Ultrastructure of thylakoid membranes in C. reinhardtii: Evidence for variations in the partition coefficient of the light-harvesting complex-containing particles upon membrane fracture

The ultrastructure of the thylakoid membranes of Chlamydomonas reinhardtii was investigated using cell cultures grown under light intensities of 200 and 4000 lx, respectively. A significant difference in the size distribution of the exoplasmic fracture face (EF) particles appears upon Mg²⁺ treatment of broken cell preparations from the two light growth conditions. Particles larger than 150 Å are seen at 4000 lx only. However neither the absorption spectra of chlorophyll at 77 °K, nor the chlorophyll a/chlorophyll b ratios differ in the two cell batches. In addition, the polypeptide composition of the thylakoid membranes and the Mg²⁺ effect (spillover) on the photochemical rate of Photosystem II are the same in both conditions. We conclude that the partition coefficient between the two fracture faces of light-harvesting complex-containing particles is variable. It depends on Mg²⁺ ion concentration in the incubating medium of the membranes and on the light growth conditions of the cell cultures. Our results suggest that 60- to 80-Å protoplasmic fracture face (PF) particles containing the light-harvesting complexes can aggregate either in larger PF particles (100–120 Å) or in EF particles larger than 120 Å which also contain the Photosystem II centers. That some light-harvesting complexes are located on the PF faces is confirmed by the analysis of the BF4 mutant of C. reinhardtii lacking in chlorophyll-protein complex II. The PF faces of the BF4 thylakoids display a reduced number of particles as compared to that in the wild type.     

Ultrastructure and freeze-fracture studies of the thylakoids ofMantoniella squamata (Prasinophyceae)

Summary The ultrastructure and the supramolecular organization of the thylakoids of the small green flagellate,Mantoniella squamata, were examined in thin sections and freeze-fracture preparations. The whole chloroplast is tightly packed with thylakoids, which show a pattern of meandering, branching and/or anastomosing membranes. In freeze-fracture preparations only two fracture-faces can be distinguished: the PF- and the EF-face. The PF-face has a much higher particle density than the EF-face (PF: 4086 particles/m²; EF: 865 particles/m²). The EF-face is not as uniform as the PF-face. The areas which are packed with particles probably correspond to closely appressed thylakoid regions or adhesive patches, noticed in thin sections in some areas. The mean particle size on both faces is also different (EF: 10.5 nm; PF: 8.6 nm), but no information about the classification of the particles to special protein complexes is available at this time.Abbreviations chl chlorophyll - EF exoplasmic fracture face - ER endoplasmic reticulum - LHC light-harvesting chlorophyll-protein complex - PF protoplasmic fracture face - PS I photosystem I - PS II photosystem II     

Development of chloroplast membranes during light induced greening ofChlorella fusca g-2

Summary The development of thylakoid protein complexes during light induced greening of a mutant ofChlorella fusca was studied. Separation of chlorophyll-protein complexes and thylakoid polypeptides by LDS-polyacrylamide gel electrophoresis show that cells grown in the dark contain proteins belonging to coupling factor and cytochrome f/b₆ complex. Parts of both reaction centers are present also. The antennae complexes are specifically lost in yellow cells. The changes in polypeptide pattern at different stages of development in the light are related to ultrastructural changes. The beginning of membrane appression can be correlated with the appearance of the light-harvesting complex II. While the average diameter of EF-particles increases throughout the greening process, their densitiesapart from the rearrangement due to membrane stacking-remain fairly constant. The kinetics of EF_u-particle enlargement are different from those of EF_s-particles.PF-faces in thylakoids grown in the dark contain particles of uniform diameter but some of them protrude more from the fracture plane than do their neighbors. During the first hours of greening, their density increases and two classes develop. From the beginning of membrane stacking, the composition of PF_u-faces remains constant and PF_s-particles increase in number for some time.Results are discussed on the basis of present knowledge of structurefunction relations in thylakoids.Abbreviations CF _o intrinsic membrane complex of the coupling factor - EF, EF _s ,EF _u exoplasmic fracture face, stacked and unstacked region, respectively - LDS lithium dodecyl sulfate - LHCII light-harvesting complex of Photosystem II - PF, PF _s ,PF _u protoplasmic fracture face, stacked and unstacked region, respectively - PS I andPS II Photosystem I and Photosystem II     

