Biochimica et Biophysica Acta 1541 (2001) 91^101

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Review

Biogenesis and origin of thylakoid membranes

Ute C. Vothknecht

a;

*, Peter Westho

Botanisches Institut der Christian-Albrechts-Universitat Kiel, Am Botanischen Garten 1^9, D-24118 Kiel, Germany
Institut fur Entwicklungs- und Molekularbiologie der Panzen, Heinrich-Heine-Universitat Dusseldorf, Universitatsstrasse 1,
D-40225 Dusseldorf, Germany
Received 25 July 2001; accepted 1 August 2001

Abstract
Thylakoids are photosynthetically active membranes found in Cyanobacteria and chloroplasts. It is likely that they
originated in photosynthetic bacteria, probably in close connection to the occurrence of photosystem II and oxygenic
photosynthesis. In higher plants, chloroplasts develop from undifferentiated proplastids. These contain very few internal
membranes and the whole thylakoid membrane system is built when chloroplast differentiation takes place. During cell and
organelle division a constant synthesis of new thylakoid membrane material is required. Also, rapid adaptation to changes in
light conditions and long term adaptation to a number of environmental factors are accomplished by changes in the lipid and
protein content of the thylakoids. Thus regulation of synthesis and assembly of all these elements is required to ensure
optimal function of these membranes. 2001 Elsevier Science B.V. All rights reserved.
Keywords: Chloroplast; Photosynthesis; Plastid development; Organelle evolution; Thylakoid formation

1. Introduction
Evolution of oxygenic photosynthesis with its CO2
xation and oxygen release enabled life on earth as
we experience it today. It is assumed that oxygenic
photosynthesis developed several billion years ago in
an ancestor of today's cyanobacteria most likely
from an already existing anoxygenic photosynthesis
apparatus [1]. The capacity to perform oxygenic photosynthesis was passed on to algae and higher plants
by an endosymbiotic event that turned a cyanobacterium into a cell organelle, the chloroplast. The photosynthetic machinery of both, cyanobacteria and

* Corresponding author. Fax: +49-431-880-4222.

E-mail address: uvothknecht@bot.uni-kiel.de (U.C.
Vothknecht).

chloroplasts, is located on a special internal membrane system, the thylakoids. Thanks to the unique
architecture of this membrane cyanobacteria and
chloroplasts convert solar energy into chemical energy with an eciency signicantly better than any
man-made photovoltaic system. Therefore, the ability of the cell to build and alter this membrane system is essential for ecient oxygenic photosynthesis.
Resulting from the combination of structural, biochemical, and genetic analysis, we have a well
founded knowledge of the ultrastructure and composition of thylakoid membranes, but despite the importance that the thylakoid membrane system has
for photosynthesis and the energy metabolism of
plants and cyanobacteria, the molecular processes
connected to the origin, synthesis, maintenance and
adaptation of the thylakoids remain elusive. In this
review we will discuss recent ndings on thylakoid

0167-4889 / 01 / $ ^ see front matter 2001 Elsevier Science B.V. All rights reserved.
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biogenesis and evolution and their impact on our

understanding. Since most studies concerning the
biogenesis of thylakoids have been performed on
chloroplasts of higher plants and green algae, this
review will focus on these organisms. The last section
will deal with internal membrane systems in bacteria,
especially the thylakoids of cyanobacteria, and the
evolution of the thylakoid membrane in these organisms.
2. Form and function in plastids
In 1848 chloroplasts were rst described by Unger
simply as pigment bound structures. Later in the
19th century Schimper [2,3] characterized these structures, which he called `Chlorophyllkorner', in greater
detail. With only the resolution of the light microscope available, he described plastids as cell components containing chlorophyll or other pigments
which develop from colorless precursors. He already
perceived the existence of various types of plastids
and made the observation that they can pass through
dierent stages during their development. Shortly
after the invention of the electron microscope, the
rst electron micrographs of chloroplasts were published [4], and soon thereafter, this new technique
was used for the rst detailed studies of dierent
plastid forms and their development. In the late 50s
the basic structure of thylakoids had been described
[5^7] and the means of thylakoid biogenesis were
discussed.
Thylakoids are the dominating structure inside
fully mature chloroplasts. The formation and alteration of the thylakoid membrane structure and composition are closely connected to the development of
the chloroplasts from simple, undierentiated proplastids. These are small round shaped organelles
hardly distinguishable from mitochondria (Fig. 1),
that contain very few internal membranes that are
often found as vesicles or small saccular structures
[7^9]. Occasionally these membranes are observed
continuous with the inner envelope.
In the presence of light proplastids develop into
mature chloroplasts. This transition has been intensively studied in grasses. The leaves of these monocotyledonous plants grow with a basal meristem and
hence form a developmental gradient. Cells found at

