Yeast 5-aminolevulinate synthase provides additional chlorophyll precursor in transgenic tobacco

Synthesis of the tetrapyrrole precursor 5-aminolevulinate (ALA) in plants starts with glutamate and is a tRNA-dependent pathway consisting of three enzymatic steps localized in plastids. In animals and yeast, ALA is formed in a single step from succinyl CoA and glycine by aminolevulinate synthase (ALA-S) in mitochondria. A gene encoding a fusion protein of yeast ALA-S with an amino-terminal transit sequence for the small subunit of ribulose bisphosphate carboxylase was introduced into the genome of wild-type tobacco and a chlorophyll-deficient transgenic line expressing glutamate 1-semi-aldehyde aminotransferase (GSA-AT) antisense RNA. Expression of ALA-S in the GSA-AT antisense transgenic line provided green-pigmented co-transformants similar to wild-type in chlorophyll content, while transformants derived from wild-type plants did not show phenotypical changes. The capacity to synthesize ALA and chlorophyll was increased in transformed plants, indicating a contribution of ALA-S to the ALA supply for chlorophyll synthesis. ALA-S activity was detected in plastids of the transformants. Preliminary evidence is presented that succinyl CoA, the substrate for ALA-S, can be synthesized and metabolized in plastids. The transgenic plants formed chlorophyll in the presence of gabaculine, an inhibitor of GSA-AT. Steady-state RNA and protein levels and, consequently, the enzyme activity of GSA-AT were reduced in plants expressing ALA-S. In analogy to the light-dependent ALA synthesis attributed to feedback regulation, a mechanism at the level of intermediates or tetrapyrrole end-products is proposed, which co-ordinates the need for heme and chlorophyll precursors and restricts synthesis of ALA by regulating GSA-AT gene expression. The genetically engineered tobacco plants containing the yeast ALA-S activity demonstrate functional complementation of the catalytic activity of the plant ALA-synthesizing pathway and open strategies for producing tolerance against inhibitors of the C5 pathway.     

Assembly of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii requires expression of the TLA2-CpFTSY gene

The truncated light-harvesting antenna2 (tla2) mutant of Chlamydomonas reinhardtii showed a lighter-green phenotype, had a lower chlorophyll (Chl) per-cell content, and higher Chl a/b ratio than corresponding wild-type strains. Physiological analyses revealed a higher intensity for the saturation of photosynthesis and greater P(max) values in the tla2 mutant than in the wild type. Biochemical analyses showed that the tla2 strain was deficient in the Chl a-b light-harvesting complex, and had a Chl antenna size of the photosystems that was only about 65% of that in the wild type. Molecular and genetic analyses showed a single plasmid insertion in the tla2 strain, causing a chromosomal DNA rearrangement and deletion/disruption of five nuclear genes. The TLA2 gene, causing the tla2 phenotype, was cloned by mapping the insertion site and upon complementation with each of the genes that were deleted. Successful complementation was achieved with the C. reinhardtii TLA2-CpFTSY gene, whose occurrence and function in green microalgae has not hitherto been investigated. Functional analysis showed that the nuclear-encoded and chloroplast-localized CrCpFTSY protein specifically operates in the assembly of the peripheral components of the Chl a-b light-harvesting antenna. In higher plants, a cpftsy null mutation inhibits assembly of both the light-harvesting complex and photosystem complexes, thus resulting in a seedling-lethal phenotype. The work shows that cpftsy deletion in green algae, but not in higher plants, can be employed to generate tla mutants. The latter exhibit improved solar energy conversion efficiency and photosynthetic productivity under mass culture and bright sunlight conditions.     

Decreased and increased expression of the subunit CHL I diminishes Mg chelatase activity and reduces chlorophyll synthesis in transgenic tobacco plants

