sp. K10, a green alga.<ref>Template:Cite journal</ref> There is also a variant of Pseudoblepharisma tenue that only contains chloroplasts from green algae and no endosymbiotic purple bacteria.<ref name="Hines">Template:Cite journal</ref>

Red algal derived chloroplasts

Secondary chloroplasts derived from red algae appear to have only been taken up only once, which then diversified into a large group called chromists or chromalveolates. Today they are found in the haptophytes, cryptomonads, heterokonts, dinoflagellates and apicomplexans (the CASH lineage).<ref name="Keeling-2010" /> Red algal secondary chloroplasts usually contain chlorophyll c and are surrounded by four membranes.<ref name="Kim-2009" />

However, chromist monophyly has been rejected, and it is considered more likely that some chromists acquired their plastids by incorporating another chromist instead of inheriting them from a common ancestor. Cryptophytes seem to have acquired plastids from red algae, which were then transmitted from them to both the Heterokontophytes and the Haptophytes, and then from these last to the Myzozoa.<ref name="Strassert Irisarri Williams Burki 2021">Template:Cite journal</ref>

Cryptophytes

File:CSIRO ScienceImage 6743 SEM Cryptophyte.jpg

Cryptophytes under SEM.

Cryptophytes, or cryptomonads, are a group of algae that contain a red-algal derived chloroplast. Cryptophyte chloroplasts contain a nucleomorph that superficially resembles that of the chlorarachniophytes.<ref name="Keeling-2004" /> Cryptophyte chloroplasts have four membranes. The outermost membrane is continuous with the rough endoplasmic reticulum. They synthesize ordinary starch, which is stored in granules found in the periplastid space—outside the original double membrane, in the place that corresponds to the ancestral red alga's cytoplasm. Inside cryptophyte chloroplasts is a pyrenoid and thylakoids in stacks of two.<ref name="Kim-2009" /> Cryptophyte chloroplasts do not have phycobilisomes,<ref name="Kim-2009" /> but they do have phycobilin pigments which they keep in the thylakoid space, rather than anchored on the outside of their thylakoid membranes.<ref name="Kim-2009" /><ref name="Keeling-2004" />

Cryptophytes may have played a key role in the spreading of red algal based chloroplasts.<ref>Template:Cite thesis</ref><ref>Template:Cite journal</ref>

Haptophytes

File:Gephyrocapsa oceanica brighter.jpg

Scanning electron micrograph of Gephyrocapsa oceanica, a haptophyte.

Haptophytes are similar and closely related to cryptophytes or heterokontophytes.<ref name="Keeling-2010" /> Their chloroplasts lack a nucleomorph,<ref name="Kim-2009" /><ref name="Keeling-2004" /> their thylakoids are in stacks of three, and they synthesize chrysolaminarin sugar, which are stored in granules completely outside of the chloroplast, in the cytoplasm of the haptophyte.<ref name="Kim-2009" />

Stramenopiles (heterokontophytes)

File:20110123 185042 Diatom.jpg

The photosynthetic pigments present in their chloroplasts make diatoms greenish-brown.

The stramenopiles, also known as heterokontophytes, are a very large and diverse group of eukaryotes. It inlcludes Ochrophyta—which includes diatoms, brown algae (seaweeds), and golden algae (chrysophytes)<ref name="Campbell-2009f" />— and Xanthophyceae (also called yellow-green algae).<ref name="Keeling-2010" />

Heterokont chloroplasts are very similar to haptophyte chloroplasts. They have a pyrenoid, triplet thylakoids, and, with some exceptions,<ref name="Kim-2009" /> four layer plastidic envelope with the outermost membrane connected to the endoplasmic reticulum. Like haptophytes, stramenopiles store sugar in chrysolaminarin granules in the cytoplasm.<ref name="Kim-2009" /> Stramenopile chloroplasts contain chlorophyll a and, with a few exceptions,<ref name="Kim-2009" /> chlorophyll c.<ref name="Keeling-2004" /> They also have carotenoids which give them their many colors.<ref name="Campbell-2009f" />

Apicomplexans, chromerids, and dinophytes

Template:See also The alveolates are a major clade of unicellular eukaryotes of both autotrophic and heterotrophic members. Many members contain a red-algal derived plastid. One notable characteristic of this diverse group is the frequent loss of photosynthesis. However, a majority of these heterotrophs continue to process a non-photosynthetic plastid.<ref name="Janouškovec-2017">Template:Cite journal</ref>

Apicomplexans

File:Plasmodium.png

Diagram of Plasmodium, including its apicoplast.

Apicomplexans are a group of alveolates. Like the helicosproidia, they're parasitic, and have a nonphotosynthetic chloroplast.<ref name="Keeling-2010" /> They were once thought to be related to the helicosproidia, but it is now known that the helicosproida are green algae rather than part of the CASH lineage.<ref name="Keeling-2010" /> The apicomplexans include Plasmodium, the malaria parasite. Many apicomplexans keep a vestigial red algal derived chloroplast<ref name="Nair-2011" /><ref name="Keeling-2010" /> called an apicoplast, which they inherited from their ancestors. Apicoplasts have lost all photosynthetic function, and contain no photosynthetic pigments or true thylakoids. They are bounded by four membranes, but the membranes are not connected to the endoplasmic reticulum.<ref name="Kim-2009" /> Other apicomplexans like Cryptosporidium have lost the chloroplast completely.<ref name="Nair-2011" /> Apicomplexans store their energy in amylopectin granules that are located in their cytoplasm, even though they are nonphotosynthetic.<ref name="Kim-2009" />

The fact that apicomplexans still keep their nonphotosynthetic chloroplast around demonstrates how the chloroplast carries out important functions other than photosynthesis. Plant chloroplasts provide plant cells with many important things besides sugar, and apicoplasts are no different—they synthesize fatty acids, isopentenyl pyrophosphate, iron-sulfur clusters, and carry out part of the heme pathway.<ref name="Nair-2011" /> The most important apicoplast function is isopentenyl pyrophosphate synthesis—in fact, apicomplexans die when something interferes with this apicoplast function, and when apicomplexans are grown in an isopentenyl pyrophosphate-rich medium, they dump the organelle.<ref name="Nair-2011" />

Chromerids

File:Vitrella brassicaformis LM Michalek 2020.png

Typical life cycle stages of Vitrella brassicaformis, a chromerid.

The chromerids are a group of algae known from Australian corals which comprise some close photosynthetic relatives of the apicomplexans. The first member, Chromera velia, was discovered and first isolated in 2001. The discovery of Chromera velia with similar structure to the apicomplexans, provides an important link in the evolutionary history of the apicomplexans and dinophytes. Their plastids have four membranes, lack chlorophyll c and use the type II form of RuBisCO obtained from a horizontal transfer event.<ref>Template:Cite journal</ref>

Dinophytes

File:Ceratium furca.jpg

Ceratium furca, a peridinin-containing dinophyte.<ref>Template:Cite journal</ref>

The dinoflagellates are yet another very large and diverse group, around half of which are at least partially photosynthetic (i.e. mixotrophic).<ref name="Campbell-2009f" /><ref name="Hackett-2004">Template:Cite journal</ref> Dinoflagellate chloroplasts have relatively complex history. Most dinoflagellate chloroplasts are secondary red algal derived chloroplasts. Many dinoflagellates have lost the chloroplast (becoming nonphotosynthetic), some of these have replaced it though tertiary endosymbiosis.<ref name="Dorrell-2011">Template:Cite journal</ref> Others replaced their original chloroplast with a green algal derived chloroplast.<ref name="Keeling-2004" /><ref name="Keeling-2010" /><ref name="Hackett-2004" /> The peridinin chloroplast is thought to be the dinophytes' "original" chloroplast,<ref name="Hackett-2004" /> which has been lost, reduced, replaced, or has company in several other dinophyte lineages.<ref name="Keeling-2010" />

The most common dinophyte chloroplast is the peridinin-type chloroplast, characterized by the carotenoid pigment peridinin in their chloroplasts, along with chlorophyll a and chlorophyll c₂.<ref name="Keeling-2004" /><ref name="Hackett-2004" /> Peridinin is not found in any other group of chloroplasts.<ref name="Hackett-2004" /> The peridinin chloroplast is bounded by three membranes (occasionally two),<ref name="Kim-2009" /> having lost the red algal endosymbiont's original cell membrane.<ref name="Keeling-2004" /><ref name="Keeling-2010" /> The outermost membrane is not connected to the endoplasmic reticulum.<ref name="Kim-2009" /><ref name="Hackett-2004" /> They contain a pyrenoid, and have triplet-stacked thylakoids. Starch is found outside the chloroplast.<ref name="Kim-2009" /> Peridinin chloroplasts also have DNA that is highly reduced and fragmented into many small circles.<ref name="Hackett-2004" /> Most of the genome has migrated to the nucleus, and only critical photosynthesis-related genes remain in the chloroplast.