The eyespot of the flagellateTetraselmis cordiformis stein (Chlorophyceae): Structural spezialization of the outer chloroplast membrane and its possible significance in phototaxis of green algae

Summary The eyespot region of the flagellateTetraselmis cordiformis Stein (Chlorophyceae) was investigated with the freeze-fracture technique. The only fracture faces observed in this region were the two complementary fracture faces (PF and EF) of the outer chloroplast envelope membrane. Intramembranous particle numbers on both fracture faces of this membrane were significantly higher in the eyespot region as compared to regions outside the eye-spot. Higher numbers of particles on the PF face in the eyespot region were mainly caused by an increase in particle numbers of the size class 6–8 mm, while on the EF face particle size distribution was not significantly different between eyespot and other regions. Functional implications are discussed and evidence is presented that the outer chloroplast envelope membrane may be the site of photoreceptor location in green algal phototaxis.     

Ultrastructural studies on the membrane systems and cell inclusions of the filamentous prochlorophyte Prochlorothrix hollandica

The outer membrane of Prochlorothrix hollandica is covered with a network of fine fibrils on its surface and separated from the cytoplasmic membrane by an electrondense peptidoglycan layer (8 to 20 nm thick). The thylakoid membranes are arranged in stacked and unstacked regions which present four characteristic fracture faces with different numbers and sizes of intramembrane particles. Cell inclusions such as polyhedral bodies (carboxysomes), ribosomes, and polyphosphate granules were found in Prochlorothrix hollandica. Another type of cell inclusions was identified by its characteristic shape (a cylindre with conical caps) and a regular striation as gas vesicles. It is concluded that the organism is in its morphological structure similar to the cyanobacteria.Abbreviations C carboxysome - CM cytoplasmic membrane - EF_s, EF_u exoplasmic fracture face of stacked and unstacked membrane area, respectively - ES exoplasmic surface - PF_s, PF_u plasmic fracture face of stacked and unstacked membrane area, respectively - PG peptidoglycan layer - TM thylakoid membrane Dedicated to Prof. Dr. D. Peters, Hamburg, on the occasion of his 75th birthday     

The fluidity of chloroplast thylakoid membranes and their constituent lipids: A comparative study by ESR

Comparative measurements were made of the fluidity of chloroplast thylakoids, total membrane lipids and polar lipids utilizing the order parameter and motion of spin labels.No significant differences were found in the fluidity of membranes or total membrane lipids from a wild type and a mutant barley (Hordeum vulgare chlorina f₂ mutant) which lacks chlorophyll $\text{b}$ and a 25 000 dalton thylakoid polypeptide. Redistribution of intrinsic, exoplasmic face (EF) membrane particles by unstacking thylakoid membranes in low salt medium also had no effect on membrane fluidity. However, heating of isolated thylakoids decreased membrane fluidity.The fluidity of vesicles composed of membrane lipids is much greater than that of the corresponding membranes. Fluidity of the membranes, however, increased during greening indicating that the rigidity of the membranes, compared with that of total membrane lipids, is not caused by chlorophyll or its associated peptides. It is concluded that the restriction of motion in the acyl chains in the thylakoids is not caused by chlorophyll or the major intrinsic polypeptide but by some other protein components.     

The Supramolecular Organization of Photosynthetic Membranes of the Thallus Stage of Porphyra leucosticta Thuret. (Bangiales,Rhodophyta) Visualized by Freeze-Fracture

The supramolecular organization of the thylakoid membranes of the thallus stage in the red alga Porphyra leucosticta is studied in replicas of rapidly frozen and fractured cells. Freeze-fractured thylakoid membranes exhibit only two types of fracture faces (EF and PF), because the lamellae in red algal chloroplasts are not stacked. The PF reveals numerous, tightly packed, but randomly distributed particles (density range from 2970 to 3550 particles/μm²). In contrast, the EF particles appear organized into parallel rows, the spacing of which is about 60–70 nm (about 8–9 particles occur along 100 nm of the line that is formed). Significant numbers of single EF particles are randomly distributed between the EF particle rows. The particles on both fracture faces (PF and EF) fall into two size classes: 10 to 11 nm (major size class) and 14 to 15 nm (minor size class).     