The chelation of Fe2+ and Mg2+ ions forms protoheme IX and Mg-protoporphyrin IX, respectively, and the latter is an intermediate in chlorophyll synthesis. Active magnesium protoporphyrin IX chelatase (Mg-chelatase) is an enzyme complex consisting of three different subunits. To investigate the function of the CHL I subunit of Mg-chelatase and the effects of modified Mg-chelatase activity on the tetrapyrrole biosynthetic pathway, we characterized N. tabacum transformants carrying gene constructs with the Chl I cDNA sequence in antisense and sense orientation under the control of the CaMV 35S promoter. Both elevated and diminished levels of Chl I mRNA and Chl I protein led to reduced Mg-chelatase activities, reflecting a perturbation of the assembly of the enzyme complex. The transformed plants did not accumulate the substrate of Mg-chelatase, protoporphyrin IX, but the leaves contained less chlorophyll and possessed increased chlorophyll a/b ratios, as well as a deficiency of light-harvesting chlorophyll binding proteins of photosystems I and II. The expression and activity of several tetrapyrrolic enzymes were reduced in parallel to lower the Mg-chelatase activity. Consistent with the lower chlorophyll contents, the rate-limiting synthesis of 5-aminolevulinate was also decreased in the transgenic lines analyzed. The consequence of reduced Mg-chelatase on early and late steps of chlorophyll synthesis, and on the organization of light harvesting complexes is discussed.     

Structural and Functional Characteristics of the Mutant Plastids from Extranuclear Variegated Forms of Sunflower

The electron-microscopic analysis of extranuclear variegated forms of sunflower demonstrated that plastids from white leaf areas are practically devoid of inner membrane structures. The mutants and the initial (wild-type) green line were compared by the contents of chlorophyll (Chl), carotenoids, and 70S ribosomes and by the activity of Rubisco. A mutant Var10 line was used to demonstrate that the primary characteristic manifestations of Chl deficiency include the synchronous retardation of the synthesis of a specific Chl precursor, 5-aminolevulinate, a decrease in the Chl a/b ratio to the level below that in the wild type, and the reduction of photosystem (PS) I Chl luminance. The progress of photodestructive processes in the mutant aggravated the listed disturbances; as a result, PSII complexes and light-harvesting complexes gradually degraded. The manifestation of the trait of Chl-deficiency notably varied depending on plant growth conditions (primarily, temperature and illumination regimes). The dependence of this manifestation on light and temperature was observed only at the early stage of development of pigment-containing tissues. When plants terminated this stage under low irradiation, they did not become Chl-deficient, and when their leaves were subsequently transferred to the conditions promoting the maximum pigment anomaly, the destructive processes did not advance in the mutant tissue. The authors assume that the plastom mutations under study impair the control over the expression of the structural plastogenes that determine the synthesis of pigment and protein components of the photosynthetic apparatus, rather than directly damage to the primary structures of plastogenes.     

Overexpression of chlorophyllide a oxygenase (CAO) enlarges the antenna size of photosystem II in Arabidopsis thaliana

The light-harvesting efficiency of a photosystem is thought to be largely dependent on its photosynthetic antenna size. It has been suggested that antenna size is controlled by the biosynthesis of chlorophyll b. To verify this hypothesis, we overexpressed the enzyme for chlorophyll b biosynthesis, chlorophyllide a oxygenase (CAO), in Arabidopsis thaliana by transforming the plant with cDNA for CAO under the control of the 35S cauliflower mosaic virus promoter. In the early de-etiolation phase, when the intrinsic CAO expression is very low, the chlorophyll a: b ratio was drastically decreased from 28 to 7.3, indicating that enhancement of chlorophyll b biosynthesis had been successfully achieved. We made the following observations in full-green rosette leaves of transgenic plants. (1) The chlorophyll a : b ratio was reduced from 2.85 to 2.65. (2) The ratio of the peripheral light-harvesting complexes (LHCII) to the core antenna complex (CPa) resolved with the green-gel system increased by 20%. (3) The ratio of the light-harvesting complex II apoproteins (LHCP) to 47-kDa chlorophyll a protein (CP47), which was estimated by the results of immunoblotting, increased by 40%. These results indicated that the antenna size increased by at least 10-20% in transgenic plants, suggesting that chlorophyll b biosynthesis controls antenna size. To the best of our knowledge, this is the first report on enlargement of the antenna size by genetic manipulations.     

A red-shifted antenna protein associated with photosystem II in Physcomitrella patens

Antenna systems of plants and green algae are made up of pigment-protein complexes belonging to the light-harvesting complex (LHC) multigene family. LHCs increase the light-harvesting cross-section of photosystems I and II and catalyze photoprotective reactions that prevent light-induced damage in an oxygenic environment. The genome of the moss Physcomitrella patens contains two genes encoding LHCb9, a new antenna protein that bears an overall sequence similarity to photosystem II antenna proteins but carries a specific motif typical of photosystem I antenna proteins. This consists of the presence of an asparagine residue as a ligand for Chl 603 (A5) chromophore rather than a histidine, the common ligand in all other LHCbs. Asparagine as a Chl 603 (A5) ligand generates red-shifted spectral forms associated with photosystem I rather than with photosystem II, suggesting that in P. patens, the energy landscape of photosystem II might be different with respect to that of most green algae and plants. In this work, we show that the in vitro refolded LHCb9-pigment complexes carry a red-shifted fluorescence emission peak, different from all other known photosystem II antenna proteins. By using a specific antibody, we localized LHCb9 within PSII supercomplexes in the thylakoid membranes. This is the first report of red-shifted spectral forms in a PSII antenna system, suggesting that this biophysical feature might have a special role either in optimization of light use efficiency or in photoprotection in the specific environmental conditions experienced by this moss.     

