Minor but key chlorophylls in Photosystem II

A ‘metal-free’ chlorophyll (Chl) a, pheophytin (Phe) a, functions as the primary electron acceptor in PS II. On the basis of Phe a/PS II = 2, Phe a content is postulated as an index for estimation of the stoichiometry of pigments and photosystems. We found Phe a in a Chl d-dominant cyanobacterium Acaryochloris marina, whereas Phe d was absent. The minimum Chl a:Phe a ratio was 2:2, indicating that the primary electron donor is Chl a, accessory is Chl d, and the primary electron acceptor is Phe a in PS II of A. marina. Chl d was artificially formed by the treatment of Chl a with papain in aqueous organic solvents. Further, we will raise a key question on the mechanisms of water oxidation in PS II.     

Pigment protein complexes and the concept of the photosynthetic unit: Chlorophyll complexes and phycobilisomes

The photosynthetic unit includes the reaction centers (RC 1 and RC 2) and the light-harvesting complexes which contribute to evolution of one O₂ molecule. The light-harvesting complexes, that greatly expand the absorptance capacity of the reactions, have evolved along three principal lines. First, in green plants distinct chlorophyll (Chl) a/b-binding intrinsic membrane complexes are associated with RC 1 and RC 2. The Chl a/b-binding complexes may add about 200 additional chromophores to RC 2. Second, cyanobacteria and red algae have a significant type of antenna (with RC 2) in the form of phycobilisomes. A phycobilisome, depending on the size and phycobiliprotein composition adds from 700 to 2300 light-absorbing chromophores. Red algae also have a sizable Chl a-binding complex associated with RC 1, contributing an additional 70 chromophores. Third, in chromophytes a variety of carotenoid-Chl-complexes are found. Some are found associated with RC 1 where they may greatly enhance the absorptance capacity. Association of complexes with RC 2 has been more difficult to ascertain, but is also expected in chromophytes. The apoprotein framework of the complexes provides specific chromophore attachment sites, which assures a directional energy transfer whithin complexes and between complexes and reaction centers. The major Chl-binding antenna proteins generally have a size of 16–28 kDa, whether of chlorophytes, chromophytes, or rhodophytes. High sequence homology observed in two of three transmembrane regions, and in putative chlorophyll-binding residues, suggests that the complexes are related and probably did not evolve from widely divergent polyphyletic lines.Abbreviations APC allophycocyanin - B phycoerythrin-large bangiophycean phycoerythrin - Chl chlorophyll - L_CM linker polypeptide in phycobilisome to thylakoid - FCP fucoxanthin Chl a/c complex - LHC(s) Chl-binding light harvesting complex(s) - LHC I Chl-binding complex of Photosystem I - LHC II Chl-binding complex of Photosystem II - PC phycocyanin - PCP peridinin Chl-binding complex - P700 photochemically active Chl a of Photosystem I - PS I Photosystem I - PS II Photosystem II - RC 1 reaction center core of PS I - RC 2 reaction center core of PS II - R phycoerythrin-large rhodophycean phycoerythrin - sPCP soluble peridinin Chl-binding complex     

Functional characteristics of chlorophyll d-predominating photosynthetic apparatus in intact cells of Acaryochloris marina

Functional organization of the photosynthetic apparatus in the unique chlorophyll d-predominating prokaryote, Acaryochloris marina, was studied using polarographic measurements of single-turnover flash yields, action spectra and optical cross sections for PS-specific reactions. O₂ evolution was indicative of PS II activity, while reversible photoinhibition of respiratory O₂ uptake under aerobic conditions in the presence of DCMU and H₂ photoevolution by anaerobically adapted cells were the indicatives of PS I activity. O₂ evolution in the cells upon single-turnover flashes followed the normal S-state cycle with a period-4 oscillation. Analysis of action spectra for the partial reactions of photosynthesis revealed that: (1) distinct spectral forms of Chl d are nonuniformly distributed between PS I and PS II, e.g. Chl d-695 and Chl d-735 are preferentially located in PS II and PS I, respectively; (2) a minor fraction of Chl a in the cells belongs mostly to PS II; (3) biliproteins transfer excitation energy both to PS II and, with a lower efficiency, PS I; (4) the efficiency of energy transfer from biliproteins to PS II depends on the light quality growth conditions and is larger in white light (WL)-grown cells compared to the red light (RL)-grown cells. Content of functional O₂ evolving PS II centers decreases 2 times in the RL-grown cells relative to the WL-grown cells, whereas content of competent PS I centers involved in photoinhibition of respiration remains almost the same in both the cultures. The effective antenna size of PS I was estimated to be 80–90 Chl d including 3–10 molecules absorbing at 735 nm. The effective optical cross-section of PS II corresponded to 90–100 Chl d and, presumably, 4 Chl a + 2 Pheo a [Mimuro et al. (1999) Biochim Biophys Acta 1412: 37–46]. Optical cross-section measurements indicated that the functional PS II units of A. marina attach one rod of four hexameric units of biliproteins. This revised version was published online in June 2006 with corrections to the Cover Date.     

