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Plant Physiol, January 2001, Vol. 125, pp. 50-53
Chloroplast Origin and Integration1
Geoffrey I.
McFadden*
Plant Cell Biology Research Centre, School of Botany, University of
Melbourne, Victoria 3010, Australia
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INTRODUCTION |
The origin of oxygenic
photosynthesis must rank just after the origin of life itself as one of
the most significant events in the history of life. The early
photosynthesizers, which Bill Schopf has shown were similar to modern
cyanobacteria (32), made earth a nicer place to live by quietly
cranking out oxygen over hundreds of millions of years. Having
established an ozone UV shield, the cyanobacteria then entered into an
extraordinary partnership with eukaryotic cells: They became
chloroplasts. Equipped with chloroplasts, the early plants were
ready to colonize the land and green the planet. This review recounts
major leaps in our understanding of chloroplast evolution from the
preceding 25 years.
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ENDOSYMBIOSIS TRIUMPHS |
In 1975 the theory of endosymbiosis (which describes the origins
of mitochondria and plastids from eubacteria-like cells living within
eukaryotic hosts) was still hanging in the balance. Major players like
Lawrence Bogorad (4) and Tom Cavalier-Smith (7) were weighing the
merits of the autogenous (nonendosymbiotic) alternatives for
chloroplasts against the early compelling scenarios of Mereschkowsky and later Lynn Margulis. However, in the same year, an innovative paper
from Linda Bonen and Ford Doolittle (5; which provided the first
quantitative measure of similarities between T1 rRNA catalogues of
cyanobacteria and chloroplasts) is now recognizable as the first ripple
in a tidal wave of chloroplast molecular data that swept the autogenous
origin hypothesis away.
Hans Kössel was one of the first to apply the recently developed
Maxan/Gilbert DNA sequencing technology to chloroplasts. Primary and
secondary structure of maize chloroplast rRNA was revealed to be more
closely related to the bacterium Escherichia coli than to
equivalent genes in the nuclei of eukaryotes (the yeast sequence only
became available to Kössel at the proof stage!), corroborating
the endosymbiotic hypothesis (34).
Flügge and Heldt recognized that integration of the endosymbiont
required a transporter for export of photosynthate and characterized a
family of phosphate translocators located in the chloroplast envelope
(12, 13). It has subsequently emerged that plants (and probably algae)
have learned to operate these transporters in reverse to "feed"
their chloroplasts at night or in organs underground. Norman Weeden
then recognized that the endosymbiont, in addition to the gift of
photosynthesis, was responsible for the introgression of several other
metabolic pathways into the host (38). Weeden's insight was the spur
for the ongoing characterization of the bacterial-type chloroplast
pathways for amino acid biosynthesis, carotenoid biosynthesis, non-Shemin tetrapyrole biosynthesis, non-mevalonate isoprenoid biosynthesis, and nitrate and sulfate assimilation. Weeden also recognized that many of the genes for these processes must have undergone intracellular transfer from plastid to nucleus.
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ONE RING TO RULE THEM... AND IN THE DARKNESS BIND THEM |
The advent of Sanger-style DNA sequencing opened the way to more
ambitious projects. Oyhama's team (28) was first to sequence a
chloroplast genome determining the 121,024 bp of the liverwort Marchantia polymorpha chloroplast. The Japanese group's
work inspired numerous other chloroplast genome sequencing projects
from which various patterns, and a few surprises, have emerged. The
Marchantia genome has proven to be a good general model.
Typical chloroplast DNAs, including those of plants and most algae, are
circular with genes organized in operons. Plastid gene content varies
from about 70 in some non-photosynthetic plastids such as that of the
malaria parasite (39) up to about 200 in red algae (30); plants
typically have around 100 genes in the chloroplast. Dinoflagellate
algal chloroplasts are the only example that so far seriously defy this trend, having single gene mini-circles that encode only a handful of
genes (40). One ring no longer binds them.