the base of the leaf are youngest and contain mainly

proplastids while the oldest cells with fully developed
chloroplasts are found close to the tip. Ultra-thin
sections revealed that during the progress of chloroplast maturation the internal membrane system
builds up in consecutive phases. First, long lamella
are formed which are later complemented by smaller,
disc-shaped structures that associate into so-called
grana stacks (Fig. 1). At the same time the typical
lens-shape form of the chloroplasts develops. Finally,
mature plant chloroplasts contain a complex and intertwined internal membrane system which was
named `thylakoids' according to the Greek word
`3eVKUYASNRf' (sack-like) [10]. In fully mature chloroplast no continuation between the inner envelope
and the thylakoids has been observed.
In the absence of light proplastids turn into etioplasts which contain very few internal membranes
but a characteristic prolamellar body [11,12]. The
prolamellar body is a paracrystalline structure consisting of lipids and essentially a single protein, the
NADPH-dependent protochlorophyllide oxidoreductase [13,14]. Shortly after the onset of illumination
the prolamellar body is dispersed and thylakoids begin to form [7,12,15]. Since the start of illumination
can easily be controlled in experimental setups, this
system has often been used to study chloroplast development. Prolamellar bodies are mainly considered
in connection with etioplasts but they are not restricted to them. Already after a short period of
darkness `secondary' prolamellar bodies form inside
fully matured chloroplasts [16,17]. This results in the
coexistence of prolamellar bodies and thylakoids and
raises questions about the function of the prolamellar
body for the mature chloroplasts.
Proplastids can further develop into chromoplasts
or leucoplasts. These are specialized forms of plastids
used for coloration or storage [18]. Chromoplasts are
carotenoid-containing plastids found in many ower
petals, fruits and roots. Coloration of these organs is
often ascribed to chromoplasts and this might even
be their main function. Leucoplasts are characterized
by a lack of coloration and they can be distinguished
by the substance that is stored, i.e. amyloplasts, proteoplasts or elaioplasts. The nal stage of a plastid's
life is the senescent or gerontoplast. These are plastids that have reached a stage of senescence that is
not reversible. All plastids, independent of their sta-

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Fig. 1. Overview of the development of chloroplasts. Chloroplast develop from undierentiated proplastids. During maturation the
complex internal thylakoid membrane network is formed. Proplastids can also develop into other plastids forms, such as etioplasts,
chromoplasts and leucoplasts. Moreover, fully dierentiated plastids retain the ability to develop into each other. Gerontoplasts are a
nal stage in plastid development in which a level of senescence is reached that is irreversible. The electron microscopic pictures of
thin sections show several stages in the development of a chloroplast.

tus, retain the ability to develop into each other (Fig.

1). Interconversion of dierent plastid forms requires
dramatic changes of the ultrastructure, including the
biogenesis, reorganization and regression of internal
membranes.
3. Structure and composition of the
thylakoid membrane
The additional compartment that the thylakoid
network creates in cyanobacterial cells and in chloroplasts is an important feature that distinguishes
these from bacteria performing anoxygenic photosynthesis. In these latter organisms, the internal

membranes are invaginations still continuous with

the plasma membrane [19,20]. In mature chloroplasts
and in cyanobacteria it is assumed that the thylakoids are no longer connected to the inner envelope
or the plasma membrane, respectively, because no
such continuum can be observed in electron microscopic pictures.
How is this unique structural composition
achieved? In cyanobacteria and many algae, thylakoids consist mainly of single layers formed by
long lamellae. The structure of the thylakoid membrane in a fully mature chloroplast is more complex
(Fig. 1). Initiated by earlier electron microscopic
studies a model for the thylakoid structure as a
huge intertwined network of stroma lamellae con-