Light harvesting in photosystem I supercomplexes

In photosynthetic membranes of cyanobacteria, algae, and higher plants, photosystem I (PSI) mediates light-driven transmembrane electron transfer from plastocyanin or cytochrome c6 to the ferredoxin-NADP complex. The oxidoreductase function of PSI is sensitized by a reversible photooxidation of primary electron donor P700, which launches a multistep electron transfer via a series of redox cofactors of the reaction center (RC). The excitation energy for the functioning of the primary electron donor in the RC is delivered via the chlorophyll core antenna in the complex with peripheral light-harvesting antennas. Supermolecular complexes of the PSI acquire remarkably different structural forms of the peripheral light-harvesting antenna complexes, including distinct pigment types and organizational principles. The PSI core antenna, being the main functional unit of the supercomplexes, provides an increased functional connectivity in the chlorophyll antenna network due to dense pigment packing resulting in a fast spread of the excitation among the neighbors. Functional connectivity within the network as well as the spectral overlap of antenna pigments allows equilibration of the excitation energy in the depth of the whole membrane within picoseconds and loss-free delivery of the excitation to primary donor P700 within 20-40 ps. Low-light-adapted cyanobacteria under iron-deficiency conditions extend this capacity via assembly of efficiently energy coupled rings of CP43-like complexes around the PSI trimers. In green algae and higher plants, less efficient energy coupling in the eukaryotic PSI-LHCI supercomplexes is probably a result of the structural adaptation of the Chl a/b binding LHCI peripheral antenna that not only extends the absorption cross section of the PSI core but participates in regulation of excitation flows between the two photosystems as well as in photoprotection.     

A small chloroplast-encoded protein as a novel architectural component of the light-harvesting antenna

A small conserved open reading frame in the plastid genome, ycf9, encodes a putative membrane protein of 62 amino acids. To determine the function of this reading frame we have constructed a knockout allele for targeted disruption of ycf9. This allele was introduced into the tobacco plastid genome by biolistic transformation to replace the wild-type ycf9 allele. Homoplasmic ycf9 knockout plants displayed no phenotype under normal growth conditions. However, under low light conditions, their growth rate was significantly reduced as compared with the wild-type, due to a lowered efficiency of the light reaction of photosynthesis. We show that this phenotype is caused by the deficiency in a pigment-protein complex of the light-harvesting antenna of photosystem II and hence by a reduced efficiency of photon capture when light availability is limiting. Our results indicate that, in contrast to the current view, light-harvesting complexes do not only consist of the classical pigment-binding proteins, but may contain small structural subunits in addition. These subunits appear to be crucial architectural factors for the assembly and/or maintenance of stable light-harvesting complexes.     

Photosystem I activity is increased in the absence of the PSI-G subunit.

PSI-G is a subunit of photosystem I in eukaryotes. The function of PSI-G was characterized in Arabidopsis plants transformed with a psaG cDNA in antisense orientation. Several plants with significantly decreased PSI-G protein content were identified. Plants with reduced PSI-G content were indistinguishable from wild type when grown under optimal conditions, despite a 40% reduction of photosystem I. This decrease of photosystem I was correlated with a similar reduction in state transitions. Surprisingly, the reduced photosystem I content was compensated for by a more effective photosystem I because the light-dependent reduction of NADP(+) in vitro was 48% higher. Photosystem I antenna size determined from flash-induced P700 absorption changes did not reveal any significant effect on the size of the photosystem I antenna in the absence of PSI-G, whereas a 17% reduction was seen in the absence of PSI-K. However, nondenaturing green gels revealed that the interaction between photosystem I and the light-harvesting complex I was less stable in the absence of PSI-G. Thus, PSI-G plays a role in stabilizing the binding of the peripheral antenna. The increased activity in the absence of PSI-G suggests that PSI-G could have an important role in regulation of photosystem I.     