Ultrastructural organization of chloroplast membranes in mutants of Pisum sativum L. with impaired activity in the photosystems

The ultrastructural organization and the photosynthesis reactions of chloroplast membranes were studied in three lethal mutants of Pisum sativum, Chl-1, Chl-19 and Chl-5, all lacking the capacity to evolve oxygen. The rates of 2,6-dichloroindophenol reduction, delayed fluorescence and electron-spin-resonance signal 1 indicate that Chl-1 and Chl-19 have an impaired activity in photosystem II (PS II), while in Chl-5 the electron transport is blocked between PS I and the reactions of CO₂ fixation. Ultrathin sectioning demonstrates the presence of giant grana in the chloroplasts of Chl-1 and Chl-19, while the chloroplast structure of the Chl-5 is very similar to that of the wild-type. The grana of the Chl-19 mutant contain large multilamellar regions of tightly packed membranes. When the chloroplast membranes were studied by freeze-fracture, the exoplasmic and protoplasmic fracture faces (EF and PF, respectively) in both stacked and unstacked membranes were found to show large differences in particle concentrations and relative population area (per m²), and also in particle size distribution, between all mutant chloroplast membranes and the wild-type. A close correlation between increasing k_mt (ratio of particle concentrations on PF/EF) and PS II activity was observed. The differences in particle concentrations on both fracture faces in different regions of the intact chloroplast membranes of the wild-type are the consequence of a rearrangement of existing membrane components by lateral particle movements since quantitative measurements demonstrate almost complete conservation of intramembrane particles in number and size during the stacking of stroma thylakoid membranes. The results indicating particle movements strongly support the concept that the chloroplast membranes have a highly dynamic structure.Abbreviations DPIP 2,6-dichloroindophenol - EF and PF exoplasmic and protoplasmic fracture faces, respectively - PS I and PS II photosystems I and II, respectively     

Spatial relationship of photosystem I, photosystem II, and the light- harvesting complex in chloroplast membranes

We have previously demonstrated (Armond, P. A., C. J. Arntzen, J.-M. Briantais, and C. Vernotte. 1976. Arch. Biochem. Biophys. 175:54-63; and Davis, D. J., P. A. Armond, E. L. Gross, and C. J. Arntzen. 1976. Arch. Biochem. Biophys. 175:64-70) that pea seedlings which were exposed to intermittent illumination contained incompletely developed chloroplasts. These plastids were photosynthetically competent, but did not contain grana. We now demonstrate that the incompletely developed plastids have a smaller photosynthetic unit size; this is primarily due to the absence of a major light-harvesting pigment-protein complex which is present in the mature membranes. Upon exposure of intermittent- light seedlings to continuous white light for periods up to 48 h, a ligh-harvesting chlorophyll-protein complex was inserted into the chloroplast membrane with a concomitant appearance of grana stacks and an increase in photosynthetic unit size. Plastid membranes from plants grown under intermediate light were examined by freeze-fracture electron microscopy. The membrane particles on both the outer (PF) and inner (EF) leaflets of the thylakoid membrane were found to be randomly distributed. The particle density of the PF fracture face was approx. four times that of the EF fracture face. While only small changes in particle density were observed during the greening process under continuous light, major changes in particle size were noted, particularly in the EF particles of stacked regions (EFs) of the chloroplast membrane. Both the changes in particle size and an observed aggregation of the EF particles into the newly stacked regions of the membrane were correlated with the insertion of light-harvesting pigment- protein into the membrane. Evidence is presented for identification of the EF particles as the morphological equivalent of a "complete" photosystem II complex, consisting of a phosochemically active "core" complex surrounded by discrete aggregates of the light-harvesting pigment protein. A model demonstrating the spatial relationships of photosystem I, photosystem II, and the light-harvesting complex in the chloroplast membrane is presented.     