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necting grana stacks was proposed which with little

alterations is still valid today [21,22]. One can distinguish two major parts, the grana and the stroma
lamellae. Grana are short, disc-shaped lamellae
closely packed to form stacks. These stacks are interconnected by stroma lamellae which also form
prolonged extensions into the stroma. Thus, the arrangement of the thylakoid membrane system creates
a single huge compartment inside the chloroplast, the
thylakoid lumen. Additionally to creating a single
internal space this structure builds a membrane surface that is much larger than a simple invagination of
the inner envelope would generate.
To understand the complexity of the task that the
formation of thylakoids presents to the cell, it is
important to gure the components that are required
to build up this special photosynthetic membrane.
Thylakoids are lipid bilayers with a unique glycerolipid composition dierent from other cell membranes. Thylakoid lipids have a high content, about
70^80%, of galactosyl diglycerides and both monogalactosyl diacylglycerol and digalactosyl diacylglycerol are lipids nearly exclusively found in plastidal
membranes [23]. Notably, these galactolipids contain
two highly unsaturated fatty acyl chains instead of
one as is common in membrane lipids and are both
non-bilayer forming lipids. Additionally the thylakoids contain phosphatidylglycerol and sulfoquinovosyl diacylglycerol together with other minor components [23]. All these lipids are not evenly
distributed along the thylakoid membrane. Instead
the lipid distribution diers between the leaet that
is exposed to the stroma and the inner leaet that
faces the thylakoid lumen [23]. It is not clear how
this asymmetrical arrangement of the lipid distribution is achieved. Yet it has to be assumed that it is
important for the function of the thylakoid membrane.
The dominant protein complexes of the thylakoids
are photosystems I [24] and II [25] and their associated light harvesting antenna, the cytochrome b6 f
complex [26] and the proton-translocating ATP synthase [27]. These complexes comprise not only many
peripheral and integral proteins but also a variety of
pigments and co-factors [28]. Their assembly is,
therefore, a complex process and requires a larger
number of auxiliary and regulatory factors [28,29].
These factors are involved in the membrane integra-

tion, modication and later degradation of the proteinaceous components and are also required for the
addition of the pigments and co-factors. To complicate matters, certain components, like the two photosystems, are unevenly distributed in the thylakoid
membrane network. While photosystem I is most
abundant in the non-stacked stroma lamellae, photosystem II is the dominating component of the grana
stacks [30]. Thus thylakoid biogenesis and maintenance have to assure not only the arrangement of a
functional but at the same time asymmetric architecture of both the lipid and the protein components of
this membrane.
4. Thylakoid membrane formation
One of the most elusive aspects of thylakoid formation is the exact mechanism by which the membrane itself is formed. In young, not yet dierentiated plastids a continuum can sometimes be observed
between the inner envelope and the developing internal membrane structures [7^9]. Thus the synthesis of
early thylakoid membranes might be achieved by invagination of the inner envelope. Even in fully mature chloroplasts the thylakoid membrane is a very
dynamic system. Short-term adaptation to changing
light conditions is obtained by movement of proteins,
especially the light harvesting complex, within the
thylakoid membrane. Long-term adaptation on the
other hand is achieved by a change in the protein
and lipid content of the thylakoids. Although in mature chloroplasts a continuum between the inner envelope and the thylakoids can no longer be observed,
the membrane material required for synthesis and
maintenance of the thylakoids originates from the
chloroplast's inner envelope [7,31,32] and not from
de novo synthesis on already existing thylakoids.
How these lipids are transferred from the inner
envelope to the thylakoids is controversially discussed. One possibility would be the transfer by
vesicles which is a common phenomenon in the cytosol, where vesicle trac is involved in many dierent cellular processes including the secretory pathway, endocytosis, neural transmission and vacuole
formation [33]. A similar vesicle transfer from
the inner envelope to the thylakoids has been implicated in the synthesis of thylakoid membranes