Light regulation of photosynthetic membrane structure,organization, and function

The light environment during plant growth determines the structural and functional properties of higher plant chloroplasts, thus revealing a dynamically regulated developmental system. Pisum sativum plants growing under intermittent illumination showed chloroplasts with fully functional photosystem (PS) II and PSI reaction centers that lacked the peripheral chlorophyll (Chi) a/b and Chl a light-harvesting complexes (LHC), respectively. The results suggest a light flux differential threshold regulation in the biosynthesis of the photosystem core and peripheral antenna complexes. Sun-adapted species and plants growing under far-red-depleted illumination showed grana stacks composed of few (3–5) thylakoids connected with long intergrana (stroma) thylakoids. They had a PSII/PSI reaction center ratio in the range 1.3–1.9. Shade-adapted species and plants growing under far-red-enrichcd illumination showed large grana stacks composed of several thylakoids, often extending across the entire chloroplast body, and short intergrana stroma thylakoids. They had a higher PSII/PSI reaction center ratio, in the range of 2.2–4.0. Thus, the relative extent of grana and stroma thylakoid formation corresponds with the relative amounts of PSII and PSI in the chloroplast, respectively. The structural and functional adaptation of the photosynthetic membrane system in response to the quality of illumination involves mainly a control on the rate of PSII and PSI complex biosynthesis.     

Functional analysis of N-terminal domains of Arabidopsis chlorophyllide a oxygenase.

Higher plants acclimate to various light environments by changing the antenna size of a light-harvesting photosystem. The antenna size of a photosystem is partly determined by the amount of chlorophyll b in the light-harvesting complexes. Chlorophyllide a oxygenase (CAO) converts chlorophyll a to chlorophyll b in a two-step oxygenation reaction. In our previous study, we demonstrated that the cellular level of the CAO protein controls accumulation of chlorophyll b. We found that the amino acids sequences of CAO in higher plants consist of three domains (A, B, and C domains). The C domain exhibits a catalytic function, and we demonstrated that the combination of the A and B domains regulates the cellular level of CAO. However, the individual function of each of A and B domain has not been determined yet. Therefore, in the present study we constructed a series of deleted CAO sequences that were fused with green fluorescent protein and overexpressed in a chlorophyll b-less mutant of Arabidopsis thaliana, ch1-1, to further dissect functions of A and B domains. Subsequent comparative analyses of the transgenic plants overexpressing B domain containing proteins and those lacking the B domain determined that there was no significant difference in CAO protein levels. These results indicate that the B domain is not involved in the regulation of the CAO protein levels. Taken together, we concluded that the A domain alone is involved in the regulatory mechanism of the CAO protein levels.     

Association of chlorophyll a/c(2) complexes to photosystem I and photosystem II in the cryptophyte Rhodomonas CS24

Photosynthetic supercomplexes from the cryptophyte Rhodomonas CS24 were isolated by a short detergent treatment of membranes from the cryptophyte Rhodomonas CS24 and studied by electron microscopy and low-temperature absorption and fluorescence spectroscopy. At least three different types of supercomplexes of photosystem I (PSI) monomers and peripheral Chl a/c(2) proteins were found. The most common complexes have Chl a/c(2) complexes at both sides of the PSI core monomer and have dimensions of about 17x24 nm. The peripheral antenna in these supercomplexes shows no obvious similarities in size and/or shape with that of the PSI-LHCI supercomplexes from the green plant Arabidopsis thaliana and the green alga Chlamydomonas reinhardtii, and may be comprised of about 6-8 monomers of Chl a/c(2) light-harvesting complexes. In addition, two different types of supercomplexes of photosystem II (PSII) dimers and peripheral Chl a/c(2) proteins were found. The detected complexes consist of a PSII core dimer and three or four monomeric Chl a/c(2) proteins on one side of the PSII core at positions that in the largest complex are similar to those of Lhcb5, a monomer of the S-trimer of LHCII, Lhcb4 and Lhcb6 in green plants.     

Kinetics and spectra of photo-induced changes in the absorption of pigment-protein complexes of photosystem 1 in a picosecond range

The energy transfer from the light-harvesting antenna chlorophylls to the reaction center molecules and subsequent charge separation were investigated using a difference picosecond spectrophotometer with selective excitation. The objects were the pigment-protein complexes of photosystem 1 (Chl/P700 = 60) isolated from bean leaves. The difference absorption spectra of the excited states of light-harvesting antenna chlorophylls and the P700 photooxidation were measured. It was shown that the excited states of antenna chlorophylls were generated within 10 ps and deactivated with three-component kinetics: tau 1 = 20--45 ps, tau 2 = 100--300 ps, tau 3 greater than 500 ps. The process of the P700 photooxidation induced by the 650 nm exciting pulse was approximately monoexponential with tau equal to 15--30 ps. It is established that the P700 photooxidation is due to the efficient transfer of excitation energy from antenna chlorophylls to reaction centers.     