The next major leap in the application of sequencing was the complete
genome of the cyanobacterium of Synechocystis (20). The
Japanese team identified 3,229 genes, which provided a reference point
for the gene content of the chloroplast endosymbiont at the outset of
endosymbiosis. For the first time we held a list of genes with which
the endosymbiont probably started. Comparisons of this list with the
present content of plastid genomes issues some weighty challenges.
The genome sequences also provided an opportunity to compare gene
synteny between chloroplasts and bacteria. Conservation of large
operons proved that chloroplast DNAs are reduced bacterial genomes
(28). In a converse manner, unique gene assemblages shared among all
chloroplasts suggest that all chloroplasts arose from a single primary
endosymbiosis (30), as first argued by Tom Cavalier-Smith (8). Many
other studies now concur, but the idea is not universally accepted
(23). Large amounts of sequence data have also allowed meta-analyses in
which multiple genes are used in determining more robust phylogenies
(25). These trees are consistent with the belief that the glaucophyte
alga Cyanophora, which has a blue-green chloroplast with a
peptidoglycan wall, is one of the earliest diverging lines to contain a
chloroplast. Meta-analysis also confirms morphological clues suggesting
that the prasinophyte Mesostigma is the closest living
relative to the unicellular alga from which the land plants are
descended (22).
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INTRACELLULAR GENE TRANSFER AND PROTEIN TARGETING |
An important outcome of comparing the gene content of a
cyanobacterium with that of chloroplasts is the detailed reconstruction of genome reduction. When it was first established that nuclear genes
encode some types of chloroplast proteins it became clear that
chloroplasts were genetically only semi-autonomous and had relinquished
genes to their hosts. The genome maps in conjunction with the phylogeny
tell us that this process occurred in a somewhat ad hoc manner,
although certain classes of genes are apparently less amenable to
transfer (25). Two intriguing questions are how many nuclear genes
service the chloroplast and are they all derived from the endosymbiont
cyanobacterium. The Arabidopsis genome project is beginning to allow us
to address these questions; early surveys indicate that as many as
2,000 nuclear genes encode proteins with plastid functions (1). The
tagged mutagenesis program also provides a means to assay the functions
of those genes whose products are predicted to be chloroplast targeted,
but for which no obvious function is apparent.
The transfer of genes from chloroplast to nucleus typically requires
return passage of the gene product to its place of function. This
mechanism is central to the establishment and refinement of
endosymbiosis. The first vagrant gene to be studied was the Rubisco
small subunit (RbcS), and it was established early that a precursor
form was produced in the cytosol and targeted. It was subsequently
established that the precursor bears an N-terminal extension, the
so-called "transit peptide," which mediates transport. There were
several key developments in the understanding of this process.
Dobberstein et al. (10) established that RbcS began as a precursor with
a small (3.5 kD) extension that is proteolytically removed within the
chloroplast prior to assembly of the holoenzyme. Gregory Schmidt and
colleagues (31) determined the leader sequence and the site of
processing. Chua and Schmidt (9) and Highfield and Ellis (19)
subsequently demonstrated that the translocation into the chloroplast
occurs post-translationally.
These experiments were important for two key reasons: first, they
showed that chloroplast targeting was different to the "signal hypothesis" for cotranslational insertion into the endomembrane system, and second, they were the germ of a system to dissect the
import process. By using in vitro translated precursors in conjunction
with isolated plastids it has been possible to manipulate the system
experimentally to identify conditions required for transport and
components of the transport machinery. Mishkind et al. (27)
demonstrated that the transit peptide was sufficient and necessary for
import of proteins. Schrier et al. (33) and van den Broek et al. (37)
established the use of transit peptides to direct foreign proteins into
chloroplasts in transgenic plants, which paved the way for targeting of
reporter proteins such as the jellyfish green fluorescent protein.
A key development in understanding the mechanisms of import was the
identification of ATP as the essential requirement for transport (16).
This dependence was exploited to interrupt transfer, which enabled the
partially transferred precursor to be used as a tag (often through
innovative use of cross linkers or affinity motifs attached to the
targeted protein) to recover components of the transport apparatus.