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[7,31,32,34,35]. Vesicles inside plastids have been observed in early electron microscopic studies [7^9].
They are common in proplastids and have also
been observed on the inner envelope of etioplasts
in dark-grown cells of the Chlamydomonas y-1 mutant, shortly after illumination when chloroplast development sets in [36]. On the other hand vesicles are
very rarely detected in mature chloroplasts. They do
accumulate in the stromal space between the inner
envelope and the thylakoids after a low temperature
incubation of leaf tissue [34,35]. A similar phenomenon is described for vesicle transport in animal cells,
i.e. enodplasmic reticulum to Golgi and Golgi to
plasma membrane, where low temperature blocks
the fusion of vesicles with their target membrane
[37]. Further indication for vesicle transfer in chloroplasts comes from mutant analysis. In several plant
mutants that are aected in thylakoid biogenesis, an
accumulation of vesicles can be observed. Others,
like the vipp1 mutant of Arabidopsis, no longer exhibited low temperature vesicle accumulation [35].
The possibility of vesicle transfer inside the chloroplast raised the additional question whether solely
membrane lipids would be transported by these
vesicles. As in vacuole formation the vesicle transport in chloroplasts could be limited to the supply
of thylakoid lipids that are either synthesized at the
inner envelope, i.e. galactolipids, or imported from
the cytosol. It is also possible that non-lipid components of the thylakoid membrane might be transported by means of vesicle trac [38,39]. Several of
the non-lipid components required for the biogenesis
and maintenance of thylakoids are synthesized on
the envelope, i.e. carotenoids, or in the cytosol
[40,41]. Especially hydrophobic components would
require a system to travel through the aqueous stroma.
During chloroplast maturation an extensive formation of thylakoid membranes occurs in concert
with the accumulation of the photosynthetic complexes. Several of the proteinaceous components are
nuclear encoded and post-translationally imported
into the chloroplasts. It was suggested that in Chlamydomonas the nuclear encoded light harvesting
complex proteins are inserted into newly developing
membranes at the inner envelope immediately upon
their entrance in the organelle [42]. Later on, the
development of the thylakoid system continues with

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the formation of grana stacking. Again, integration

of the light harvesting complex into the thylakoid
membrane might play an important role in this structural reconstruction [43]. This early speculation was
supported recently by Simidjiev and coworkers, who
showed that delipidated light harvesting complexes
would restructure into ordered lamellae by the addition of monogalactosyl diacylglycerol [44]. They concluded that the light harvesting complex together
with monogalactosyl diacylglycerol is responsible
for lamellae organization of the thylakoid membrane. Therefore interaction between thylakoid proteins and thylakoid lipids seems important for the
formation of the lipid bilayer in a membrane whose
main components are non-bilayer forming lipids.
5. Regulation of thylakoid biogenesis
How is the formation of the thylakoid lipid bilayer
coordinated with the expression of proteins and the
biosynthesis of pigments and co-factors? It became
obvious quite early after the identication of DNA
and genome structure that plastid development and
thylakoid formation is controlled by both the genome of the cell (nucleome) and the organelle (plastome). Plastids contain up to several hundred copies
of a circular chromosome with a size between 120
and 220 kb. Encoded on the plastome is an average
of about 100^200 proteins in addition to a full set of
ribosomal and transfer RNAs [45,46]. Chloroplasts
are, however, estimated to house about 2000^5000
dierent proteins; consequently only 5^10% of the
plastidal proteins are encoded within the plastome
[46,47] and the majority of proteins required for plastid development and function are encoded in the nucleus. These nuclear encoded proteins are translated
on cytoplasmic ribosomes and have to be post-translationally transported to the chloroplast ([48]; Jarvis
and Soll, this issue).
Many protein complexes and biosynthetic pathways of the chloroplast contain components encoded
both in the nucleome and in the plastome and virtually all chloroplast functions require the concerted
action of nuclear and plastidal encoded factors (Fig.
2). Complex regulatory processes are required to ensure that gene expression of proteins encoded in the
nucleome is properly coordinated with the expression