Pigment exchange of Photosystem II reaction center by chlorophyll d

Pigment exchanges among photosystem reaction centers (RCs) are useful for the identification and functional analysis of chromophores in photosynthetic organisms. Pigment replacement within the spinach Photosystem II RC was performed with Chl d derived from the oxygenic alga Acaryochloris marina, using a protocol similar to that reported previously [Gall et al. (1998) FEBS Lett 434: 88–92] based on the incubation of reaction centers with an excess of other pigments. In this study, we analyzed Chl d-modified monomeric RC which was separated from Chl d-modified dimeric RC by size-exclusion chromatography. Based on the assumption of a constant ratio of two Pheo a molecules per RC, the number of Chl a molecules in Chl d-modified monomeric RCs was found to decrease from six to four. The absorption spectrum of the Chl d-modified monomeric RC at room temperature showed a large peak at 699.5 nm originating from Chl d and a small peak at 672.5 nm orignating from Chl a. Photoaccumulation of the Pheo a⁻ in Chl d-modified monomeric RC, in the presence of sodium dithionate and methyl viologen, did not differ significantly from that in control RC, showing that the Chl d-modified monomeric RC retains its charge separation activity and photochemically active Pheo a.     

Excitation energy transfer in Photosystem I from oxygenic organisms

This Review discusses energy transfer pathways in Photosystem I (PS I) from oxygenic organisms. In the trimeric PS I core from cyanobacteria, the efficiency of solar energy conversion is largely determined by ultrafast excitation transfer processes in the core chlorophyll a (Chl a) antenna network and efficient photochemical trapping in the reaction center (RC). The role of clusters of Chl a in energy equilibration and photochemical trapping in the PS I core is discussed. Dimers of the longest-wavelength absorbing (red) pigments with strongest excitonic interactions localize the excitation in the PS I core antenna. Those dimers that are located closer to the RC participate in a fast energy equilibration with coupled pigments of the RC. This suggests that the function of the red pigments is to concentrate the excitation near the RC. In the PS I holocomplex from algae and higher plants, in addition to the red pigments of the core antenna, spectrally distinct red pigments are bound to the peripheral Chl a/b-binding light-harvesting antenna (LHC I), specifically to the Lhca4 subunit of the LHC I-730 complex. Intramonomeric energy equilibration between pools of Chl b and Chl a in Lhca1 and Lhca4 monomers of the LHC I-730 heterodimer are as fast as the energy equilibration processes within the PS I core. In contrast to the structural stability of the PS I core, the flexible subunit structure of the LHC I would probably determine the observed slow excitation energy equilibration processes in the range of tens of picoseconds. The red pigments in the LHC I are suggested to function largely as photoprotective excitation sinks in the peripheral antenna of PS I. This revised version was published online in August 2006 with corrections to the Cover Date.     

Endolithic chlorophyll d-containing phototrophs

Cyanobacteria in the genus Acaryochloris are the only known oxyphototrophs that have exchanged chlorophyll a (Chl a) with Chl d as their primary photopigment, facilitating oxygenic photosynthesis with near infrared (NIR) light. Yet their ecology and natural habitats are largely unknown. We used hyperspectral and variable chlorophyll fluorescence imaging, scanning electron microscopy, photopigment analysis and DNA sequencing to show that Acaryochloris-like cyanobacteria thrive underneath crustose coralline algae in a widespread endolithic habitat on coral reefs. This finding suggests an important role of Chl d-containing cyanobacteria in a range of hitherto unexplored endolithic habitats, where NIR light-driven oxygenic photosynthesis may be significant.     