Approaches based on this strategy have identified numerous
transport-related proteins (known as Tocs and Tics) located in the
inner and outer chloroplast membranes such as the receptors (36) and
the channel protein (24).
The targeting of proteins within the confines of the chloroplast was
first studied by Smeekens et al. (35). They demonstrated that
plastocyanin was targeted into the chloroplast stroma by a typical
transit peptide, but that a second signal, this time homologous to
bacterial secretion peptides, directed the protein across the thylakoid
membranes. In other words, the original bacterial mechanisms for
targeting proteins across the inner membrane have been retained in
chloroplasts, a phenomenon now termed "conservative sorting." The
transit peptide was thus perceived as a prefix appended to the protein
to return it to the organelle for internal distribution.
The intense study of Rubisco allowed plant biologists to discover a
major unifying principle for the assembly of oligomeric protein
structures. John Ellis and colleagues (3) identified a 60-kD protein
essential for Rubisco assembly. Sequencing of the so-called
"Rubisco-binding protein" and the GroEL heat shock proteins of
E. coli demonstrated that these proteins, which Ellis dubbed chaperones, are fundamental and ubiquitous components of protein folding and protein/protein interactions (18). This discovery is one of
the rare instances where plant science forged the way into a unifying
concept with application across all cell biology.
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SECOND-HAND CHLOROPLASTS |
Plant chloroplasts have two bounding membranes, but chloroplasts
of many algae have three or four membranes. Sally Gibbs (15) first
articulated the idea that these multi-membraned, or complex, chloroplasts were acquired indirectly, not by the classic mechanism of
endosymbiosis of a cyanobacterial-like prokaryote. Gibbs proposed that
the Euglena chloroplast, for instance, derived from a
eukaryotic alga that had been engulfed by a eukaryotic phagotroph (15).
The engulfed cell underwent drastic reduction such that in most cases the only residues are the chloroplast and the extra membranes created
by the engulfment. Acquisition of chloroplasts in this manner occurred
multiple times (the number of acquisitions is argued hotly) and thus
explains the patchy distribution of chloroplasts across the eukaryotic
tree. The process was first verified in cryptomonads where a minute
residue of the endosymbiont nucleus has been identified (11). This
process of acquisition is known as secondary endosymbiosis, as distinct
from primary endosymbiosis between a eukaryote host and a prokaryote
endosymbiont. Secondary endosymbiosis allowed lateral transfers of
chloroplasts into non-photosynthetic lineages, including unexpected
lineages such as the malaria parasite (26, 39). Gibbs also predicted
(correctly) how proteins could be targeted across the multiple bounding
membranes by initial utilization of the secretory pathway to cross the
outermost membrane.
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REFORGING THE RING |
During its hundreds of millions of years tenure within the host,
the chloroplast genome has undergone substantial modification. Human
intervention is now beginning to make even more drastic modifications
to chloroplast DNA. Transgenics is an immensely powerful tool for
biological understanding. Chloroplast transformation was first achieved
in the green alga Chlamydomonas using a "gene gun"
approach in which selectable markers were literally blasted into the
cells (6). However, engineering of chloroplast genomes has not really
achieved widespread application. Nevertheless, chloroplast
transformation has opened doors to some exciting developments. Abrogation of the chloroplast's indigenous transcription system (knockouts of a cyanobacterial-like rpo gene in chloroplast
DNA) provided definitive evidence for the existence of an elusive
alternate polymerase (2). A single subunit, phage-type polymerase,
encoded by the nucleus and similar to that used for mitochondrial
transcription, was subsequently shown to be responsible for a major
fraction of chloroplast gene transcription (17). The dual transcription systems are thought to allow regulation of chloroplast development a hypothesis whose testing will require further transgenics.