Most dinophyte chloroplasts contain form II RuBisCO, at least the photosynthetic pigments chlorophyll a, chlorophyll c₂, beta-carotene, and at least one dinophyte-unique xanthophyll (peridinin, dinoxanthin, or diadinoxanthin), giving many a golden-brown color.<ref name="Janouškovec-2017" /><ref name="Hackett-2004" /> All dinophytes store starch in their cytoplasm, and most have chloroplasts with thylakoids arranged in stacks of three.<ref name="Kim-2009" />

Haptophyte-derived chloroplasts

File:Karenia brevis.jpg

Karenia brevis is a fucoxanthin-containing dynophyte responsible for algal blooms called "red tides".<ref name="Hackett-2004" />

The fucoxanthin dinophyte lineages (including Karlodinium and Karenia)<ref name="Keeling-2010" /> lost their original red algal derived chloroplast, and replaced it with a new chloroplast derived from a haptophyte endosymbiont, making these tertiary plastids. Karlodinium and Karenia probably took up different endosymbionts.<ref name="Keeling-2010" /> Because the haptophyte chloroplast has four membranes, tertiary endosymbiosis would be expected to create a six membraned chloroplast, adding the haptophyte's cell membrane and the dinophyte's phagosomal vacuole.<ref name="Tengs-2000" /> However, the haptophyte was heavily reduced, stripped of a few membranes and its nucleus, leaving only its chloroplast (with its original double membrane), and possibly one or two additional membranes around it.<ref name="Keeling-2010" /><ref name="Tengs-2000">Template:Cite journal</ref>

Fucoxanthin-containing chloroplasts are characterized by having the pigment fucoxanthin (actually 19′-hexanoyloxy-fucoxanthin and/or 19′-butanoyloxy-fucoxanthin) and no peridinin. Fucoxanthin is also found in haptophyte chloroplasts, providing evidence of ancestry.<ref name="Hackett-2004" />

Diatom-derived chloroplasts

File:Durinskia baltica.jpg

Durinskia is a genus significant to the study of endosymbiotic events and organelle integration.<ref name="Zerdoner2017">Template:Cite journal</ref>

Some dinophytes, like Kryptoperidinium and Durinskia,<ref name="Keeling-2010" /> have a diatom (heterokontophyte)-derived chloroplast.<ref name="Keeling-2004" /> These chloroplasts are bounded by up to five membranes,<ref name="Keeling-2004" /> (depending on whether the entire diatom endosymbiont is counted as the chloroplast, or just the red algal derived chloroplast inside it). The diatom endosymbiont has been reduced relatively little—it still retains its original mitochondria,<ref name="Keeling-2010" /> and has endoplasmic reticulum, ribosomes, a nucleus, and of course, red algal derived chloroplasts—practically a complete cell,<ref name="Schnepf-1999">Template:Cite journal</ref> all inside the host's endoplasmic reticulum lumen.<ref name="Keeling-2010" /> However the diatom endosymbiont can't store its own food—its storage polysaccharide is found in granules in the dinophyte host's cytoplasm instead.<ref name="Kim-2009" /><ref name="Schnepf-1999" /> The diatom endosymbiont's nucleus is present, but it probably can't be called a nucleomorph because it shows no sign of genome reduction, and might have even been expanded.<ref name="Keeling-2010" /> Diatoms have been engulfed by dinoflagellates at least three times.<ref name="Keeling-2010" />

The diatom endosymbiont is bounded by a single membrane,<ref name="Hackett-2004" /> inside it are chloroplasts with four membranes. Like the diatom endosymbiont's diatom ancestor, the chloroplasts have triplet thylakoids and pyrenoids.<ref name="Schnepf-1999" />

In some of these genera, the diatom endosymbiont's chloroplasts aren't the only chloroplasts in the dinophyte. The original three-membraned peridinin chloroplast is still around, converted to an eyespot.<ref name="Keeling-2004" /><ref name="Keeling-2010" />

Kleptoplasty

File:Dinophysis acuminata.jpg

In some groups of mixotrophic protists, like some dinoflagellates (e.g. Dinophysis), chloroplasts are separated from a captured alga and used temporarily. These klepto chloroplasts may only have a lifetime of a few days and are then replaced.<ref name="Skovgaard-1998">Template:Cite journal</ref><ref>Template:Cite journal</ref>

Cryptophyte-derived dinophyte chloroplast

Dinophysis acuminata has chloroplasts taken from a cryptophyte.<ref name="Keeling-2004" />

Members of the genus Dinophysis have a phycobilin-containing<ref name="Tengs-2000" /> chloroplast taken from a cryptophyte.<ref name="Keeling-2004" /> However, the cryptophyte is not an endosymbiont—only the chloroplast seems to have been taken, and the chloroplast has been stripped of its nucleomorph and outermost two membranes, leaving just a two-membraned chloroplast. Cryptophyte chloroplasts require their nucleomorph to maintain themselves, and Dinophysis species grown in cell culture alone cannot survive, so it is possible (but not confirmed) that the Dinophysis chloroplast is a kleptoplast—if so, Dinophysis chloroplasts wear out and Dinophysis species must continually engulf cryptophytes to obtain new chloroplasts to replace the old ones.<ref name="Hackett-2004" />

Chloroplast DNA

Template:Main Template:See also

Chloroplasts, like other endosymbiotic organelles, contain a genome separate from that in the cell nucleus. The existence of chloroplast DNA (cpDNA) was identified biochemically in 1959,<ref name="Stocking-1959" /> and confirmed by electron microscopy in 1962.<ref name="Plaut-1962" /> The discoveries that the chloroplast contains ribosomes<ref name="Lyttleton-1962" /> and performs protein synthesis<ref name="Heber, U.-1962" /> revealed that the chloroplast is genetically semi-autonomous. Chloroplast DNA was first sequenced in 1986.<ref name="UniHamburg">Template:Cite web</ref> Since then, hundreds of chloroplast genomes from various species have been sequenced, but they are mostly those of land plants and green algae—glaucophytes, red algae, and other algal groups are extremely underrepresented, potentially introducing some bias in views of "typical" chloroplast DNA structure and content.<ref name="Sandelius-2009">Template:Cite book</ref>

Template:Chloroplast DNA

Molecular structure

With few exceptions, most chloroplasts have their entire chloroplast genome combined into a single large circular DNA molecule,<ref name="Sandelius-2009" /> typically 120,000–170,000 base pairs long<ref name="Dann-2002">Template:Cite book</ref><ref name="Clegg-1994" /><ref name="Shaw-2007" /><ref name="Milo">Template:Cite web</ref> and a mass of about 80–130 million daltons.<ref name="Burgess-1989b">Template:Cite book</ref> While chloroplast genomes can almost always be assembled into a circular map, the physical DNA molecules inside cells take on a variety of linear and branching forms.<ref name="Sandelius-2009" /><ref>Template:Cite journal</ref> New chloroplasts may contain up to 100 copies of their genome,<ref name="Dann-2002" /> though the number of copies decreases to about 15–20 as the chloroplasts age.<ref name="PlantBiochem-2005">Template:Cite book</ref>

Chloroplast DNA is usually condensed into nucleoids, which can contain multiple copies of the chloroplast genome. Many nucleoids can be found in each chloroplast.<ref name="Burgess-1989b" /> In primitive red algae, the chloroplast DNA nucleoids are clustered in the center of the chloroplast, while in green plants and green algae, the nucleoids are dispersed throughout the stroma.<ref name="Kobayashi-2002" /> Chloroplast DNA is not associated with true histones, proteins that are used to pack DNA molecules tightly in eukaryote nuclei.<ref name="Campbell-2009c" /> Though in red algae, similar proteins tightly pack each chloroplast DNA ring in a nucleoid.<ref name="Kobayashi-2002">Template:Cite journal</ref>

Many chloroplast genomes contain two inverted repeats, which separate a long single copy section (LSC) from a short single copy section (SSC).<ref name="Shaw-2007">Template:Cite journal</ref> A given pair of inverted repeats are rarely identical, but they are always very similar to each other, apparently resulting from concerted evolution.<ref name="Sandelius-2009" /> The inverted repeats vary wildly in length, ranging from 4,000 to 25,000 base pairs long each and containing as few as four or as many as over 150 genes.<ref name="Sandelius-2009" /> The inverted repeat regions are highly conserved in land plants, and accumulate few mutations.<ref name="Shaw-2007" /><ref name="Kolodner-1979">Template:Cite journal</ref>

Similar inverted repeats exist in the genomes of cyanobacteria and the other two chloroplast lineages (glaucophyta and rhodophyceae), suggesting that they predate the chloroplast.<ref name="Sandelius-2009" /> Some chloroplast genomes have since lost<ref name="Kolodner-1979" /><ref name="Palmer-1982" /> or flipped the inverted repeats (making them direct repeats).<ref name="Sandelius-2009" /> It is possible that the inverted repeats help stabilize the rest of the chloroplast genome, as chloroplast genomes which have lost some of the inverted repeat segments tend to get rearranged more.<ref name="Palmer-1982">Template:Cite journal</ref>

DNA repair and replication

In chloroplasts of the moss Physcomitrella patens, the DNA mismatch repair protein Msh1 interacts with the recombinational repair proteins RecA and RecG to maintain chloroplast genome stability.<ref name="Odahara-2017">Template:Cite journal</ref> In chloroplasts of the plant Arabidopsis thaliana the RecA protein maintains the integrity of the chloroplast's DNA by a process that likely involves the recombinational repair of DNA damage.<ref name="Rowan-2010">Template:Cite journal</ref>

File:CpDNA Replication.png

Chloroplast DNA replication via multiple D-loop mechanisms. Adapted from Krishnan NM, Rao BJ's paper "A comparative approach to elucidate chloroplast genome replication."

The mechanism for chloroplast DNA (cpDNA) replication has not been conclusively determined, but two main models have been proposed. Scientists have attempted to observe chloroplast replication via electron microscopy since the 1970s.<ref name="Krishnan-2009">Template:Cite journal</ref><ref name="Heinhorst-1993">Template:Cite journal</ref> The results of the microscopy experiments led to the idea that chloroplast DNA replicates using a double displacement loop (D-loop). As the D-loop moves through the circular DNA, it adopts a theta intermediary form, also known as a Cairns replication intermediate, and completes replication with a rolling circle mechanism.<ref name="Krishnan-2009"/><ref name="Bendich-2004">Template:Cite journal</ref> Transcription starts at specific points of origin. Multiple replication forks open up, allowing replication machinery to transcribe the DNA. As replication continues, the forks grow and eventually converge. The new cpDNA structures separate, creating daughter cpDNA chromosomes.