Kinetic Studies on the Xanthophyll Cycle in Barley Leaves (Influence of Antenna Size and Relations to Nonphotochemical Chlorophyll Fluorescence Quenching)

Xanthophyll-cycle kinetics as well as the relationship between the xanthophyll de-epoxidation state and Stern-Volmer type nonphotochemical chlorophyll (Chl) fluorescence quenching (qN) were investigated in barley (Hordeum vulgare L.) leaves comprising a stepwise reduced antenna system. For this purpose plants of the wild type (WT) and the Chl b-less mutant chlorina 3613 were cultivated under either continuous (CL) or intermittent light (IML). Violaxanthin (V) availability varied from about 70% in the WT up to 97 to 98% in the mutant and IML-grown plants. In CL-grown mutant leaves, de-epoxidation rates were strongly accelerated compared to the WT. This is ascribed to a different accessibility of V to the de-epoxidase due to the existence of two V pools: one bound to light-harvesting Chl a/b-binding complexes (LHC) and the other one not bound. Epoxidation rates (k) were decreased with reduction in LHC protein contents: kWT > kmutant >> kIML plants. This supports the idea that the epoxidase activity resides on certain LHC proteins. Irrespective of huge zeaxanthin and antheraxanthin accumulation, the capacity to develop qN was reduced stepwise with antenna size. The qN level obtained in dithiothreitol-treated CL- and IML-grown plants was almost identical with that in untreated IML-grown plants. The findings provide evidence that structural changes within the LHC proteins, mediated by xanthophyll-cycle operation, render the basis for the development of a major proportion of qN.     

Loss of alpha-tocopherol in tobacco plants with decreased geranylgeranyl reductase activity does not modify photosynthesis in optimal growth conditions but increases sensitivity to high-light stress

The enzyme geranylgeranyl reductase (CHL P) catalyses the reduction of geranylgeranyl diphosphate to phytyl diphosphate in higher-plant chloroplasts and provides phytol for both chlorophyll (Chl) and tocopherol synthesis. The reduction in CHL P activity in transgenic tobacco (Nicotiana tabacum L.) plants is accompanied by the reduction in total Chl and tocopherol content and the accumulation of geranylgeranylated Chl (ChlGG). The photosynthetic performance and the susceptibility to photo-oxidative stress have been investigated in these transgenic plants. The reduced total Chl content in Chl P antisense plants resulted in the reduction of electron transport chains per leaf area without a concomitant effect on the stoichiometry, composition and activity of both photosystems. However, Chl P antisense plants were much more sensitive to light stress. Analyses of Chl fluorescence quenching indicated an increased photoinhibitory quenching at the expense of the pH-dependent fluorescence quenching after short illumination (15 min) at moderate light intensities. Prolonged illumination (up to 1 h) at saturating light intensities induced an increased photoinactivation from which the Chl P antisense plants could not recover or could only partially recover during a subsequent low light phase. Our data imply that the presence of ChlGG has no influence on harvesting and transfer of light energy in either photosystem. However, the reduced tocopherol content of the thylakoid membrane is a limiting factor for defensive reactions to photo-oxidative stress.     

Short- and long-term operation of the lutein-epoxide cycle in light-harvesting antenna complexes