The second major development stemming from chloroplast transformation
was the laboratory reconstruction of intracellular gene relocation. Pal
Maliga's group was able to delete the chloroplast gene for RbcL and
complement the deletion by inserting a nuclear copy, complete with a
motif to target the product, into the chloroplast (21). Perhaps the
greatest promise for chloroplast transformation lies in commercial
applications (14). The chloroplast genome has many features that make
it an ideal site for insertion of useful genes. Chloroplast DNA is
multicopy per cell, which ensures a high gene dosage and generally high
expression levels for transgenes. Unlike nuclear plant transformation,
genes can be targeted to specific sites in chloroplasts. This precision
avoids the poorly understood "position effect" whereby the site of
insertion of transgenes influences transgene behavior in an
unpredictable manner. Chloroplast transgenics makes genetic engineering
of plants more controlled. Last, because chloroplasts are maternally
inherited in angiosperms, the risk of transgene spread into the
environment through pollen is greatly reduced.
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THE FUTURE |
The breakthroughs outlined here position us to tackle some
fundamental questions in the next 25 years. We will soon have full gene
complements for the host and endosymbiont and the ability to manipulate
both genomes and target foreign proteins from the host to the
endosymbiont (there are no leads on mechanisms to do the reverse).
These approaches will be central in developing our understanding of
still mysterious processes such as chloroplast division (29), the
molecular signals regulating plastid differentiation, and the
mechanisms of cross talk between the plant cell and its little green
slaves. Armed with this information we may be in a position to
undertake some very bold experiments. We may even be able to
reconstruct endosymbiosis in the laboratory, putting chloroplasts into
non-photosynthetic hosts.
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ACKNOWLEDGMENTS |
Apologies to those whose work I could not cite. Thanks to Bill
Martin who shared his literature knowledge so enthusiastically.
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FOOTNOTES |
1
G.I.M. is supported by the Australian Research
Council and is an International Scholar of the Howard Hughes Medical Institute.
*
E-mail g.mcfadden{at}botany.unimelb.edu.au; fax
61-3-9347-1071.
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LITERATURE CITED |
-
Abdallah F, Salamini F, Leister D
(2000)
Trends Plant Sci
5: 141-142
[CrossRef][ISI][Medline]
-
Allison LA, Simon LD, Maliga P
(1996)
EMBO J
15: 2802-2809
[ISI][Medline]
-
Barraclough R, Ellis R
(1980)
Biochim Biophys Acta
608: 19-31
[Medline]
-
Bogorad L
(1975)
Science
188: 891-898
[Free Full Text]
-
Bonen L, Doolittle W
(1975)
Proc Natl Acad Sci USA
72: 2310-2314
[Abstract/Free Full Text]
-
Boynton JE, Gillham N, Harris E, Hosler JP, Johnson AM, Jones AR, Randolph-Anderson BL, Robertson D, Klein TM, Shark KB
(1988)
Science
240: 1534-1538
[Abstract/Free Full Text]
-
Cavalier-Smith T
(1975)
Nature
256: 463-468
[Medline]
-
Cavalier-Smith T
(1982)
Biol J Linn Soc
17: 289-306
-
Chua N-H, Schmidt GW
(1978)
Proc Natl Acad Sci USA
75: 6110-6114
[Abstract/Free Full Text]
-
Dobberstein B, Blobel G, Chua N-H