Developmental Loss of Photosystem II Activity and Structure in a Chloroplast-Encoded Tobacco Mutant, Lutescens-1

Lutescens-1, a tobacco mutant with a maternally inherited dysfunction, displayed an unusual developmental phenotype. In vivo measurement of chlorophyll fluorescence revealed deterioration in photosystem II (PSII) function as leaves expanded. Analysis of thylakoid membrane proteins by polyacrylamide gel electrophoresis indicated the physical loss of nuclear- and chloroplast-encoded polypeptides comprising the PSII core complex concomitant with loss of activity. Freeze fracture electron micrographs of mutant thylakoids showed a reduced density, compared to wild type, of the EF_s particles which have been shown previously to be the structural entity containing PSII core complexes and associated pigment-proteins. The selective loss of PSII cores from thylakoids resulted in a higher ratio of antenna chlorophyll to reaction centers and an altered 77 K chlorophyll fluorescence emission spectra; these data are interpreted to indicate functional isolation of light-harvesting chlorophyll a/b complexes in the absence of PSII centers. Examination of PSII reaction centers (which were present at lower levels in mutant membranes) by monitoring the light-dependent phosphorylation of PSII polypeptides and flash-induced O₂ evolution patterns demonstrated that the PSII cores which were assembled in mutant thylakoids were functionally identical to those of wild type. We conclude that the lutescens-1 mutation affected the correct stoichiometry of PSII centers, in relation to other membrane constituents, by disrupting the proper assembly and maintenance of PSII complexes in lutescens-1 thylakoid membranes.     

Biogenesis of thylakoid membranes with emphasis on the process in Chlamydomonas

Recent results obtained by electron microscopic and biochemical analyses of greening Chlamydomonas reinhardtii y1 suggest that localized expansion of the plastid envelope is involved in thylakoid biogenesis. Kinetic analyses of the assembly of light-harvesting complexes and development of photosynthetic function when degreened cells of the alga are exposed to light suggest that proteins integrate into membrane at the level of the envelope. Current information, therefore, supports the earlier conclussion that the chloroplast envelope is a major biogenic structure, from which thylakoid membranes emerge. Chloroplast development in Chlamydomonas provides unique opportunities to examine in detail the biogenesis of thylakoids.Abbreviations Rubisco ribulose bisphosphate carboxylase/oxygenase - CAB Chl a/b-binding (proteins) - Chlide chlorophyllide - LHC I light-harvesting complex of PS I - LHC II light-harvesting complex of PS II - Pchlide protochlorophyllide     

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.     

Relationship between chloroplast structure and O2 evolution rate of leaf discs in plants from different biotopes in South Finland

Abstract The chloroplast ultrastructure, especially the thylakoid organization, the polypeptide composition of the thylakoid membranes and photosynthetic O₂ evolution rate, chlorophyll (Chl) content and Chi a/b ratio were studied in leaves of nine plants growing in contrasting biotopes in the wild in South Finland. All the measurements were made at the beginning of the period of main growth on leaves approaching full expansion, when the CO₂-saturated O₂ evolution rate (measured at 20°C and 1500 μmol photons m^?2s^?1) was at a maximum, ranging from 19.2 to 6.9 μmol O₂ cm^?2 h^?1. Among the species, the Chi a/b ratio varied between 3.75 and 2.71. In the mesophyll chloroplasts, the ratio of the total length of appressed to non-appressed thylakoid membranes varied between 1.07 and 1.79, the number of partitions per granum varied between 2.8 and 12.0 and the grana area between 21 and 42% of the chloroplast area. There was a significant relationship between the rate of O₂ evolution of the leaf discs and the thylakoid organization in the mesophyll chloroplasts. The higher the O₂ evolution rate, the lower was the ratio of the total length of appressed to non-appressed thylakoid membranes and also the lower the grana area. Although the relationship of the photosynthetic rate with the Chi content and the Chi a/b ratio of the leaves was not as clear, a significant negative correlation existed between the Chi a/b ratio and the ratio of appressed to non-appressed thylakoid membranes, indicating lateral heterogeneity in the distribution of different Chl- protein complexes.     