In addition to the early microscopy experiments, this model is also supported by the amounts of deamination seen in cpDNA.<ref name="Krishnan-2009"/> Deamination occurs when an amino group is lost and is a mutation that often results in base changes. When adenine is deaminated, it becomes hypoxanthine. Hypoxanthine can bind to cytosine, and when the XC base pair is replicated, it becomes a GC (thus, an A → G base change).<ref name=Biocyclopedia>Template:Cite web</ref>

File:Adenine Deaminates to Guanine.png

Over time, base changes in the DNA sequence can arise from deamination mutations. When adenine is deaminated, it becomes hypoxanthine, which can pair with cytosine. During replication, the cytosine will pair with guanine, causing an A --> G base change.

In cpDNA, there are several A → G deamination gradients. DNA becomes susceptible to deamination events when it is single stranded. When replication forks form, the strand not being copied is single stranded, and thus at risk for A → G deamination. Therefore, gradients in deamination indicate that replication forks were most likely present and the direction that they initially opened (the highest gradient is most likely nearest the start site because it was single stranded for the longest amount of time).<ref name="Krishnan-2009"/> This mechanism is still the leading theory today; however, a second theory suggests that most cpDNA is actually linear and replicates through homologous recombination. It further contends that only a minority of the genetic material is kept in circular chromosomes while the rest is in branched, linear, or other complex structures.<ref name="Krishnan-2009"/><ref name="Bendich-2004"/>

One of competing model for cpDNA replication asserts that most cpDNA is linear and participates in homologous recombination and replication structures similar to the linear and circular DNA structures of bacteriophage T4.<ref name="Bendich-2004"/><ref name="Bernstein-1973">Template:Cite journal</ref> It has been established that some plants have linear cpDNA, such as maize, and that more species still contain complex structures that scientists do not yet understand.<ref name="Bendich-2004"/> When the original experiments on cpDNA were performed, scientists did notice linear structures; however, they attributed these linear forms to broken circles.<ref name="Bendich-2004"/> If the branched and complex structures seen in cpDNA experiments are real and not artifacts of concatenated circular DNA or broken circles, then a D-loop mechanism of replication is insufficient to explain how those structures would replicate.<ref name="Bendich-2004"/> At the same time, homologous recombination does not expand the multiple A --> G gradients seen in plastomes.<ref name="Krishnan-2009"/> Because of the failure to explain the deamination gradient as well as the numerous plant species that have been shown to have circular cpDNA, the predominant theory continues to hold that most cpDNA is circular and most likely replicates via a D loop mechanism.

Gene content and protein synthesis

The ancestral cyanobacteria that led to chloroplasts probably had a genome that contained over 3000 genes, but only approximately 100 genes remain in contemporary chloroplast genomes.<ref name="Milo" /><ref name="McFadden-2001">Template:Cite journal</ref><ref name="Clegg-1994">Template:Cite journal</ref> These genes code for a variety of things, mostly to do with the protein pipeline and photosynthesis. As in prokaryotes, genes in chloroplast DNA are organized into operons.<ref name="McFadden-2001" /> Unlike prokaryotic DNA molecules, chloroplast DNA molecules contain introns (plant mitochondrial DNAs do too, but not human mtDNAs).<ref name="Alberts-2002b" />

Among land plants, the contents of the chloroplast genome are fairly similar.<ref name="Shaw-2007" />

Chloroplast genome reduction and gene transfer

Over time, many parts of the chloroplast genome were transferred to the nuclear genome of the host,<ref name="Dann-2002" /><ref name="Clegg-1994" /><ref>Template:Cite journal</ref> a process called endosymbiotic gene transfer. As a result, the chloroplast genome is heavily reduced compared to that of free-living cyanobacteria. Chloroplasts may contain 60–100 genes whereas cyanobacteria often have more than 1500 genes in their genome.<ref name="Martin-2002">Template:Cite journal</ref> Recently, a plastid without a genome was found, demonstrating chloroplasts can lose their genome during endosymbiotic the gene transfer process.<ref>Template:Cite journal</ref>

Endosymbiotic gene transfer is how we know about the lost chloroplasts in many CASH lineages. Even if a chloroplast is eventually lost, the genes it donated to the former host's nucleus persist, providing evidence for the lost chloroplast's existence. For example, while diatoms (a heterokontophyte) now have a red algal derived chloroplast, the presence of many green algal genes in the diatom nucleus provide evidence that the diatom ancestor had a green algal derived chloroplast at some point, which was subsequently replaced by the red chloroplast.<ref name="Moustafa-2009" />

In land plants, some 11–14% of the DNA in their nuclei can be traced back to the chloroplast,<ref name="Nowack-2011" /> up to 18% in Arabidopsis, corresponding to about 4,500 protein-coding genes.<ref name="Archibald-2006">Template:Cite journal</ref> There have been a few recent transfers of genes from the chloroplast DNA to the nuclear genome in land plants.<ref name="Clegg-1994" />

Of the approximately 3000 proteins found in chloroplasts, some 95% of them are encoded by nuclear genes. Many of the chloroplast's protein complexes consist of subunits from both the chloroplast genome and the host's nuclear genome. As a result, protein synthesis must be coordinated between the chloroplast and the nucleus. The chloroplast is mostly under nuclear control, though chloroplasts can also give out signals regulating gene expression in the nucleus, called retrograde signaling.<ref name="Koussevitzky-2007">Template:Cite journal

Template:Cite magazine</ref> Recent research indicates that parts of the retrograde signaling network once considered characteristic for land plants emerged already in an algal progenitor,<ref>Template:Cite journal</ref><ref>Template:Cite journal</ref><ref>Template:Cite journal</ref> integrating into co-expressed cohorts of genes in the closest algal relatives of land plants.<ref>Template:Cite journal</ref>

Protein synthesis

Template:See also Protein synthesis within chloroplasts relies on two RNA polymerases. One is coded by the chloroplast DNA, the other is of nuclear origin. The two RNA polymerases may recognize and bind to different kinds of promoters within the chloroplast genome.<ref name="Hedtke-1997">Template:Cite journal</ref> The ribosomes in chloroplasts are similar to bacterial ribosomes.<ref name="Harris-1994">Template:Cite journal</ref>

Protein targeting and import

Because so many chloroplast genes have been moved to the nucleus, many proteins that would originally have been translated in the chloroplast are now synthesized in the cytoplasm of the plant cell. These proteins must be directed back to the chloroplast, and imported through at least two chloroplast membranes.<ref name="Soll-2004">Template:Cite journal</ref>

Curiously, around half of the protein products of transferred genes aren't even targeted back to the chloroplast. Many became exaptations, taking on new functions like participating in cell division, protein routing, and even disease resistance. A few chloroplast genes found new homes in the mitochondrial genome—most became nonfunctional pseudogenes, though a few tRNA genes still work in the mitochondrion.<ref name="Martin-2002" /> Some transferred chloroplast DNA protein products get directed to the secretory pathway,<ref name="Martin-2002" /> though many secondary plastids are bounded by an outermost membrane derived from the host's cell membrane, and therefore topologically outside of the cell because to reach the chloroplast from the cytosol, the cell membrane must be crossed, which signifies entrance into the extracellular space. In those cases, chloroplast-targeted proteins do initially travel along the secretory pathway.<ref name="Keeling-2010" />

Because the cell acquiring a chloroplast already had mitochondria (and peroxisomes, and a cell membrane for secretion), the new chloroplast host had to develop a unique protein targeting system to avoid having chloroplast proteins being sent to the wrong organelle.<ref name="Soll-2004" />

File:Chloroplast in leaf of Anemone sp TEM 12000x.png

In most, but not all cases, nuclear-encoded chloroplast proteins are translated with a cleavable transit peptide that's added to the N-terminus of the protein precursor. Sometimes the transit sequence is found on the C-terminus of the protein,<ref name="Lung-2012">Template:Cite journal</ref> or within the functional part of the protein.<ref name="Soll-2004" />

Transport proteins and membrane translocons

After a chloroplast polypeptide is synthesized on a ribosome in the cytosol, an enzyme specific to chloroplast proteins<ref name="Waegemann-1996">Template:Cite journal</ref> phosphorylates, or adds a phosphate group to many (but not all) of them in their transit sequences.<ref name="Soll-2004" /> Phosphorylation helps many proteins bind the polypeptide, keeping it from folding prematurely.<ref name="Soll-2004" /> This is important because it prevents chloroplast proteins from assuming their active form and carrying out their chloroplast functions in the wrong place—the cytosol.<ref name="May-2000" /><ref name="Jarvis-2001">Template:Cite journal</ref> At the same time, they have to keep just enough shape so that they can be recognized by the chloroplast.<ref name="May-2000">Template:Cite journal</ref> These proteins also help the polypeptide get imported into the chloroplast.<ref name="Soll-2004" />

From here, chloroplast proteins bound for the stroma must pass through two protein complexes—the TOC complex, or translocon on the outer chloroplast membrane, and the TIC translocon, or translocon on the inner chloroplast membrane translocon.<ref name="Soll-2004" /> Chloroplast polypeptide chains probably often travel through the two complexes at the same time, but the TIC complex can also retrieve preproteins lost in the intermembrane space.<ref name="Soll-2004" />

Structure

Transmission electron microscope image of a chloroplast. Grana of thylakoids and their connecting lamellae are clearly visible.