Kinetic and Energetic Model for the Primary Processes in Photosystem II

A detailed model for the kinetics and energetics of the exciton trapping, charge separation, charge recombination, and charge stabilization processes in photosystem (PS) II is presented. The rate constants describing these processes in open and closed reaction centers (RC) are calculated on the basis of picosecond data (Schatz, G. H., H. Brock, and A. R. Holzwarth. 1987. Proc. Natl. Acad. Sci. USA. 84:8414-8418) obtained for oxygen-evolving PS II particles from Synechococcus sp. with ~80 chlorophylls/P₆₈₀. The analysis gives the following results. (a) The PS II reaction center donor chlorophyll P₆₈₀ constitutes a shallow trap, and charge separation is overall trap limited. (b) The rate constant of charge separation drops by a factor of ~6 when going from open (Q-oxidized) to closed (Q-reduced) reaction centers. Thus the redox state of Q controls the yield of radical pair formation and the exciton lifetime in the Chl antenna. (c) The intrinsic rate constant of charge separation in open PS II reaction centers is calculated to be ~2.7 ps^-1. (d) In particles with open RC the charge separation step is exergonic with a decrease in standard free energy of ~38 meV. (e) In particles with closed RC the radical pair formation is endergonic by ~12 meV. We conclude on the basis of these results that the long-lived (nanoseconds) fluorescence generally observed with closed PS II reaction centers is prompt fluorescence and that the amount of primary radical pair formation is decreased significantly upon closing of the RC.     

Chromatic photoacclimation, photosynthetic electron transport and oxygen evolution in the Chlorophyll d-containing oxyphotobacterium Acaryochloris marina

Changes in photosynthetic pigment ratios showed that the Chlorophyll d-dominated oxyphotobacterium Acaryochloris marina was able to photoacclimate to different light regimes. Chl d per cell were higher in cultures grown under low irradiance and red or green light compared to those found when grown under high white light, but phycocyanin/Chl d and carotenoid/Chl d indices under the corresponding conditions were lower. Chl a, considered an accessory pigment in this organism, decreased respective to Chl d in low irradiance and low intensity non-white light sources. Blue diode PAM (Pulse Amplitude Modulation) fluorometry was able to be used to measure photosynthesis in Acaryochloris. Light response curves for Acaryochloris were created using both PAM and O₂ electrode. A linear relationship was found between electron transport rate (ETR), measured using a PAM fluorometer, and oxygen evolution (net and gross photosynthesis). Gross photosynthesis and ETR were directly proportional to one another. The optimum light for white light (quartz halogen) was about 206 ± 51 μmol m^− 2 s^− 1 (PAR) (Photosynthetically Active Radiation), whereas for red light (red diodes) the optimum light was lower (109 ± 27 μmol m^− 2 s^− 1 (PAR)). The maximum mean gross photosynthetic rate of Acaryochloris was 73 ± 7 μmol mg Chl d^− 1 h^− 1. The gross photosynthesis/respiration ratio (P_g/R) of Acaryochloris under optimum conditions was about 4.02 ± 1.69. The implications of our findings will be discussed in relation to how photosynthesis is regulated in Acaryochloris.     

Pigment organization in the photosynthetic apparatus of Roseiflexus castenholzii

The light-harvesting-reaction center (LHRC) complex from the chlorosome-lacking filamentous anoxygenic phototroph (FAP), Roseiflexus castenholzii (R. castenholzii) was purified and characterized for overall pigment organization. The LHRC is a single complex that is comprised of light harvesting (LH) and reaction center (RC) polypeptides as well as an attached c-type cytochrome. The dominant carotenoid found in the LHRC is keto-γ-carotene, which transfers excitation to the long wavelength antenna band with 35% efficiency. Linear dichroism and fluorescence polarization measurements indicate that the long wavelength antenna pigments absorbing around 880 nm are perpendicular to the membrane plane, with the corresponding Q_y transition dipoles in the plane of the membrane. The antenna pigments absorbing around 800 nm, as well as the bound carotenoid, are oriented at a large angle with respect to the membrane. The antenna pigments spectroscopically resemble the well-studied LH2 complex from purple bacteria, however the close association with the RC makes the light harvesting component of this complex functionally more like LH1.     