(1977)
Proc Natl Acad Sci USA
74: 1082-1085
[Abstract/Free Full Text]
-
Douglas SE, Murphy CA, Spencer DF, Gray MW
(1991)
Nature
350: 148-151
[CrossRef][Medline]
-
Fliege R, Flügge U, Werdan K, Heldt H
(1978)
Biochim Biophys Acta
502: 232-247
[Medline]
-
Flügge U, Fischer K, Gross A, Sebald W, Lottspeich F, Eckerskorn C
(1989)
EMBO J
8: 39-46
[ISI][Medline]
-
Garcia JS, Graves J, Hajdukiewicz P, Hunter P, Nehra N, Paradkar V, Schlittler M, Carroll J, Spatola L, Ward D, Ye G, Russell D
(2000)
Nat Biotechnol
18: 333-338
[CrossRef][ISI][Medline]
-
Gibbs SP
(1978)
Can J Bot
56: 2882-2889
-
Grossman A, Bartlett S, Chua N-H
(1980)
Nature
285: 625-628
[CrossRef]
-
Hedtke B, Borner T, Weihe A
(1997)
Science
277: 809-811
[Abstract/Free Full Text]
-
Hemmingsen S, Woolford C, Vries Svd, Tilly K, Dennis D,
Georgopoulos C, hendrix R, Ellis R (1988) Nature 330-333
-
Highfield PE, Ellis RJ
(1978)
Nature
271: 420-424
[CrossRef][Medline]
-
Kaneko T, Sato S, Kotani H, Tanaka A, Asamizu E, Nakamura Y, Miyajima N, Hirosawa M, Sugiura M, Sasamoto S, Kimura T, Hosouchi T, Matsuno A, Muraki A, Nakazaki N, Naruo K, Okumura S, Shimpo S, Takeuchi C, Wada T, Watanabe A, Yamada M, Yasuda M, Tabata S
(1996)
DNA Res
3: 109-116
[Abstract], 185-209
-
Kanevski I, Maliga P
(1994)
Proc Natl Acad Sci USA
91: 1969-1973
[Abstract/Free Full Text]
-
Lemieux C, Otis C, Turmel M
(2000)
Nature
403: 649-652
[CrossRef][Medline]
-
Lockhart PJ, Steel MA, Hendy MD, Penny D
(1994)
Mol Biol Evol
11: 605-612
[ISI]
-
Ma Y, Kouranov A, LaSalsa SE, Schnell DJ
(1996)
J Cell Biol
134: 315-327
[Abstract/Free Full Text]
-
Martin W, Stoebe B, Goremykin V, Hansmann S, Hasegawa M, Kowallik K
(1998)
Nature
393: 162-165
[CrossRef][Medline]
-
McFadden GI, Reith M, Munholland J, Lang-Unnasch N
(1996)
Nature
381: 482
[CrossRef][Medline]
-
Mishkind M, Wessler S, Schmidt G
(1985)
J Cell Biol
100: 226-234
[Abstract/Free Full Text]
-
Ohyama K, Fukuzawa H, Kohchi T, Shirai H, Sano T, Sano S, Umesono K, Shiki Y, Takeuchi M, Chang Z, Aota S, Inokuchi H, Ozeki H
(1986)
Nature
322: 572-574
[CrossRef][ISI]
-
Osteryoung KW, Vierling E
(1995)
Nature
376: 473-474
[Medline]
-
Reith M, Munholland J
(1995)
Plant Mol Biol Rep
13: 333-335
-
Schmidt G, Devillers-Thiery A, Desruisseaux H, Blobel G, Chua N-H
(1979)
J Cell Biol
83: 615-622
[Abstract/Free Full Text]
-
Schopf JW
(1993)
Science
260: 640-646
[Abstract/Free Full Text]
-
Schreier PH, Sefotr EA, Schell J, Bohnert HJ
(1985)
EMBO J
4: 25-32
[ISI][Medline]
-
Schwarz Z, Kössel H
(1980)
Nature
283: 739-742
[CrossRef]
-
Smeekens S, Bauerie C, Hageman J, Keegstra K, Weisbeek P
(1986)
Cell
46: 365-375
[CrossRef][ISI][Medline]
-
Sveshnikova N, Soll J, Schleiff E
(2000)
Proc Natl Acad Sci U.S.A
97: 4973-4978
[Abstract/Free Full Text]
-
van den Broek G, Timko M, Kausch A, Cahmore A, Montagu M, Herrera-Estrella L
(1985)
Nature
313: 358-363
[CrossRef][Medline]
-
Weeden N
(1981)
J Mol Evol
17: 133-139
[CrossRef][ISI][Medline]
-
Wilson RJM, Denny PW, Preiser PR, Rangachari K, Roberts K, Roy A, Whyte A, Strath M, Moore DJ, Moore PW, Williamson DH
(1996)
J Mol Biol
261: 155-172
[CrossRef][ISI][Medline]
-
Zhang Z, Green B, Cavalier-Smith T
(1999)
Nature
400: 155-159
[CrossRef][Medline]
© 2001 American Society of Plant Physiologists
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TOP
INTRODUCTION
ENDOSYMBIOSIS TRIUMPHS