In land plants, chloroplasts are generally lens-shaped, 3–10 μm in diameter and 1–3 μm thick.<ref name="Wise-2007b">Template:Cite book</ref><ref name="Milo"/> Corn seedling chloroplasts are ≈20 μm³ in volume.<ref name="Milo"/> Greater diversity in chloroplast shapes exists among the algae, which often contain a single chloroplast<ref name="Kim-2009" /> that can be shaped like a net (e.g., Oedogonium),<ref>Template:Cite web</ref> a cup (e.g., Chlamydomonas),<ref>Template:Cite web</ref> a ribbon-like spiral around the edges of the cell (e.g., Spirogyra),<ref>Template:Cite web</ref> or slightly twisted bands at the cell edges (e.g., Sirogonium).<ref>Template:Cite web</ref> Some algae have two chloroplasts in each cell; they are star-shaped in Zygnema,<ref>Template:Cite web</ref> or may follow the shape of half the cell in order Desmidiales.<ref>Template:Cite web</ref> In some algae, the chloroplast takes up most of the cell, with pockets for the nucleus and other organelles,<ref name="Kim-2009">Template:Cite book</ref> for example, some species of Chlorella have a cup-shaped chloroplast that occupies much of the cell.<ref>Template:Cite book</ref>

All chloroplasts have at least three membrane systems—the outer chloroplast membrane, the inner chloroplast membrane, and the thylakoid system. The two innermost lipid-bilayer membranes<ref name="Fuks-1996">Template:Cite journal</ref> that surround all chloroplasts correspond to the outer and inner membranes of the ancestral cyanobacterium's gram negative cell wall,<ref name="Keeling-2004" /><ref>Template:Cite journal</ref><ref>Template:Cite encyclopedia</ref> and not the phagosomal membrane from the host, which was probably lost.<ref name="Keeling-2004" /> Chloroplasts that are the product of secondary endosymbiosis may have additional membranes surrounding these three.<ref name="Chaal-2005" /> Inside the outer and inner chloroplast membranes is the chloroplast stroma, a semi-gel-like fluid<ref name="Wise-2006b">Template:Cite book</ref> that makes up much of a chloroplast's volume, and in which the thylakoid system floats.

There are some common misconceptions about the outer and inner chloroplast membranes. The fact that chloroplasts are surrounded by a double membrane is often cited as evidence that they are the descendants of endosymbiotic cyanobacteria. This is often interpreted as meaning the outer chloroplast membrane is the product of the host's cell membrane infolding to form a vesicle to surround the ancestral cyanobacterium—which is not true—both chloroplast membranes are homologous to the cyanobacterium's original double membranes.<ref name="Keeling-2004" />

The chloroplast double membrane is also often compared to the mitochondrial double membrane. This is not a valid comparison—the inner mitochondria membrane is used to run proton pumps and carry out oxidative phosphorylation across to generate ATP energy. The only chloroplast structure that can be considered analogous to it is the internal thylakoid system. Even so, in terms of "in-out", the direction of chloroplast [[hydronium|HTemplate:Sup]] ion flow is in the opposite direction compared to oxidative phosphorylation in mitochondria.<ref name="Wise-2006b" /><ref name="Campbell-2009b">Template:Cite book</ref> In addition, in terms of function, the inner chloroplast membrane, which regulates metabolite passage and synthesizes some materials, has no counterpart in the mitochondrion.<ref name="Wise-2006b" /> Template:Clear

Outer chloroplast membrane

The outer chloroplast membrane is a semi-porous membrane that small molecules and ions can easily diffuse across.<ref>Template:Cite journal</ref> However, it is not permeable to larger proteins, so chloroplast polypeptides being synthesized in the cell cytoplasm must be transported across the outer chloroplast membrane by the TOC complex, or translocon on the outer chloroplast membrane.<ref name="Soll-2004" />

The chloroplast membranes sometimes protrude out into the cytoplasm, forming a stromule, or stroma-containing tubule. Stromules are very rare in chloroplasts, and are much more common in other plastids like chromoplasts and amyloplasts in petals and roots, respectively.<ref name="Köhler-2000">Template:Cite journal</ref><ref name="Gray-2001">Template:Cite journal</ref> They may exist to increase the chloroplast's surface area for cross-membrane transport, because they are often branched and tangled with the endoplasmic reticulum.<ref name="Schattat-2011">Template:Cite journal</ref> When they were first observed in 1962, some plant biologists dismissed the structures as artifactual, claiming that stromules were just oddly shaped chloroplasts with constricted regions or dividing chloroplasts.<ref name="Schattat-2012">Template:Cite journal</ref> However, there is a growing body of evidence that stromules are functional, integral features of plant cell plastids, not merely artifacts.<ref>Template:Cite journal</ref>

Intermembrane space and peptidoglycan wall

File:Glaucocystis sp.jpg

Instead of an intermembrane space, glaucophyte algae have a peptidoglycan wall between their inner and outer chloroplast membranes.

Usually, a thin intermembrane space about 10–20 nanometers thick exists between the outer and inner chloroplast membranes.<ref name="Burgess-1989a">Template:Cite book</ref>

Glaucophyte algal chloroplasts have a peptidoglycan layer between the chloroplast membranes. It corresponds to the peptidoglycan cell wall of their cyanobacterial ancestors, which is located between their two cell membranes. These chloroplasts are called muroplasts (from Latin "mura", meaning "wall"). Other chloroplasts were assumed to have lost the cyanobacterial wall, leaving an intermembrane space between the two chloroplast envelope membranes,<ref name="Wise-2006b" /> but has since been found also in moss, lycophytes and ferns.<ref>Plant peptidoglycan precursor biosynthesis: Conservation between moss chloroplasts and Gram-negative bacteria</ref>

Inner chloroplast membrane

The inner chloroplast membrane borders the stroma and regulates passage of materials in and out of the chloroplast. After passing through the TOC complex in the outer chloroplast membrane, polypeptides must pass through the TIC complex (translocon on the inner chloroplast membrane) which is located in the inner chloroplast membrane.<ref name="Soll-2004" />

In addition to regulating the passage of materials, the inner chloroplast membrane is where fatty acids, lipids, and carotenoids are synthesized.<ref name="Wise-2006b" />

Peripheral reticulum

Some chloroplasts contain a structure called the chloroplast peripheral reticulum.<ref name="Burgess-1989a" /> It is often found in the chloroplasts of [[C4 plant|Template:C4 plants]], though it has also been found in some Template:C3 angiosperms,<ref name="Wise-2006b" /> and even some gymnosperms.<ref name="Whatley-1994">Template:Cite journal</ref> The chloroplast peripheral reticulum consists of a maze of membranous tubes and vesicles continuous with the inner chloroplast membrane that extends into the internal stromal fluid of the chloroplast. Its purpose is thought to be to increase the chloroplast's surface area for cross-membrane transport between its stroma and the cell cytoplasm. The small vesicles sometimes observed may serve as transport vesicles to shuttle stuff between the thylakoids and intermembrane space.<ref name="Wise-2007a" />

Stroma

The protein-rich,<ref name="Wise-2006b" /> alkaline,<ref name="Campbell-2009b" /> aqueous fluid within the inner chloroplast membrane and outside of the thylakoid space is called the stroma,<ref name="Wise-2006b" /> which corresponds to the cytosol of the original cyanobacterium. Nucleoids of chloroplast DNA, chloroplast ribosomes, the thylakoid system with plastoglobuli, starch granules, and many proteins can be found floating around in it. The Calvin cycle, which fixes [[CO2|COTemplate:Sub]] into G3P takes place in the stroma.

Chloroplast ribosomes

Template:Plain image with caption Chloroplasts have their own ribosomes, which they use to synthesize a small fraction of their proteins. Chloroplast ribosomes are about two-thirds the size of cytoplasmic ribosomes (around 17 nm vs 25 nm).<ref name="Burgess-1989a" /> They take mRNAs transcribed from the chloroplast DNA and translate them into protein. While similar to bacterial ribosomes,<ref name="Campbell-2009c" /> chloroplast translation is more complex than in bacteria, so chloroplast ribosomes include some chloroplast-unique features.<ref name="Manuell-2007">Template:Cite journal</ref><ref name="Bieri-2017">Template:Cite journal</ref>

Small subunit ribosomal RNAs in several Chlorophyta and euglenid chloroplasts lack motifs for Shine-Dalgarno sequence recognition,<ref name="Lim-2014">Template:Cite journal</ref> which is considered essential for translation initiation in most chloroplasts and prokaryotes.<ref>Template:Cite journal</ref><ref>Template:Cite journal</ref> Such loss is also rarely observed in other plastids and prokaryotes.<ref name="Lim-2014"/><ref>Template:Cite journal</ref> An additional 4.5S rRNA with homology to the 3' tail of 23S is found in "higher" plants.<ref name="Bieri-2017"/>

Plastoglobuli

Plastoglobuli (singular plastoglobulus, sometimes spelled plastoglobule(s)), are spherical bubbles of lipids and proteins<ref name="Wise-2006b" /> about 45–60 nanometers across.<ref name="Austin-2006" /> They are surrounded by a lipid monolayer.<ref name="Austin-2006" /> Plastoglobuli are found in all chloroplasts,<ref name="Burgess-1989a" /> but become more common when the chloroplast is under oxidative stress,<ref name="Austin-2006">Template:Cite journal</ref> or when it ages and transitions into a gerontoplast.<ref name="Wise-2006b" /> Plastoglobuli also exhibit a greater size variation under these conditions.<ref name="Austin-2006" /> They are also common in etioplasts, but decrease in number as the etioplasts mature into chloroplasts.<ref name="Austin-2006" />

Plastoglobuli contain both structural proteins and enzymes involved in lipid synthesis and metabolism. They contain many types of lipids including plastoquinone, vitamin E, carotenoids and chlorophylls.<ref name="Austin-2006" />

Plastoglobuli were once thought to be free-floating in the stroma, but it is now thought that they are permanently attached either to a thylakoid or to another plastoglobulus attached to a thylakoid, a configuration that allows a plastoglobulus to exchange its contents with the thylakoid network.<ref name="Austin-2006" /> In normal green chloroplasts, the vast majority of plastoglobuli occur singularly, attached directly to their parent thylakoid. In old or stressed chloroplasts, plastoglobuli tend to occur in linked groups or chains, still always anchored to a thylakoid.<ref name="Austin-2006" />