Fine structure of the chloroplast membranes ofEuglena gracilis as revealed by freeze-cleaving and deep-etching techniques

Summary The chloroplasts ofEuglena gracilis have been examined by freeze-cleaving and deep-etching techniques.The two chloroplast envelope membranes exhibit distinct fracture faces which do not resemble any of the thylakoid fracture faces.Freeze-cleaved thylakoid membranes reveal four split inner faces. Two of these faces correspond to stacked membrane regions, and two to unstacked regions. Analysis of particle sizes on the exposed faces has revealed certain differences from other chloroplast systems, which are discussed. Thylakoid membranes inEuglena are shown to reveal a constant number of particles per unit area (based on the total particle number for both complementary faces) whether they are stacked or unstacked.Deep-etchedEuglena thylakoid membranes show two additional faces, which correspond to true inner and outer thylakoid surfaces. Both of these surfaces carry very uniform populations of particles. Those on the external surface (the A surface) are round and possess a diameter of approximately 9.5 nm. Those on the inner surface (the D surface) appear rectangular (as paired subunits) and measure approximately 10 nm in width and 18 nm in length. Distribution counts of particles show that the number of particles per unit area revealed by freeze-cleaving within the thylakoid membrane approximates closely the number of particles exposed on the external thylakoid surface (the A surface) by deep-etching. The possible significance of this correlation is discussed. The distribution of rectangular particles on the inner surface of the thylakoid sac (D surface) seems to be the same in both stacked and unstacked membrane regions. We have found no correlation between the D surface particles and any clearly defined population of particles on internal, freeze-cleaved membrane faces. These and other observations suggest that stacked and unstacked membranes are similar, if not identical in internal structure.     

Effects of pigment-deficient mutants on the accumulation of photosynthetic proteins in maize

We have monitored the accumulation of photosynthetic proteins in developing pigment-deficient mutants of Zea mays. The proteins examined are the CO₂-fixing enzymes, phoshoenolpyruvate carboxylase (E.C. 4.1.1.31) and ribulose-1,5-bisphosphate carboxylase (E.C.4.1.1.39), and three thylakoid membrane proteins, the light-harvesting chlorophyll a/b binding protein (LHCP) of photosystem II, the 65 kilodalton chlorophyll a binding protein of photosystem I and the alpha subunit polypeptide of coupling factor I. Using a sensitive protein-blot technique, we have compared the relative quantities of each protein in mutants and their normal siblings. Carboxylase accumulation was found to be independent of chlorophyll content, while the amounts of the thylakoid proteins increase at about the same time as chlorophyll in delayed-greening mutants. The relative quantity of LHCP is closely correlated with the relative quantity of chlorophyll at all stages of development in all mutants. Because pigment-deficient mutants are arrested at early stages in chloroplast development, these findings suggest that the processes of chloroplast development, chlorophyll synthesis and thylakoid protein accumulation are coordinated during leaf development but that carboxylase accumulation is controlled by different regulatory mechanisms. A white leaf mutant was found to contain low levels of LHCP mRNA, demonstrating that the accumulation of LHCP mRNA is not controlled exclusively by phytochrome.     

Photochemical Apparatus Organization in the Chloroplasts of Two Beta vulgaris Genotypes

The composition and structural organization of thylakoid membranes of a low chlorophyll mutant of Beta vulgaris was investigated using spectroscopic, kinetic and electrophoretic techniques. The data obtained were compared with those of a standard F1 hybrid of the same species. The mutant was depleted in chlorophyll b relative to the hybrid and it had a higher photosystem II/photosystem I reaction center (Q/P₇₀₀) ratio and a smaller functional chlorophyll antenna size. Analysis of thylakoid membranes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the mutant lacked a portion of the chlorophyll a/b light-harvesting complex but was enriched in the photosystem II reaction center chlorophyll protein complex. Comparison of functional antenna sizes and of photosystem stoichiometries determined electrophoretically were in good agreement with those determined spectroscopically. Both approaches indicated that about 30% of the total chlorophyll was associated with photosystem I and about 70% with photosystem II. A greater proportion of photosystem II_β was detected in the mutant. The results suggest that a higher photosystem II to photosystem I ratio in the sugar beet mutant has apparently compensated for the smaller photosystem II chlorophyll light-harvesting antenna in its chloroplasts. Moreover, a lack of chlorophyll a/b light-harvesting complex correlates with the abundance of photosystem II_β. It is proposed that a developmental relationship exists between the two types of photosystem II where photosystem II_β is a precursor form of photosystem II_α occurring prior to the addition of the chlorophyll a/b light-harvesting complex and grana formation.     