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Fig. 2. Schematic display of nucleus^chloroplast interaction. Synthesis of plastid encoded proteins is regulated by nuclear encoded factors from the point of gene expression and translation until the nal incorporation into the thylakoid membrane. At the same time
the chloroplast signals the nucleus about its state of development. This signal inuences the expression of nuclear encoded genes. This
gure is based on a scheme presented by Rochaix [102].

of plastome encoded proteins. At the same time the

coordinated development of all plastids in one cell
has to be guaranteed.
Thylakoids become photochemically competent
very early in their development [49,50]. The level of
transcription, which is quite low in proplastids, increases drastically when the chloroplast begins to
mature [51]. At the same time the translational apparatus inside the plastids is built up [52]. It is believed that the nucleus has the control over the onset
of chloroplast dierentiation and also takes the leading part in further developmental stages. To execute
this control most regulatory components have been
transferred to the nucleus [53]. At the same time the
plastids signal back their development stage and condition to the nucleus. These signals, often called the
`plastidal factor', inuence the expression of nuclear
encoded plastid proteins [54^56]. The biogenesis and
function of the chloroplast are therefore an integral
part of the plant cell and the development of the cell
and its organelle are interdependent [57,58]. This is
supported by the fact that plastids cannot easily be
exchanged into a dierent cell background [59^61].
This interdependence of the cell and its organelle is
further strengthened by the fact that two dierent
RNA polymerases are required to transcribe plasti-

dal genes [62,63]. This includes a phage-type RNA

polymerase of nuclear origin [64] and an eubacterial,
multisubunit enzyme whose core subunits are encoded by the plastome [63] while its sigma factor
subunits are encoded by nuclear genes [65]. The nuclear encoded RNA polymerase is primarily responsible for transcription of so-called `housekeeping'
genes of the chloroplasts, while the bacterial-type
enzyme preferentially transcribes genes encoding
components of the photosynthetic machinery [66].
Very little is known so far about the regulation of
plastidal import in relation to plastid development.
Most studies on the regulation of plastidal import
have been done on fully mature, photosynthetically
active chloroplasts (Jarvis and Soll, this issue). A
recent publication indicates a direct inuence of assembly of the light harvesting complex on the import
of the chlorophyll binding protein into the chloroplast [42]. A similar regulation could be envisioned
for other nuclear encoded chloroplast proteins since
also in mature chloroplasts the thylakoid composition is very dynamic and undergoes constant changes
in order to adapt to changing environmental conditions. The ability for adaptation is specially important since plants are not mobile and can therefore
not escape unfavorable conditions. Only a constant

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communication between the organelle and the nucleus can ensure a coordinated supply of all the different factors required.
6. Analysis of thylakoid biogenesis through mutants
Mutants are a powerful tool to study the involvement of gene products on specic processes. Many
dierent mutants that display deciencies in plastid
development and thylakoid formation exist in a wide
range of species. Many of these mutants are randomly occurring natural variations, others are manmade. In early work, new mutants were produced by
treatment of plants or algae with radiation or chemical mutagens [43,67]. Later, the necessary genetic
tools became available for random insertional mutagenesis by T-DNA of Agrobacterium tumefaciens
[68^70] or transposable elements. Only a limited set
of these mutants can be discussed within this section.
For a more detailed summary of mutants see
[43,71,72].
There are many dierent types of mutations that
aect both plastid development and thylakoid formation and the eect that a mutation has on either
is often dicult to distinguish. Often these mutants
are blocked in a step of a biosynthetic pathway located inside the chloroplasts. The resulting loss of a
functional component of the plastid then extends its
eect on the macromolecular structures. In other
mutants structural components of the thylakoid
membrane are missing or defective. Mutations can
aect plastids in all stages of thylakoid formation.
In several cases plastids are blocked very early in
development. These mutants include dcl from tomato
[73], dag from Antirrhinum [74], cla1-1 from Arabidopsis thaliana [75] and several albina mutants of
barley [65,76]. Plastids in dcl, dag and cla1-1 seem
to be arrested in the proplastid stage while plastids in
some of the barley albina mutants can reach the size
of mature chloroplasts but remain fully depleted of
internal membrane structures except for vesicles that
accumulate in some of them.
A similar phenotype can be observed in vrpoA, B
and C1 mutants that lack the bacterial-type RNA
polymerase and consequently the ability to transcribe
the photosynthetic genes which encode subunits of
thylakoid protein complexes [62,77]. Other mutants