Plastoglobuli form when a bubble appears between the layers of the lipid bilayer of the thylakoid membrane, or bud from existing plastoglobuli—though they never detach and float off into the stroma.<ref name="Austin-2006" /> Practically all plastoglobuli form on or near the highly curved edges of the thylakoid disks or sheets. They are also more common on stromal thylakoids than on granal ones.<ref name="Austin-2006" />

Template:Main Template:Plain image with caption

Starch granules

Starch granules are very common in chloroplasts, typically taking up 15% of the organelle's volume,<ref name="Crumpton-Taylor-2012" /> though in some other plastids like amyloplasts, they can be big enough to distort the shape of the organelle.<ref name="Burgess-1989a" /> Starch granules are simply accumulations of starch in the stroma, and are not bounded by a membrane.<ref name="Burgess-1989a" />

Starch granules appear and grow throughout the day, as the chloroplast synthesizes sugars, and are consumed at night to fuel respiration and continue sugar export into the phloem,<ref name="Zeeman-2007">Template:Cite journal</ref> though in mature chloroplasts, it is rare for a starch granule to be completely consumed or for a new granule to accumulate.<ref name="Crumpton-Taylor-2012">Template:Cite journal</ref>

Starch granules vary in composition and location across different chloroplast lineages. In red algae, starch granules are found in the cytoplasm rather than in the chloroplast.<ref name="Rochaix-1998">Template:Cite book</ref> In [[C4 plant|Template:C4 plants]], mesophyll chloroplasts, which do not synthesize sugars, lack starch granules.<ref name="Wise-2006b" />

RuBisCO

The chloroplast stroma contains many proteins, though the most common and important is RuBisCO, which is probably also the most abundant protein on the planet.<ref name="Campbell-2009b" /> RuBisCO is the enzyme that fixes [[CO2|COTemplate:Sub]] into sugar molecules. In [[C3 plant|Template:C3 plants]], RuBisCO is abundant in all chloroplasts, though in [[C4 plant|Template:C4 plants]], it is confined to the bundle sheath chloroplasts, where the Calvin cycle is carried out in Template:C4 plants.<ref name="Gunning-1996a" />

Pyrenoids

The chloroplasts of some hornworts<ref>Template:Cite journal</ref> and algae contain structures called pyrenoids. They are not found in higher plants.<ref name="Ma-2011">Template:Cite journal</ref> Pyrenoids are roughly spherical and highly refractive bodies which are a site of starch accumulation in plants that contain them. They consist of a matrix opaque to electrons, surrounded by two hemispherical starch plates. The starch is accumulated as the pyrenoids mature.<ref name="Retallack-1970" /> In algae with carbon concentrating mechanisms, the enzyme RuBisCO is found in the pyrenoids. Starch can also accumulate around the pyrenoids when CO₂ is scarce.<ref name="Ma-2011" /> Pyrenoids can divide to form new pyrenoids, or be produced "de novo".<ref name="Retallack-1970">Template:Cite journal</ref><ref name="Brown-1970">Template:Cite journal</ref>

Thylakoid system

File:Lettuce Chloroplast STEM.jpg

Scanning transmission electron microscope imaging of a chloroplast
(Top) 10-nm-thick STEM tomographic slice of a lettuce chloroplast. Grana stacks are interconnected by unstacked stromal thylakoids, called "stroma lamellae". Round inclusions associated with the thylakoids are plastoglobules. Scalebar=200 nm. See.<ref name="Bussi-2019" />
(Bottom) Large-scale 3D model generated from segmentation of tomographic reconstructions by STEM. grana=yellow; stroma lamellae=green; plastoglobules=purple; chloroplast envelope=blue. See.<ref name="Bussi-2019" />

Thylakoids (sometimes spelled thylakoïds),<ref>Template:Cite journal</ref> are small interconnected sacks which contain the membranes that the light reactions of photosynthesis take place on. The word thylakoid comes from the Greek word thylakos which means "sack".<ref>Template:Cite web</ref>

Suspended within the chloroplast stroma is the thylakoid system, a highly dynamic collection of membranous sacks called thylakoids where chlorophyll is found and the light reactions of photosynthesis happen.<ref name="Campbell-2009g" /> In most vascular plant chloroplasts, the thylakoids are arranged in stacks called grana,<ref name="Mustárdy-2008" /> though in certain [[C4 plant|Template:C4 plant]] chloroplasts<ref name="Gunning-1996a" /> and some algal chloroplasts, the thylakoids are free floating.<ref name="Kim-2009" />

Thylakoid structure

File:Thylakoid Structure.jpg

Granum-stroma assembly structure The prevailing model of the granum-stroma assembly is stacks of granal thylakoids wrapped by right-handed helical stromal thylakoids which are connected to large parallel sheets of stromal thylakoids and adjacent right-handed helices by left-handed helical structures. (Based on<ref name="Bussi-2019" />).

Using a light microscope, it is just barely possible to see tiny green granules—which were named grana.<ref name="Burgess-1989a" /> With electron microscopy, it became possible to see the thylakoid system in more detail, revealing it to consist of stacks of flat thylakoids which made up the grana, and long interconnecting stromal thylakoids which linked different grana.<ref name="Burgess-1989a" /> In the transmission electron microscope, thylakoid membranes appear as alternating light-and-dark bands, 8.5 nanometers thick.<ref name="Burgess-1989a" />

The three-dimensional structure of the thylakoid membrane system has been disputed. Many models have been proposed, the most prevalent being the helical model, in which granum stacks of thylakoids are wrapped by helical stromal thylakoids.<ref name="Paolillo-1970">Template:Cite journal</ref> Another model known as the 'bifurcation model', which was based on the first electron tomography study of plant thylakoid membranes, depicts the stromal membranes as wide lamellar sheets perpendicular to the grana columns which bifurcates into multiple parallel discs forming the granum-stroma assembly.<ref name="Reich Z-2005">Template:Cite journal</ref> The helical model was supported by several additional works,<ref name="Mustárdy-2008">Template:Cite journal</ref><ref name="Austin-2011">Template:Cite journal</ref> but ultimately it was determined in 2019 that features from both the helical and bifurcation models are consolidated by newly discovered left-handed helical membrane junctions.<ref name="Bussi-2019">Template:Cite journal</ref> Likely for ease, the thylakoid system is still commonly depicted by older "hub and spoke" models where the grana are connected to each other by tubes of stromal thylakoids.<ref>Template:Cite web</ref>

Grana consist of a stacks of flattened circular granal thylakoids that resemble pancakes. Each granum can contain anywhere from two to a hundred thylakoids,<ref name="Burgess-1989a" /> though grana with 10–20 thylakoids are most common.<ref name="Mustárdy-2008" /> Wrapped around the grana are multiple parallel right-handed helical stromal thylakoids, also known as frets or lamellar thylakoids. The helices ascend at an angle of ~20°, connecting to each granal thylakoid at a bridge-like slit junction.<ref name="Mustárdy-2008" /><ref name="Austin-2011" /><ref name="Bussi-2019" />

The stroma lamellae extend as large sheets perpendicular to the grana columns. These sheets are connected to the right-handed helices either directly or through bifurcations that form left-handed helical membrane surfaces.<ref name="Bussi-2019" /> The left-handed helical surfaces have a similar tilt angle to the right-handed helices (~20°), but ¼ the pitch. Approximately 4 left-handed helical junctions are present per granum, resulting in a pitch-balanced array of right- and left-handed helical membrane surfaces of different radii and pitch that consolidate the network with minimal surface and bending energies.<ref name="Bussi-2019" /> While different parts of the thylakoid system contain different membrane proteins, the thylakoid membranes are continuous and the thylakoid space they enclose form a single continuous labyrinth.<ref name="Mustárdy-2008" />

Thylakoid composition

Embedded in the thylakoid membranes are important protein complexes which carry out the light reactions of photosynthesis. Photosystem II and photosystem I contain light-harvesting complexes with chlorophyll and carotenoids that absorb light energy and use it to energize electrons. Molecules in the thylakoid membrane use the energized electrons to pump hydrogen ions into the thylakoid space, decreasing the pH and turning it acidic. ATP synthase is a large protein complex that harnesses the concentration gradient of the hydrogen ions in the thylakoid space to generate ATP energy as the hydrogen ions flow back out into the stroma—much like a dam turbine.<ref name="Campbell-2009b" />

There are two types of thylakoids—granal thylakoids, which are arranged in grana, and stromal thylakoids, which are in contact with the stroma. Granal thylakoids are pancake-shaped circular disks about 300–600 nanometers in diameter. Stromal thylakoids are helicoid sheets that spiral around grana.<ref name="Mustárdy-2008" /> The flat tops and bottoms of granal thylakoids contain only the relatively flat photosystem II protein complex. This allows them to stack tightly, forming grana with many layers of tightly appressed membrane, called granal membrane, increasing stability and surface area for light capture.<ref name="Mustárdy-2008" />

In contrast, photosystem I and ATP synthase are large protein complexes which jut out into the stroma. They can't fit in the appressed granal membranes, and so are found in the stromal thylakoid membrane—the edges of the granal thylakoid disks and the stromal thylakoids. These large protein complexes may act as spacers between the sheets of stromal thylakoids.<ref name="Mustárdy-2008" />

The number of thylakoids and the total thylakoid area of a chloroplast is influenced by light exposure. Shaded chloroplasts contain larger and more grana with more thylakoid membrane area than chloroplasts exposed to bright light, which have smaller and fewer grana and less thylakoid area. Thylakoid extent can change within minutes of light exposure or removal.<ref name="Wise-2007a" />

Pigments and chloroplast colors

Inside the photosystems embedded in chloroplast thylakoid membranes are various photosynthetic pigments, which absorb and transfer light energy. The types of pigments found are different in various groups of chloroplasts, and are responsible for a wide variety of chloroplast colorations. Other plastid types, such as the leucoplast and the chromoplast, contain little chlorophyll and do not carry out photosynthesis.