Isolation and spectral characterization of Photosystem II reaction center from Synechocystis sp. PCC 6803

We isolated highly-purified photochemically active photosystem (PS) II reaction center (RC) complexes from the cyanobacterium Synechocystis sp. PCC 6803 using a histidine-tag introduced to the 47 kDa chlorophyll protein, and characterized their spectroscopic properties. Purification was carried out in a one-step procedure after isolation of PS II core complex. The RC complexes consist of five polypeptides, the same as in spinach. The pigment contents per two molecules of pheophytin a were 5.8 +/- 0.3 chlorophyll (Chl) a and 1.8 +/- 0.1 beta-carotene; one cytochrome b(559) was found per 6.0 Chl a molecules. Overall absorption and fluorescence properties were very similar to those of spinach PS II RCs; our preparation retains the best properties so far isolated from cyanobacteria. However, a clear band-shift of pheophytin a and beta-carotene was observed. Reasons for these differences, and RC composition, are discussed on the basis of the three-dimensional structure of complexes.     

Photosystem I complexes associated with fucoxanthin-chlorophyll-binding proteins from a marine centric diatom, Chaetoceros gracilis

Diatoms occupy a key position as a primary producer in the global aquatic ecosystem. We developed methods to isolate highly intact thylakoid membranes and the photosystem I (PS I) complex from a marine centric diatom, Chaetoceros gracilis. The PS I reaction center (RC) was purified as a super complex with light-harvesting fucoxanthin-chlorophyll (Chl)-binding proteins (FCP). The super complex contained 224 Chl a, 22 Chl c, and 55 fucoxanthin molecules per RC. The apparent molecular mass of the purified FCP-PS I super complex (∼ 1000 kDa) indicated that the super complex was composed of a monomer of the PS I RC complex and about 25 copies of FCP. The complex contained menaquinone-4 as the secondary electron acceptor A₁ instead of phylloquinone. Time-resolved fluorescence emission spectra at 77 K indicated that fast (16 ps) energy transfer from a Chl a band at 685 nm on FCP to Chls on the PS I RC complex occurs. The ratio of fucoxanthin to Chl a on the PS I-bound FCP was lower than that of weakly bound FCP, suggesting that PS I-bound FCP specifically functions as the mediator of energy transfer between weakly bound FCPs and the PS I RC.     

Chlorophylls <Emphasis Type="Italic">d</Emphasis> and <Emphasis Type="Italic">f</Emphasis> and their role in primary photosynthetic processes of cyanobacteria

The finding of unique Chl d- and Chl f-containing cyanobacteria in the last decade was a discovery in the area of biology of oxygenic photosynthetic organisms. Chl b, Chl c, and Chl f are considered to be accessory pigments found in antennae systems of photosynthetic organisms. They absorb energy and transfer it to the photosynthetic reaction center (RC), but do not participate in electron transport by the photosynthetic electron transport chain. However, Chl d as well as Chl a can operate not only in the light-harvesting complex, but also in the photosynthetic RC. The long-wavelength (Q_y) Chl d and Chl f absorption band is shifted to longer wavelength (to 750 nm) compared to Chl a, which suggests the possibility for oxygenic photosynthesis in this spectral range. Such expansion of the photosynthetically active light range is important for the survival of cyanobacteria when the intensity of light not exceeding 700 nm is attenuated due to absorption by Chl a and other pigments. At the same time, energy storage efficiency in photosystem 2 for cyanobacteria containing Chl d and Chl f is not lower than that of cyanobacteria containing Chl a. Despite great interest in these unique chlorophylls, many questions related to functioning of such pigments in primary photosynthetic processes are still not elucidated. This review describes the latest advances in the field of Chl d and Chl f research and their role in primary photosynthetic processes of cyanobacteria.     

An overview on chlorophylls and quinones in the photosystem I-type reaction centers

Minor but key chlorophylls (Chls) and quinones in photosystem (PS) I-type reaction centers (RCs) are overviewed in regard to their molecular structures. In the PS I-type RCs, the prime-type chlorophylls, namely, bacteriochlorophyll (BChl) a′ in green sulfur bacteria, BChl g′ in heliobacteria, Chl a′ in Chl a-type PS I, and Chl d′ in Chl d-type PS I, function as the special pairs, either as homodimers, (BChl a′)₂ and (BChl g′)₂ in anoxygenic organisms, or heterodimers, Chl a/a′ and Chl d/d′ in oxygenic photosynthesis. Conversions of BChl g to Chl a and Chl a to Chl d take place spontaneously under mild condition in vitro. The primary electron acceptors, A ₀, are Chl a-derivatives even in anoxygenic PS I-type RCs. The secondary electron acceptors are naphthoquinones, whereas the side chains may have been modified after the birth of cyanobacteria, leading to succession from menaquinone to phylloquinone in oxygenic PS I.     