The lutein-5,6-epoxide (Lx) cycle operates in some plants between lutein (L) and its monoepoxide, Lx. Whereas recent studies have established the photoprotective roles of the analogous violaxanthin cycle, physiological functions of the Lx cycle are still unknown. In this article, we investigated the operation of the Lx cycle in light-harvesting antenna complexes (Lhcs) of Inga sapindoides Willd, a tropical tree legume accumulating substantial Lx in shade leaves, to identify the xanthophyll-binding sites involved in short- and long-term responses of the Lx cycle and to analyze the effects on light-harvesting efficiency. In shade leaves, Lx was converted into L upon light exposure, which then replaced Lx in the peripheral V1 site in trimeric Lhcs and the internal L2 site in both monomeric and trimeric Lhcs, leading to xanthophyll composition resembling sun-type Lhcs. Similar to the violaxanthin cycle, the Lx cycle was operating in both photosystems, yet the light-induced Lx --> L conversion was not reversible overnight. Interestingly, the experiments using recombinant Lhcb5 reconstituted with different Lx and/or L levels showed that reconstitution with Lx results in a significantly higher fluorescence yield due to higher energy transfer efficiencies among chlorophyll (Chl) a molecules, as well as from xanthophylls to Chl a. Furthermore, the spectroscopic analyses of photosystem I-LHCI from I. sapindoides revealed prominent red-most Chl forms, having the lowest energy level thus far reported for higher plants, along with reduced energy transfer efficiency from antenna pigments to Chl a. These results are discussed in the context of photoacclimation and shade adaptation.     

Photoinhibitory damage is modulated by the rate of photosynthesis and by the photosystem II light-harvesting chlorophyll antenna size

We investigated the effect of photosynthetic electron transport and of the photosystem II (PSII) chlorophyll (Chl) antenna size on the rate of PSII photoinhibitory damage. To modulate the rate of photosynthesis and the light-harvesting capacity in the unicellular chlorophyte Dunaliella salina Teod., we varied the amount of inorganic carbon in the culture medium. Cells were grown under high irradiance either with a limiting supply of inorganic carbon, provided by an initial concentration of 25 mM NaHCO₃, or with supplemental CO₂ bubbled in the form of 3% CO₂ in air. The NaHCO₃-grown cells displayed slow rates of photosynthesis and had a small PSII light-harvesting Chl antenna size (60 Chl molecules). The half-time of PSII photodamage was 40 min. When switched to supplemental CO₂ conditions, the rate of photodamage was retarded to a t_1/2 = 70 min. Conversely, CO₂-supplemented cells displayed faster rates of photosynthesis and a larger PSII light-harvesting Chl antenna size (500 Chl molecules). They also showed a rate of photodamage with t_1/2 = 40 min. When depleted of CO₂, the rate of photodamage was accelerated (t_1/2  = 20 min). These results indicate that the in-vivo susceptibility to photodamage is modulated by the rate of forward electron transport through PSII. Moreover, a large Chl antenna size enhances the rate of light absorption and photodamage and, therefore, counters the mitigating effect of forward electron transport. We propose that under steady-state photosynthesis, the rate of light absorption (determined by incident light intensity and PS Chl antenna size) and the rate of forward electron transport (determined by CO₂ availability) modulate the oxidation/reduction state of the primary PSII acceptor Q_A, which in turn defines the low/high probability for photodamage in the PSII reaction center. Received: 14 August 1997 / Accepted: 26 September 1997     

Synthesis of chlorophyll <Emphasis Type="Italic">b</Emphasis>: Localization of chlorophyllide <Emphasis Type="Italic">a</Emphasis>oxygenase and discovery of a stable radical in the catalytic subunit

Background  

Assembly of stable light-harvesting complexes (LHCs) in the chloroplast of green algae and plants requires synthesis of chlorophyll (Chl) b, a reaction that involves oxygenation of the 7-methyl group of Chl a to a formyl group. This reaction uses molecular oxygen and is catalyzed by chlorophyllide a oxygenase (CAO). The amino acid sequence of CAO predicts mononuclear iron and Rieske iron-sulfur centers in the protein. The mechanism of synthesis of Chl b and localization of this reaction in the chloroplast are essential steps toward understanding LHC assembly.     

Efficient assembly of photosystem II in Chlamydomonas reinhardtii requires Alb3.1p, a homolog of Arabidopsis ALBINO3

Alb3 homologs Oxa1 and YidC have been shown to be required for the integration of newly synthesized proteins into membranes. Here, we show that although Alb3.1p is not required for integration of the plastid-encoded photosystem II core subunit D1 into the thylakoid membrane of Chlamydomonas reinhardtii, the insertion of D1 into functional photosystem II complexes is retarded in the Alb3.1 deletion mutant ac29. Alb3.1p is associated with D1 upon its insertion into the membrane, indicating that Alb3.1p is essential for the efficient assembly of photosystem II. Furthermore, levels of nucleus-encoded light-harvesting proteins are vastly reduced in ac29; however, the remaining antenna systems are still connected to photosystem II reaction centers. Thus, Alb3.1p has a dual function and is required for the accumulation of both nucleus- and plastid-encoded protein subunits in photosynthetic complexes of C. reinhardtii.