Paper chromatography of some spinach leaf extract shows the various pigments present in their chloroplasts.

Xanthophylls

Chlorophyll a

Chlorophyll b

Chlorophylls

Chlorophyll a is found in all chloroplasts, as well as their cyanobacterial ancestors. Chlorophyll a is a blue-green pigment<ref name="Campbell-2009e">Template:Cite book</ref> partially responsible for giving most cyanobacteria and chloroplasts their color. Other forms of chlorophyll exist, such as the accessory pigments chlorophyll b, chlorophyll c, chlorophyll d,<ref name="Kim-2009" /> and chlorophyll f.

Chlorophyll b is an olive green pigment found only in the chloroplasts of plants, green algae, any secondary chloroplasts obtained through the secondary endosymbiosis of a green alga, and a few cyanobacteria.<ref name="Kim-2009" /> It is the chlorophylls a and b together that make most plant and green algal chloroplasts green.<ref name="Campbell-2009e" />

Chlorophyll c is mainly found in secondary endosymbiotic chloroplasts that originated from a red alga, although it is not found in chloroplasts of red algae themselves. Chlorophyll c is also found in some green algae and cyanobacteria.<ref name="Kim-2009" />

Chlorophylls d and f are pigments found only in some cyanobacteria.<ref name="Kim-2009" /><ref name="UniSydney-2010">Template:Cite web</ref>

Carotenoids

In addition to chlorophylls, another group of yellow–orange<ref name="Campbell-2009e" /> pigments called carotenoids are also found in the photosystems. There are about thirty photosynthetic carotenoids.<ref name="Takaichi-2011">Template:Cite journal</ref> They help transfer and dissipate excess energy,<ref name="Kim-2009" /> and their bright colors sometimes override the chlorophyll green, like during the fall, when the leaves of some land plants change color.<ref>Template:Cite web</ref> β-carotene is a bright red-orange carotenoid found in nearly all chloroplasts, like chlorophyll a.<ref name="Kim-2009" /> Xanthophylls, especially the orange-red zeaxanthin, are also common.<ref name="Takaichi-2011" /> Many other forms of carotenoids exist that are only found in certain groups of chloroplasts.<ref name="Kim-2009" />

Phycobilins

Phycobilins are a third group of pigments found in cyanobacteria, and glaucophyte, red algal, and cryptophyte chloroplasts.<ref name="Kim-2009" /><ref name="Howe-2008">Template:Cite journal</ref> Phycobilins come in all colors, though phycoerytherin is one of the pigments that makes many red algae red.<ref>Template:Cite web</ref> Phycobilins often organize into relatively large protein complexes about 40 nanometers across called phycobilisomes.<ref name="Kim-2009" /> Like photosystem I and ATP synthase, phycobilisomes jut into the stroma, preventing thylakoid stacking in red algal chloroplasts.<ref name="Kim-2009" /> Cryptophyte chloroplasts and some cyanobacteria don't have their phycobilin pigments organized into phycobilisomes, and keep them in their thylakoid space instead.<ref name="Kim-2009" />

Photosynthetic pigments. Presence of pigments across chloroplast groups and cyanobacteria. Colored cells represent pigment presence. Chl = chlorophyll<ref name="Kim-2009" /><ref name="Takaichi-2011" /><ref name="Howe-2008" />
	Chl a	Chl b	Chl c	Chl d and f	Xanthophylls	α-carotene	β-carotene	Phycobilins
Land plants
Green algae
Euglenophytes and Chlorarachniophytes
Euglenophytes and Chlorarachniophytes
Multicellular red algae
Unicellular red algae
Haptophytes and Dinophytes
Haptophytes and Dinophytes
Cryptophytes
Glaucophytes
Cyanobacteria

Specialized chloroplasts in Template:C4 plants

To fix carbon dioxide into sugar molecules in the process of photosynthesis, chloroplasts use an enzyme called RuBisCO. RuBisCO has trouble distinguishing between carbon dioxide and oxygen, so at high oxygen concentrations, RuBisCO starts accidentally adding oxygen to sugar precursors. This has the result of ATP energy being wasted and Template:CO2 being released, all with no sugar being produced. This is a big problem, since OTemplate:Sub is produced by the initial light reactions of photosynthesis, causing issues down the line in the Calvin cycle which uses RuBisCO.<ref name="Campbell-2009d">Template:Cite book</ref>

[[C4 plants|Template:C4 plants]] evolved a way to solve this—by spatially separating the light reactions and the Calvin cycle. The light reactions, which store light energy in ATP and NADPH, are done in the mesophyll cells of a Template:C4 leaf. The Calvin cycle, which uses the stored energy to make sugar using RuBisCO, is done in the bundle sheath cells, a layer of cells surrounding a vein in a leaf.<ref name="Campbell-2009d" />

As a result, chloroplasts in Template:C4 mesophyll cells and bundle sheath cells are specialized for each stage of photosynthesis. In mesophyll cells, chloroplasts are specialized for the light reactions, so they lack RuBisCO, and have normal grana and thylakoids,<ref name="Gunning-1996a">Template:Cite book</ref> which they use to make ATP and NADPH, as well as oxygen. They store Template:CO2 in a four-carbon compound, which is why the process is called [[C4 carbon fixation|Template:C4 photosynthesis]]. The four-carbon compound is then transported to the bundle sheath chloroplasts, where it drops off Template:CO2 and returns to the mesophyll. Bundle sheath chloroplasts do not carry out the light reactions, preventing oxygen from building up in them and disrupting RuBisCO activity.<ref name="Campbell-2009d" /> Because of this, they lack thylakoids organized into grana stacks—though bundle sheath chloroplasts still have free-floating thylakoids in the stroma where they still carry out cyclic electron flow, a light-driven method of synthesizing ATP to power the Calvin cycle without generating oxygen. They lack photosystem II, and only have photosystem I—the only protein complex needed for cyclic electron flow.<ref name="Gunning-1996a" /><ref name="Campbell-2009d" /> Because the job of bundle sheath chloroplasts is to carry out the Calvin cycle and make sugar, they often contain large starch grains.<ref name="Gunning-1996a" />

Both types of chloroplast contain large amounts of chloroplast peripheral reticulum,<ref name="Gunning-1996a" /> which they use to get more surface area to transport stuff in and out of them.<ref name="Whatley-1994" /><ref name="Wise-2007a">Template:Cite book</ref> Mesophyll chloroplasts have a little more peripheral reticulum than bundle sheath chloroplasts.<ref name="Lawton-1988">Template:Cite journal</ref>

Function and chemistry

Guard cell chloroplasts

Unlike most epidermal cells, the guard cells of plant stomata contain relatively well-developed chloroplasts.<ref name="Lawson">Lawson T. and J. I. L. Morison. Essay 10.1 Guard Cell Photosynthesis. Plant Physiology and Development, Sixth Edition [1]</ref> However, exactly what they do is controversial.<ref>Template:Cite journal</ref>

Plant innate immunity

Plants lack specialized immune cells—all plant cells participate in the plant immune response. Chloroplasts, along with the nucleus, cell membrane, and endoplasmic reticulum,<ref name="Padmanabhan-2010">Template:Cite journal</ref> are key players in pathogen defense. Due to its role in a plant cell's immune response, pathogens frequently target the chloroplast.<ref name="Padmanabhan-2010" />

Plants have two main immune responses—the hypersensitive response, in which infected cells seal themselves off and undergo programmed cell death, and systemic acquired resistance, where infected cells release signals warning the rest of the plant of a pathogen's presence. Chloroplasts stimulate both responses by purposely damaging their photosynthetic system, producing reactive oxygen species. High levels of reactive oxygen species will cause the hypersensitive response. The reactive oxygen species also directly kill any pathogens within the cell. Lower levels of reactive oxygen species initiate systemic acquired resistance, triggering defense-molecule production in the rest of the plant.<ref name="Padmanabhan-2010" />

In some plants, chloroplasts are known to move closer to the infection site and the nucleus during an infection.<ref name="Padmanabhan-2010" />

Chloroplasts can serve as cellular sensors. After detecting stress in a cell, which might be due to a pathogen, chloroplasts begin producing molecules like salicylic acid, jasmonic acid, nitric oxide and reactive oxygen species which can serve as defense-signals. As cellular signals, reactive oxygen species are unstable molecules, so they probably don't leave the chloroplast, but instead pass on their signal to an unknown second messenger molecule. All these molecules initiate retrograde signaling—signals from the chloroplast that regulate gene expression in the nucleus.<ref name="Padmanabhan-2010" />

In addition to defense signaling, chloroplasts, with the help of the peroxisomes,<ref name="Katsir-2008">Template:Cite journal</ref> help synthesize an important defense molecule, jasmonate. Chloroplasts synthesize all the fatty acids in a plant cell<ref name="Padmanabhan-2010" /><ref name="Schnurr-2002">Template:Cite journal</ref>—linoleic acid, a fatty acid, is a precursor to jasmonate.<ref name="Padmanabhan-2010" />

Photosynthesis

One of the main functions of the chloroplast is its role in photosynthesis, the process by which light is transformed into chemical energy, to subsequently produce food in the form of sugars. Water (H₂O) and carbon dioxide (CO₂) are used in photosynthesis, and sugar and oxygen (O₂) are made, using light energy. Photosynthesis is divided into two stages—the light reactions, where water is split to produce oxygen, and the dark reactions, or Calvin cycle, which builds sugar molecules from carbon dioxide. The two phases are linked by the energy carriers adenosine triphosphate (ATP) and nicotinamide adenine dinucleotide phosphate (NADP⁺).<ref name="Pearson-2009" /><ref name="Campbell-2006">Template:Cite book Template:Page needed</ref>

Light reactions

The light reactions take place on the thylakoid membranes. They take light energy and store it in NADPH, a form of NADP⁺, and ATP to fuel the dark reactions.