18O Labeling of Chlorophyll d in Acaryochloris marina Reveals That Chlorophyll a and Molecular Oxygen Are Precursors

The cyanobacterium Acaryochloris marina was cultured in the presence of either H₂¹⁸O or ¹⁸O₂, and the newly synthesized chlorophylls (Chl a and Chl d) were isolated using high performance liquid chromatography and analyzed by mass spectroscopy. In the presence of H₂¹⁸O, newly synthesized Chl a and d, both incorporated up to four isotopic ¹⁸O atoms. Time course H₂¹⁸O labeling experiments showed incorporation of isotopic ¹⁸O atoms originating from H₂¹⁸O into Chl a, with over 90% of Chl a ¹⁸O-labeled at 48 h. The incorporation of isotopic ¹⁸O atoms into Chl d upon incubation in H₂¹⁸O was slower compared with Chl a with ∼50% ¹⁸O-labeled Chl d at 115 h. The rapid turnover of newly synthesized Chl a suggested that Chl a is the direct biosynthetic precursor of Chl d. In the presence of ¹⁸O₂ gas, one isotopic ¹⁸O atom was incorporated into Chl a with approximately the same kinetic incorporation rate observed in the H₂¹⁸O labeling experiment, reaching over 90% labeling intensity at 48 h. The incorporation of two isotopic ¹⁸O atoms derived from molecular oxygen (¹⁸O₂) was observed in the extracted Chl d, and the percentage of double isotopic ¹⁸O-labeled Chl d increased in parallel with the decrease of non-isotopic-labeled Chl d. This clearly indicated that the oxygen atom in the C3¹-formyl group of Chl d is derived from dioxygen via an oxygenase-type reaction mechanism.     

Incorporation of the chlorophyll d-binding light-harvesting protein from Acaryochloris marina and its localization within the photosynthetic apparatus of Synechocystis sp. PCC6803

The gene encoding a chlorophyll d-binding light-harvesting protein, pcbA from Acaryochloris marina (now called as accessory Chlorophyll Binding Protein CBPII) marked with a His-tag was transformed into the genome of Synechocystis PCC6803. Protein gel electrophoresis and western blotting confirmed that this foreign chlorophyll d-binding protein CBPII was expressed and integrated into the thylakoid membrane and bound with chlorophyll a, the only type of chlorophyll present in Synechocystis PCC 6803. Native electrophoresis suggested that CBPII interacts with photosystem II of Synechocystis PCC 6803. Surprisingly, spectral analyses showed that the phycobiliproteins were suppressed in the transformed Synechocystis pcbA⁺, with a lower ratio of phycobilins to chlorophyll a. These results suggest that there are competitive interactions between the external antenna system of phycobiliproteins and the integral antenna system of chlorophyll-bound protein complexes.     

Chromatic Regulation of Photosystem Composition in the Photosynthetic System of Red and Blue-Green Algae

A chromatic adaptation in the photosynthetic quantum yield forthe light mainly absorbed by chlorophyll a (Chl a light) firstfound by Yocum (1951) was studied with one red and three blue-greenalgal strains. When the cells were grown under a weak Chl alight, the quantum yield in all the strains increased. Comparisonof photosystem (PS) compositions, including phycobilin (PBP)and Chl a antennae, reaction centers I and II, in the cellsgrown under the light mainly absorbed by PBP and Chl a revealedthat changes in quantum yield could be attributed to changesin the ratio of PS I/II; PS I/II becomes larger than 1 underPBP light but decreases to 1 in most cases under Chl a light.The change in the PS I/II ratio is due solely to the changesin the PS I population in the cell; PS II remains constant.These results are similar to the intensity-dependent responsein PS composition. A common hypothesis for both the chromatic and intensity-inducedregulation of PS composition was proposed based on the ideaof balance between the electron flow from H₂O to NADP drivenby PS I and II and the cyclic one driven by PS I. (Received May 16, 1985; Accepted September 4, 1985)     

Assembly and composition of the chlorophyll a-b light-harvesting complex of barley (Hordeum vulgare L.): Immunochemical analysis of chlorophyll b-less and chlorophyll b-deficient mutants