A chlorophyll b-less mutant of Chlamydomonas reinhardii lacking in the light-harvesting chlorophyll complex but not in its apoproteins

The mutant pg 113, derived from Chlamydomonas reinhardii, arg₂⁻ mt⁺ (parent strain), completely lacks chlorophyll (Chl) b but is still able to grow under autotrophic conditions. The light-harvesting Chl complex (LHCP) is absent. This is shown (a) by the lack of the corresponding signal in the CD spectrum of thylakoids and (b) by the absence of the band of the LHCP after electrophoresis of partially solubilized thylakoid membranes on lithium dodecyl sulfate polyacrylamide gels. All the other chlorophyll-protein complexes are present. In spite of the absence of the LHCP, all the polypeptide components of this complex are present in the mutant in the same ratios as in the parent strain, although in slightly reduced amounts. The LHC apoproteins are synthesized, processed and transported into the thylakoid membrane of the mutant. Moreover, the phosphorylation of thylakoid membrane polypeptides, which is related to the regulation of the energy distribution between Photosystem I and II, is the same in the mutant and in the parent strain, indicating that phosphorylation is not dependent on the presence of Chl b. Electron micrographs of thin sections of whole cells show that there are stacked regions of thylakoids in both the mutant and the parent strain chloroplasts. However, in the mutant, stacks are located near the chloroplast envelope, while long stretches or sometimes circles of unstacked membranes are found in the interior, mostly around the pyrenoid.     

Inside-out membrane vesicles isolated from spinach thylakoids

Inside-out thylakoid vesicles have been separated from right-side-out material after press disruption of chloroplast lamellae. The separation was obtained by partition in an aqueous dextran-polyethylene glycol two-phase system, a method which utilizes differences in surface properties for separation of membrane particles. The isolated thylakoid vesicles showed the following inside-out properties: (1) light-induced reversible proton extrusion into the surrounding medium when supplied with the Photosystem II electron acceptor phenyl-p-benzoquinone; (2) a pH rise in the internal phase accompanying the external proton release, (3) sensitivity to trypsin treatment different from that of thylakoid membranes of normal orientation; (4) concave EF and convex PF freeze-fracture faces.     

Enhanced thermal tolerance of photosynthesis and altered chloroplast ultrastructure in a mutant of Arabidopsis deficient in lipid desaturation

A mutant of Arabidopsis thaliana, deficient in activity of the chloroplast n-6 desaturase, accumulated high levels of C_16:1 and C_18:1 lipids and had correspondingly reduced levels of polyunsaturated lipids. The altered lipid composition of the mutant had pronounced effects on chloroplast ultrastructure, thylakoid membrane protein and chlorophyll content, electron transport rates, and the thermal stability of the photosynthetic membranes. The change in chloroplast ultrastructure was due to a 48% decrease in the amount of appressed membranes that was not compensated for by an increased amount of nonappressed membrane. This resulted in a net loss of 36% of the thylakoid membrane per chloroplast and a corresponding reduction in chlorophyll and protein content. Electrophoretic analysis of the chlorophyll-protein complexes further revealed a small decrease in the amount of light-harvesting complex. Relative levels of whole chain and protosystem II electron transport rates were also reduced in the mutant. In addition, the mutation resulted in enhanced thermal stability of photosynthetic electron transport. These observations suggest a central role of polyunsaturated lipids in determining chloroplast structure and maintaining normal photosynthetic function and demonstrate that lipid unsaturation directly affects the thermal stability of photosynthetic membranes.     

Molecular cloning of a chloroplastic proteinassociated with both the envelope and thylakoid membranes

Chloroplasts consist of six morphologically distinct compartments. Each compartment has a specific set of polypeptides that perform distinct biochemical functions. We report here the identification of a membrane-associated protein with a novel localization. This protein was synthesized as a 37 kDa precursor and was processed to a mature protein of 30 kDa after being imported into isolated pea chloroplasts. Fractionation of chloroplasts showed that the 30 kDa mature protein was associated with both of the envelope membranes as well as with thylakoid membranes. Immunocyto-chemical localization of the 30 kDa protein revealed that the protein occurred in clusters in the vicinity of both the envelope and the thylakoid. Possible functions of this 30 kDa protein, inferred from its novel localization pattern, are discussed.Abbreviations CAB light-harvesting chlorophyll a/b-binding protein of photosystem II - prCAB precursor protein to CAB - SS small subunit of ribulose-1,5-bisphosphate carboxylase - prSS precursor protein to SS - RCF relative centrifugation force