Energy carriers

ATP is the phosphorylated version of adenosine diphosphate (ADP), which stores energy in a cell and powers most cellular activities. ATP is the energized form, while ADP is the (partially) depleted form. NADP⁺ is an electron carrier which ferries high energy electrons. In the light reactions, it gets reduced, meaning it picks up electrons, becoming NADPH.

Photophosphorylation

Like mitochondria, chloroplasts use the potential energy stored in an H⁺, or hydrogen ion, gradient to generate ATP energy. The two photosystems capture light energy to energize electrons taken from water, and release them down an electron transport chain. The molecules between the photosystems harness the electrons' energy to pump hydrogen ions into the thylakoid space, creating a concentration gradient, with more hydrogen ions (up to a thousand times as many)<ref name="Campbell-2009b" /> inside the thylakoid system than in the stroma. The hydrogen ions in the thylakoid space then diffuse back down their concentration gradient, flowing back out into the stroma through ATP synthase. ATP synthase uses the energy from the flowing hydrogen ions to phosphorylate adenosine diphosphate into adenosine triphosphate, or ATP.<ref name="Campbell-2009b" /><ref>Template:Cite journal</ref> Because chloroplast ATP synthase projects out into the stroma, the ATP is synthesized there, in position to be used in the dark reactions.<ref name="Berg-2002b">Template:Cite book</ref>

NADP⁺ reduction

Electrons are often removed from the electron transport chains to charge NADP⁺ with electrons, reducing it to NADPH. Like ATP synthase, ferredoxin-NADP⁺ reductase, the enzyme that reduces NADP⁺, releases the NADPH it makes into the stroma, right where it is needed for the dark reactions.<ref name="Berg-2002b" />

Because NADP⁺ reduction removes electrons from the electron transport chains, they must be replaced—the job of photosystem II, which splits water molecules (H₂O) to obtain the electrons from its hydrogen atoms.<ref name="Campbell-2009b" /><ref name="Pearson-2009">Template:Cite book</ref>

Cyclic photophosphorylation

While photosystem II photolyzes water to obtain and energize new electrons, photosystem I simply reenergizes depleted electrons at the end of an electron transport chain. Normally, the reenergized electrons are taken by NADP⁺, though sometimes they can flow back down more H⁺-pumping electron transport chains to transport more hydrogen ions into the thylakoid space to generate more ATP. This is termed cyclic photophosphorylation because the electrons are recycled. Cyclic photophosphorylation is common in [[C4 plants|Template:C4 plants]], which need more ATP than NADPH.<ref name="Campbell-2009d" />

Dark reactions

The Calvin cycle, also known as the dark reactions, is a series of biochemical reactions that fixes CO₂ into G3P sugar molecules and uses the energy and electrons from the ATP and NADPH made in the light reactions. The Calvin cycle takes place in the stroma of the chloroplast.<ref name="Campbell-2009d" />

While named "the dark reactions", in most plants, they take place in the light, since the dark reactions are dependent on the products of the light reactions.<ref name="Campbell-2009g" />

Carbon fixation and G3P synthesis

The Calvin cycle starts by using the enzyme RuBisCO to fix CO₂ into five-carbon Ribulose bisphosphate (RuBP) molecules. The result is unstable six-carbon molecules that immediately break down into three-carbon molecules called 3-phosphoglyceric acid, or 3-PGA. The ATP and NADPH made in the light reactions is used to convert the 3-PGA into glyceraldehyde-3-phosphate, or G3P sugar molecules. Most of the G3P molecules are recycled back into RuBP using energy from more ATP, but one out of every six produced leaves the cycle—the end product of the dark reactions.<ref name="Campbell-2009d" />

Sugars and starches

Glyceraldehyde-3-phosphate can double up to form larger sugar molecules like glucose and fructose. These molecules are processed, and from them, the still larger sucrose, a disaccharide commonly known as table sugar, is made, though this process takes place outside of the chloroplast, in the cytoplasm.<ref name="Berg-2002a">Template:Cite book</ref>

Alternatively, glucose monomers in the chloroplast can be linked together to make starch, which accumulates into the starch grains found in the chloroplast.<ref name="Berg-2002a" /> Under conditions such as high atmospheric CO₂ concentrations, these starch grains may grow very large, distorting the grana and thylakoids. The starch granules displace the thylakoids, but leave them intact.<ref name="Wample-1983" /> Waterlogged roots can also cause starch buildup in the chloroplasts, possibly due to less sucrose being exported out of the chloroplast (or more accurately, the plant cell). This depletes a plant's free phosphate supply, which indirectly stimulates chloroplast starch synthesis.<ref name="Wample-1983">Template:Cite journal</ref> While linked to low photosynthesis rates, the starch grains themselves may not necessarily interfere significantly with the efficiency of photosynthesis,<ref>Template:Cite journal</ref> and might simply be a side effect of another photosynthesis-depressing factor.<ref name="Wample-1983" />

Photorespiration

Photorespiration can occur when the oxygen concentration is too high. RuBisCO cannot distinguish between oxygen and carbon dioxide very well, so it can accidentally add O₂ instead of CO₂ to RuBP. This process reduces the efficiency of photosynthesis—it consumes ATP and oxygen, releases CO₂, and produces no sugar. It can waste up to half the carbon fixed by the Calvin cycle.<ref name="Pearson-2009" /> Several mechanisms have evolved in different lineages that raise the carbon dioxide concentration relative to oxygen within the chloroplast, increasing the efficiency of photosynthesis. These mechanisms are called carbon dioxide concentrating mechanisms, or CCMs. These include Crassulacean acid metabolism, [[C4 carbon fixation|Template:C4 carbon fixation]],<ref name="Pearson-2009" /> and pyrenoids. Chloroplasts in Template:C4 plants are notable as they exhibit a distinct chloroplast dimorphism.

pH

Because of the H⁺ gradient across the thylakoid membrane, the interior of the thylakoid is acidic, with a pH around 4,<ref>Template:Cite book</ref> while the stroma is slightly basic, with a pH of around 8.<ref name="Hauser-1995">Template:Cite journal</ref> The optimal stroma pH for the Calvin cycle is 8.1, with the reaction nearly stopping when the pH falls below 7.3.<ref name="Werdan-1975" />

CO₂ in water can form carbonic acid, which can disturb the pH of isolated chloroplasts, interfering with photosynthesis, even though CO₂ is used in photosynthesis. However, chloroplasts in living plant cells are not affected by this as much.<ref name="Hauser-1995" />

Chloroplasts can pump K⁺ and H⁺ ions in and out of themselves using a poorly understood light-driven transport system.<ref name="Hauser-1995" />

In the presence of light, the pH of the thylakoid lumen can drop up to 1.5 pH units, while the pH of the stroma can rise by nearly one pH unit.<ref name="Werdan-1975">Template:Cite journal</ref>

Amino acid synthesis

Chloroplasts alone make almost all of a plant cell's amino acids in their stroma<ref name="Burgess-1989e" /> except the sulfur-containing ones like cysteine and methionine.<ref name="Ferro-2002">Template:Cite journal</ref><ref name="Rolland-1992" /> Cysteine is made in the chloroplast (the proplastid too) but it is also synthesized in the cytosol and mitochondria, probably because it has trouble crossing membranes to get to where it is needed.<ref name="Rolland-1992">Template:Cite journal</ref> The chloroplast is known to make the precursors to methionine but it is unclear whether the organelle carries out the last leg of the pathway or if it happens in the cytosol.<ref name="Ravanel-1998">Template:Cite journal</ref>

Other nitrogen compounds

Chloroplasts make all of a cell's purines and pyrimidines—the nitrogenous bases found in DNA and RNA.<ref name="Burgess-1989e" /> They also convert nitrite (NO₂⁻) into ammonia (NH₃) which supplies the plant with nitrogen to make its amino acids and nucleotides.<ref name="Burgess-1989e" />

Other chemical products

The plastid is the site of diverse and complex lipid synthesis in plants.<ref name="Buchanan BB">Buchanan BB, Gruissem W, Jones RL (Eds.). 2015. Biochemistry & Molecular Biology of Plants. Wiley Blackwell.</ref><ref name="Joyard-2010">Template:Cite journal</ref> The carbon used to form the majority of the lipid is from acetyl-CoA, which is the decarboxylation product of pyruvate.<ref name="Buchanan BB" /> Pyruvate may enter the plastid from the cytosol by passive diffusion through the membrane after production in glycolysis.<ref name="Proudlove-1981">Template:Cite journal</ref> Pyruvate is also made in the plastid from phosphoenolpyruvate, a metabolite made in the cytosol from pyruvate or PGA.<ref name="Buchanan BB" /> Acetate in the cytosol is unavailable for lipid biosynthesis in the plastid.<ref name="Bao-2000">Bao X, Focke M, Pollard M, Ohlrogge J. 2000. Understanding in vivo carbon precursor supply for fatty acid synthesis in leaf tissue. Plant Journal 22, 39–50.</ref> The typical length of fatty acids produced in the plastid are 16 or 18 carbons, with 0-3 cis double bonds.<ref name="Ohlrogge-1995">Template:Cite journal</ref>

The biosynthesis of fatty acids from acetyl-CoA primarily requires two enzymes. Acetyl-CoA carboxylase creates malonyl-CoA, used in both the first step and the extension steps of synthesis. Fatty acid synthase (FAS) is a large complex of enzymes and cofactors including acyl carrier protein (ACP) which holds the acyl chain as it is synthesized. The initiation of synthesis begins with the condensation of malonyl-ACP with acetyl-CoA to produce ketobutyryl-ACP. 2 reductions involving the use of NADPH and one dehydration creates butyryl-ACP. Extension of the fatty acid comes from repeated cycles of malonyl-ACP condensation, reduction, and dehydration.<ref name="Buchanan BB" />

Other lipids are derived from the methyl-erythritol phosphate (MEP) pathway and consist of gibberelins, sterols, abscisic acid, phytol, and innumerable secondary metabolites.<ref name="Buchanan BB" />

Location

Distribution in a plant

Not all cells in a multicellular plant contain chloroplasts. All green parts of a plant contain chloroplasts as the color comes from the chlorophyll.<ref name="Campbell-2009g" /> The plant cells which contain chloroplasts are usually parenchyma cells, though chloroplasts can also be found in collenchyma tissue.<ref>Template:Cite book</ref> A plant cell which contains chloroplasts is known as a chlorenchyma cell. A typical chlorenchyma cell of a land plant contains about 10 to 100 chloroplasts.