The chlorina-f2 mutant of barley (Hordeum vulgare L.) contains no chlorophyll b in its light-harvesting antenna, whereas the chlorina-103 mutant contains approximately 10% of the chlorophyll b found in wild-type. The absolute chlorophyll antenna size for Photosystem-II in wild-type, chlorina-103 and chlorina-f2 mutant was 250, 58 and 50 chlorophyll molecules, respectively. The absolute chlorophyll antenna size for Photosystem-I in wild-type, chlorina-103 and chlorina-f2 mutant was 210, 137 and 150 chlorophyll molecules, respoectively. In spite of the smaller PS I antenna size in the chlorina mutants, immunochemical analysis showed the presence of polypeptide components of the LHC-I auxiliary antenna with molecular masses of 25, 19.5 and 19 kDa. The chlorophyll a-b-binding LHC-II auxiliary antenna of PS II contained five polypeptide subunits in wild-type barley, termed a, b, c, d and e, with molecular masses of 30, 28, 27, 24 and 21 kDa, respectively. The polypeptide composition of the LHC-II auxiliary antenna of PS II was found to be identical in the two mutants, with only the 24 kDa subunit d present at an equal copy number per PS II in each of the mutants and in the wild-type barley. This d subunit assembles stably in the thylakoid membrane even in the absence of chlorophyll b and exhibits flexibility in its complement of bound chlorophylls. We suggest that polypeptide subunit d binds most of the chlorophyll associated with the residual PS II antenna in the chlorina mutants and that is proximal to the PS II-core complex.Abbreviations CP chlorophyll-protein - LHC the chlorophyll a-b binding light-harvesting complex - LHC-II subunit a the Lhcb4/5 gene product - subunit b the Lhcb1 gene product - subunit c Lhcb2 the gene product - subunit d the Lhcb3 gene product - subunit e the Lhcb6 gene product - PMSF phenylmethane sulphonyl fluoride - RC reaction center - Q_A the primary quinone electron acceptor of Photosystem-II - P700 the reaction center of PS I     

Antenna ring around trimeric Photosystem I in chlorophyll b containing cyanobacterium Prochlorothrix hollandica

Prochlorothrix hollandica is one of the three known species of an unusual clade of cyanobacteria (formerly called “prochlorophytes”) that contain chlorophyll a and b molecules bound to intrinsic light-harvesting antenna proteins. Here, we report the structural characterization of supramolecular complex consisting of Photosystem I (PSI) associated with the chlorophyll a/b-binding Pcb proteins. Electron microscopy and single particle image analysis of negatively stained preparations revealed that the Pcb-PSI supercomplex consists of a central trimeric PSI surrounded by a ring of 18 Pcb subunits. We conclude that the formation of the Pcb ring around trimeric PSI represents a mechanism for increasing the light-harvesting efficiency in chlorophyll b-containing cyanobacteria.     

Modulation of the light-harvesting chlorophyll antenna size in Chlamydomonas reinhardtii by TLA1 gene over-expression and RNA interference

Truncated light-harvesting antenna 1 (TLA1) is a nuclear gene proposed to regulate the chlorophyll (Chl) antenna size in Chlamydomonas reinhardtii. The Chl antenna size of the photosystems and the chloroplast ultrastructure were manipulated upon TLA1 gene over-expression and RNAi downregulation. The TLA1 over-expressing lines possessed a larger chlorophyll antenna size for both photosystems and contained greater levels of Chl b per cell relative to the wild type. Conversely, TLA1 RNAi transformants had a smaller Chl antenna size for both photosystems and lower levels of Chl b per cell. Western blot analyses of the TLA1 over-expressing and RNAi transformants showed that modulation of TLA1 gene expression was paralleled by modulation in the expression of light-harvesting protein, reaction centre D1 and D2, and VIPP1 genes. Transmission electron microscopy showed that modulation of TLA1 gene expression impacts the organization of thylakoid membranes in the chloroplast. Over-expressing lines showed well-defined grana, whereas RNAi transformants possessed loosely held together and more stroma-exposed thylakoids. Cell fractionation suggested localization of the TLA1 protein in the inner chloroplast envelope and potentially in association with nascent thylakoid membranes, indicating a role in Chl antenna assembly and thylakoid membrane biogenesis. The results provide a mechanistic understanding of the Chl antenna size regulation by the TLA1 gene.