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can be found that are blocked in later stages of plastid development, anywhere from the proplastid to
mature chloroplasts. Because of the close connection
between plastid development and thylakoid formation it is often dicult to distinguish pleiotropic effects of these mutations. In some cases mutants seem
to suer from a secondary destruction of the internal
membrane structure rather than a defect in thylakoid
synthesis [78,79].
Defects in thylakoid formation are often caused by
mutations that result in a depletion of major proteinaceous components of the thylakoid membrane, e.g.
major components of the photosystems. For instance, the hcf136 mutant of A. thaliana cannot assemble a functional photosystem II, and this defect is
associated with a drastically disturbed thylakoid
membrane system [80]. Mutations of the protein import apparatus of chloroplasts cause similar defects
in thylakoid formation [81]. Other mutations that
have a great impact on thylakoid formation are mutations that aect the import pathways by which
proteins are inserted into the thylakoid membrane
[82^84]. Examples for such mutants can be found
in maize in the form of tha1 and tha5 which inhibit
the SecA-type import pathway and hcf106 and tha4
where the vph or Tat pathway is disrupted [85,86].
Not surprisingly, mutants that aect the synthesis of
important thylakoid lipids display alterations in the
chloroplast ultrastructure. Arabidopsis dgd1 and
mdg1 mutants lack the enzymes monogalactosyl diacylglycerol synthase or digalactosyl diacylglycerol
synthase that are required for the formation of the
two major thylakoid membrane lipids. These mutants show a wide range of alterations including
changes in the chloroplast ultrastructure and protein
composition [87^89].
Also very common is the connection between deciencies in thylakoid formation and disruption of
pigment biosynthesis [43,67,90]. While pleiotropic effects of these mutations cannot be excluded in some
cases, many investigations have supported the potential inuence of chlorophyll production on chloroplast development [43,90^93]. This connection is especially interesting in light of the `plastidal factor'
that is discussed as a signal from the chloroplasts
to the nucleus (Fig. 2). As described above, the `plastidal factor' is thought to signal the developmental
stage of the plastid to the nucleus and aect the

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expression of many dierent nuclear encoded genes

[55,56]. So far the nature of this plastidal factor remained elusive and indication for the existence of
more than one signaling pathways exists. A recent
paper by Chory and coworkers on the Arabidopsis
mutant uncoupled 5, together with earlier studies by
other groups, provides evidence that one of these
factors might have been found ([94] and references
therein). Their ndings indicate that a subunit of
Mg-chelatase, the enzyme that converts protoporphyrin IX into Mg-protoporphyrin, has an additional, distinct function in the plastid^nucleus signaling
pathway.
Especially interesting are mutants that aect thylakoid formation in otherwise fully developed chloroplasts. One recent example is the vipp1 mutants of
Arabidopsis and Synechocystis. Mutant analysis
showed that the gene product of vipp1 is involved
in the biogenesis of thylakoids in Arabidopsis and
cyanobacteria [35,95]. Interruption of the vipp1
gene locus results in a complete loss of thylakoid
membranes. It seems that Vipp1 is directly involved
in the process of thylakoid biogenesis. Even more,
phylogenetic analysis indicated that the presence of
this protein might be a prerequisite to the ability of
cyanobacteria and chloroplasts to form internal
membranes. Interestingly, the Arabidopsis vipp1 mutant additionally lost the ability for vesicle formation. A vesicle transport system might thus be important for thylakoid formation in mature chloroplasts.
7. Evolution of the thylakoid membrane system
Cyanobacteria are the only phototrophic prokaryotes that carry out oxygenic photosynthesis with two
photosystems. They very much resemble chloroplasts
and it is assumed that at the time of the endosymbiotic event they had already invented oxygenic photosynthesis and developed most of the photosynthetic features found in chloroplasts today. Like
chloroplasts, most cyanobacteria contain an internal
membrane system in which the photosynthetic apparatus is located. Extensive stacking of grana lamellae
is not found in these organisms. Their thylakoids are
organized in layers often paralleling the contour of
the cells. Algae are probably the organism most similar to the early endosymbiotic cells. Similar to cya-

nobacteria, most algae do not contain grana stacks.