In some plants such as cacti, chloroplasts are found in the stems,<ref>Template:Cite book</ref> though in most plants, chloroplasts are concentrated in the leaves. One square millimeter of leaf tissue can contain half a million chloroplasts.<ref name="Campbell-2009g" /> Within a leaf, chloroplasts are mainly found in the mesophyll layers of a leaf, and the guard cells of stomata. Palisade mesophyll cells can contain 30–70 chloroplasts per cell, while stomatal guard cells contain only around 8–15 per cell, as well as much less chlorophyll. Chloroplasts can also be found in the bundle sheath cells of a leaf, especially in [[C4 plants|CTemplate:Sub plants]], which carry out the Calvin cycle in their bundle sheath cells. They are often absent from the epidermis of a leaf.<ref name="Lawson" />

Cellular location

Chloroplast movement

The chloroplasts of plant and algal cells can orient themselves to best suit the available light. In low-light conditions, they will spread out in a sheet—maximizing the surface area to absorb light. Under intense light, they will seek shelter by aligning in vertical columns along the plant cell's cell wall or turning sideways so that light strikes them edge-on. This reduces exposure and protects them from photooxidative damage.<ref name="Wells-1979">Template:Cite journal</ref> This ability to distribute chloroplasts so that they can take shelter behind each other or spread out may be the reason why land plants evolved to have many small chloroplasts instead of a few big ones.<ref name="Glynn-2007" /> Chloroplast movement is considered one of the most closely regulated stimulus-response systems that can be found in plants.<ref name="Dong-1998">Template:Cite journal</ref> Mitochondria have also been observed to follow chloroplasts as they move.<ref name="Takagi-2003">Template:Cite journal</ref>

In higher plants, chloroplast movement is run by phototropins, blue light photoreceptors also responsible for plant phototropism. In some algae, mosses, ferns, and flowering plants, chloroplast movement is influenced by red light in addition to blue light,<ref name="Wells-1979" /> though very long red wavelengths inhibit movement rather than speeding it up. Blue light generally causes chloroplasts to seek shelter, while red light draws them out to maximize light absorption.<ref name="Takagi-2003" />

Studies of Vallisneria gigantea, an aquatic flowering plant, have shown that chloroplasts can get moving within five minutes of light exposure, though they don't initially show any net directionality. They may move along microfilament tracks, and the fact that the microfilament mesh changes shape to form a honeycomb structure surrounding the chloroplasts after they have moved suggests that microfilaments may help to anchor chloroplasts in place.<ref name="Dong-1998" /><ref name="Takagi-2003" />

Differentiation, replication, and inheritance

Chloroplasts are a special type of a plant cell organelle called a plastid, though the two terms are sometimes used interchangeably. There are many other types of plastids, which carry out various functions. All chloroplasts in a plant are descended from undifferentiated proplastids found in the zygote,<ref name="Burgess-1989e">Template:Cite book</ref> or fertilized egg. Proplastids are commonly found in an adult plant's apical meristems. Chloroplasts do not normally develop from proplastids in root tip meristems<ref name="Gunning-1996b">Template:Cite book</ref>—instead, the formation of starch-storing amyloplasts is more common.<ref name="Burgess-1989e" />

In shoots, proplastids from shoot apical meristems can gradually develop into chloroplasts in photosynthetic leaf tissues as the leaf matures, if exposed to the required light.<ref name="Alberts-2002a" /> This process involves invaginations of the inner plastid membrane, forming sheets of membrane that project into the internal stroma. These membrane sheets then fold to form thylakoids and grana.<ref name="Burgess-1989d">Template:Cite book</ref>

If angiosperm shoots are not exposed to the required light for chloroplast formation, proplastids may develop into an etioplast stage before becoming chloroplasts. An etioplast is a plastid that lacks chlorophyll, and has inner membrane invaginations that form a lattice of tubes in their stroma, called a prolamellar body. While etioplasts lack chlorophyll, they have a yellow chlorophyll precursor stocked.<ref name="Alberts-2002a" /> Within a few minutes of light exposure, the prolamellar body begins to reorganize into stacks of thylakoids, and chlorophyll starts to be produced. This process, where the etioplast becomes a chloroplast, takes several hours.<ref name="Burgess-1989d" /> Gymnosperms do not require light to form chloroplasts.<ref name="Burgess-1989d" />

Light, however, does not guarantee that a proplastid will develop into a chloroplast. Whether a proplastid develops into a chloroplast some other kind of plastid is mostly controlled by the nucleus<ref name="Alberts-2002a" /> and is largely influenced by the kind of cell it resides in.<ref name="Burgess-1989e" />

Plastid interconversion

Plastid differentiation is not permanent, in fact many interconversions are possible. Chloroplasts may be converted to chromoplasts, which are pigment-filled plastids responsible for the bright colors seen in flowers and ripe fruit. Starch storing amyloplasts can also be converted to chromoplasts, and it is possible for proplastids to develop straight into chromoplasts. Chromoplasts and amyloplasts can also become chloroplasts, like what happens when a carrot or a potato is illuminated. If a plant is injured, or something else causes a plant cell to revert to a meristematic state, chloroplasts and other plastids can turn back into proplastids. Chloroplast, amyloplast, chromoplast, proplastid are not absolute; state—intermediate forms are common.<ref name="Burgess-1989e" />

Division

Most chloroplasts in a photosynthetic cell do not develop directly from proplastids or etioplasts. In fact, a typical shoot meristematic plant cell contains only 7–20 proplastids. These proplastids differentiate into chloroplasts, which divide to create the 30–70 chloroplasts found in a mature photosynthetic plant cell. If the cell divides, chloroplast division provides the additional chloroplasts to partition between the two daughter cells.<ref name="Burgess-1989c">Template:Cite book</ref>

In single-celled algae, chloroplast division is the only way new chloroplasts are formed. There is no proplastid differentiation—when an algal cell divides, its chloroplast divides along with it, and each daughter cell receives a mature chloroplast.<ref name="Burgess-1989d" />

Almost all chloroplasts in a cell divide, rather than a small group of rapidly dividing chloroplasts.<ref name="Possingham-1976">Template:Cite journal</ref> Chloroplasts have no definite S-phase—their DNA replication is not synchronized or limited to that of their host cells.<ref name="Cannon-1993">Template:Cite journal</ref> Much of what we know about chloroplast division comes from studying organisms like Arabidopsis and the red alga Cyanidioschyzon merolæ.<ref name="Glynn-2007" />

The division process starts when the proteins FtsZ1 and FtsZ2 assemble into filaments, and with the help of a protein ARC6, form a structure called a Z-ring within the chloroplast's stroma.<ref name="Glynn-2007">Template:Cite journal</ref><ref name="Miyagishima-2003">Template:Cite journal</ref> The Min system manages the placement of the Z-ring, ensuring that the chloroplast is cleaved more or less evenly. The protein MinD prevents FtsZ from linking up and forming filaments. Another protein ARC3 may also be involved, but it is not very well understood. These proteins are active at the poles of the chloroplast, preventing Z-ring formation there, but near the center of the chloroplast, MinE inhibits them, allowing the Z-ring to form.<ref name="Glynn-2007" />

Next, the two plastid-dividing rings, or PD rings form. The inner plastid-dividing ring is located in the inner side of the chloroplast's inner membrane, and is formed first.<ref name="Glynn-2007" /> The outer plastid-dividing ring is found wrapped around the outer chloroplast membrane. It consists of filaments about 5 nanometers across,<ref name="Glynn-2007" /> arranged in rows 6.4 nanometers apart, and shrinks to squeeze the chloroplast. This is when chloroplast constriction begins.<ref name="Miyagishima-2003" />
In a few species like Cyanidioschyzon merolæ, chloroplasts have a third plastid-dividing ring located in the chloroplast's intermembrane space.<ref name="Glynn-2007" /><ref name="Miyagishima-2003" />

Late into the constriction phase, dynamin proteins assemble around the outer plastid-dividing ring,<ref name="Miyagishima-2003" /> helping provide force to squeeze the chloroplast.<ref name="Glynn-2007" /> Meanwhile, the Z-ring and the inner plastid-dividing ring break down.<ref name="Miyagishima-2003" /> During this stage, the many chloroplast DNA plasmids floating around in the stroma are partitioned and distributed to the two forming daughter chloroplasts.<ref name="Hashimoto-1989">Template:Cite journal</ref>

Later, the dynamins migrate under the outer plastid dividing ring, into direct contact with the chloroplast's outer membrane,<ref name="Miyagishima-2003" /> to cleave the chloroplast in two daughter chloroplasts.<ref name="Glynn-2007" />

A remnant of the outer plastid dividing ring remains floating between the two daughter chloroplasts, and a remnant of the dynamin ring remains attached to one of the daughter chloroplasts.<ref name="Miyagishima-2003" />

Of the five or six rings involved in chloroplast division, only the outer plastid-dividing ring is present for the entire constriction and division phase—while the Z-ring forms first, constriction does not begin until the outer plastid-dividing ring forms.<ref name="Miyagishima-2003" />