Chloroplasts of red algae contain a simple thylakoid
structure similar to cyanobacteria. In green and
brown algae regions of closely appressed thylakoid
membranes occur similar to grana stacks in chloroplasts of higher plants [15]. Also many algae contain
only a single chloroplast per cell. These structural
similarities t well with an evolutionary position between the cyanobacterial endosymbiont and higher
plants.
It is still a point of debate where photosynthesis
developed in the rst place. Recent results favor an
origin of photosynthesis in anoxygenic bacteria [1].
Phototropic green and purple bacteria carry out anoxygenic photosynthesis with a single photosystem
strongly resembling photosystem I. In green-sulfur
bacteria the photosynthetic machinery is located in
the cytoplasmic membrane and the antenna complexes reside in a special non-membranous structure,
the chlorosomes, closely attached to the cytoplasmic
membrane [96]. Purple bacteria on the other hand
often display strong invagination of the cytoplasmic
membrane and their photosystems are concentrated
in these intracytoplasmic membrane regions [97,98].
It is believed that these membranes are not fully
separated from the cytoplasmic membrane and still
form a continuum with the latter [20,21]. It is therefore tempting to speculate that the development of
oxygenic photosynthesis is connected to two dierent events: the invention of the second photosystem
and the biogenesis of an internal membrane system
disconnected from the cell membrane. Support for
this speculation arose from the identication of
Vipp1, a protein essential for thylakoid formation
in higher plant chloroplasts and cyanobacteria
[35,95]. Phylogenetic analysis showed that Vipp1
can be found in organisms that carry out oxygenic
photosynthesis, i.e. plants, algae and cyanobacteria.
No Vipp1 homologue has been found so far in bacteria including those that are capable of anoxygenic
photosynthesis, such as Rhodobacter or Chlorobium.
Vipp1 shares sequence homology with a subunit of
the bacterial phage shock, pspA, and might have
originated from a gene duplication of the latter in
an ancestor of cyanobacteria. It subsequently obtained an additional C-terminal domain that seems
essential for its function in thylakoid formation. In
Arabidopsis the vipp1 mutation also interrupts

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vesicle trac between the inner envelope and the

thylakoids. No such vesicle transport has yet been
shown in any prokaryotic organism including cyanobacteria. Further studies are needed to show
whether vesicle transport is a feature that developed
only in chloroplasts.
At least one cyanobacterium performs oxygenic
photosynthesis without having thylakoids. Gloeobacter violaceus was rst isolated in 1972 from a
limestone rock in Switzerland [99]. Electron microscopic studies revealed the complete lack of internal
membranes. Not even invaginations of the plasma
membrane were observed. Nevertheless, these cells
perform oxygenic photosynthesis [100,101]. The photosystems are located on the plasma membrane and,
similar to purple and green-sulfur bacteria, they form
their proton gradient along the plasma membrane.
This organism might be a cyanobacterium at a stage
before biogenesis of thylakoids was invented or has
resulted from a secondary loss of thylakoid membranes. Compared to cyanobacteria with thylakoids
their photosynthetic capacity is very low. Thus, ecient oxygenic photosynthesis may require the presence of an internal membrane system.
While it is easy to envision the evolution of chloroplasts from a cyanobacterium, it is much more
dicult to understand the evolutionary processes
that created the multiple forms of plastids. There is
no indication that the structures found in proplastids, chromoplasts or leucoplasts have been part of
the genetic plan that the endosymbiont transferred to
the host cell. It must be assumed that this development took place after the endosymbiotic event and
was imposed on the plastid by the host cell. It will be
for future research to elucidate the evolutionary true
origin of the thylakoid membrane and its evolution
from simple single membrane layers to the complex
system present in plant chloroplasts.
Acknowledgements
The authors would like to thank Prof. Dr. J. Soll
for helpful suggestions and discussions. We would
also like to thank S. Westphal and C. Glockman
for the electron microscopic pictures in Fig. 1. Financial support by the `Deutsche Forschungsgemeinschaft' SFB TR1 is acknowledged.

99

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