Autotrophic cells of the <Emphasis Type="Italic">Synechocystis psbH</Emphasis> deletion mutant are deficient in synthesis of CP47 and accumulate inactive PS II core complexes

Cells of the psbH deletion mutant IC7 of the cyanobacterium Synechocystis PCC 6803 grown in the absence of glucose contain strongly reduced levels of chlorophyll when compared with cells grown in the presence of glucose, or compared with wild-type (WT) cells. Low-temperature fluorescence emission spectra revealed decreased content of both active PS II (Photosystem II) and PS I (Photosystem I) complexes. Analysis of thylakoid membrane complexes of IC7 by native electrophoresis showed a similar set of chlorophyll–proteins, namely a PS II core complex and trimeric and monomeric PS II complexes, as in WT. However, in contrast to WT, the ³⁵S-methionine protein labeling pattern of the mutant exhibited no preferential labeling of the D1 protein in the PS II core complexes, and the labeled D1 and D2 proteins accumulated predominantly in the PS II reaction center lacking CP47. The results show that in autotrophically grown cells of the psbH deletion mutant, selective D1 turnover is inhibited and synthesis of CP47 becomes a limiting step in the PS II assembly.     

Prolonged high light treatment of plant cells results in changes of the amount,the localization and the electrophoretic behavior of several thylakoid membrane proteins

The effect of a 30 h high light treatment on the amount and the localization of thylakoid proteins was analysed in low light grown photoautotrophic cells of Marchantia polymorpha and Chenopodium rubrum. High light treatment resulted in a net loss of D1 protein which was accompanied by comparable losses of other proteins of the PS II core (reaction center with inner antenna). LHC II proteins were not reduced correspondingly, indicating that these complexes are less affected by prolonged high light. High light influenced the distribution of PS II components between the grana and the stroma region of the thylakoid membrane, probably by translocation of the respective PS II proteins. Additionally, modifications of several thylakoid proteins were detected in high light treated cells of C. rubrum. These effects are discussed in relation to photoinhibitory damage and repair processes.Abbreviations BCA bioinchonic acid - chl chlorophyll - CF₁ coupling factor - CYC cycloheximide - GT grana thylakoids - HL high light - LL low light - PAGE polyacrylamide gel electrophoresis - PFD photon flux density - PS I Photosystem I - PS II Photosystem II - RC reaction center - SDS sodium dodecylsulfate - ST stroma thylakoids - Thyl unfractionated thylakoids     

Characterization of a Synechococcus sp. strain PCC 7002 mutant lacking Photosystem I. Protein assembly and energy distribution in the absence of the Photosystem I reaction center core complex

A Synechococcus sp. strain PCC 7002 psaAB::cat mutant has been constructed by deletional interposon mutagenesis of the psaA and psaB genes through selection and segregation under low-light conditions. This strain can grow photoheterotrophically with glycerol as carbon source with a doubling time of 25 h at low light intensity (10 E m^–2 s^–1). No Photosystem I (PS I)-associated chlorophyll fluorescence emission peak was detected in the psaAB::cat mutant. The chlorophyll content of the psaAB::cat mutant was approximately 20% that of the wild-type strain on a per cell basis. In the absence of the PsaA and PsaB proteins, several other PS I proteins do not accumulate to normal levels. Assembly of the peripheral PS I proteins PsaC,PsaD, PsaE, and PsaL is dependent on the presence of the PsaA and PsaB heterodimer core. The precursor form of PsaF may be inserted into the thylakoid membrane but is not processed to its mature form in the absence of PsaA and PsaB. The absence of PS I reaction centers has no apparent effect on Photosystem II (PS II) assembly and activity. Although the mutant exhibited somewhat greater fluorescence emission from phycocyanin, most of the light energy absorbed by phycobilisomes was efficiently transferred to the PS II reaction centers in the absence of the PS I. No light state transition could be detected in the psaAB::cat strain; in the absence of PS I, cells remain in state 1. Development of this relatively light-tolerant strain lacking PS I provides an important new tool for the genetic manipulation of PS I and further demonstrates the utility of Synechococcus sp. PCC 7002 for structural and functional analyses of the PS I reaction center.Abbreviations ATCC American type culture collection - Chl chlorophyll - DCMU 3-(3,4-dichlorophyl)-1,1-dimethylurea - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - HEPES N-[2-hydroxyethyl]piperazine-N-[2-ethanesulfonic acid] - PCC Pasteur culture collection - PS I Photosystem I - PS II Photosystem II - SDS sodium dodecyl sulfate     

Topographical Studies on Subunit Polypeptides of the PS I Reaction Center Complex in the Thylakoid Membranes of the Thermophilic Cyanobacterium Synechococcus sp.

The permeability to protein molecules of the outer limitingmembranes and the thylakoid membranes in hypotonically shockedprotoplasts of the thermophilic cyanobacterium Synechococcussp. was studied by examining the effects of NaBr-washing andpronase E-digestion on phycobiliproteins and a 35 kDa proteinwhich are associated with the outer and inner surface of thethylakoid membranes, respectively, and by measuring photooxidationof added cytochrome c. All the results obtained indicate thatthe shocked protoplasts are in essence a homogenous right side-outthylakoid membrane preparation; the outer limiting membranesare leaky to protein molecules, whereas the thylakoid membranesare still impermeable to proteins. The thylakoid membranes becamepermeable to proteins when the protoplasts were mechanicallydisrupted. Following on from these findings, the membrane topology of subunitpolypeptides of the photosystem I reaction center complex wasstudied. Proteolytic digestion of shocked protoplasts with trypsinand pronase E indicated that four of the five subunit polypeptidesof the PS I reaction center complex are exposed at the stromalsurface of thylakoid membranes; two subunits of 14 and 13 kDawere selectively digested by trypsin, whereas two chlorophyll-bindingsubunits of 62 and 60 kDa were preferentially attacked by pronaseE. However, a 10 kDa subunit appears to be strongly resistantto the proteases. Experiments with mechanically disrupted protoplastsfailed to provide evidence for a uniform transmembrane organizationof the PS I subunits. (Received March 31, 1986; Accepted August 18, 1986)     

Immunocytochemical localization of IdiA, a protein expressed under iron or manganese limitation in the mesophilic cyanobacterium Synechococcus PCC 6301 and the thermophilic cyanobacterium Synechococcus elongatus

Iron-deficiency-induced protein A (IdiA) with a calculated molecular mass of 35 kDa has previously been shown to be essential under manganese- and iron-limiting conditions in the cyanobacteria Synechococcus PCC 6301 and PCC 7942. Studies of mutants indicated that in the absence of IdiA mainly photosystem II becomes damaged, suggesting that the major function of IdiA is in Mn and not Fe metabolism (Michel et al. 1996, Microbiology 142: 2635–2645). To further elucidate the function of IdiA, the immunocytochemical localization of IdiA in the cell was examined. These investigations provided evidence that under mild Fe deficiency IdiA is intracellularly localized and is mainly associated with the thylakoid membrane in Synechococcus PCC 6301. The protein became distributed throughout the cell under severe Fe limitation when substantial morphological changes had already occurred. For additional verification of a preferential thylakoid membrane association of IdiA, these investigations were extended to the thermophilic Synechococcus elongatus. In this cyanobacterium Mn deficiency could be obtained more rapidly than in the mesophilic Synechococcus PCC 6301 and PCC 7942, and the thylakoid membrane structure proved to be more stable under limiting growth conditions. The immunocytochemical investigations with this cyanobacterium clearly supported a thylakoid membrane association of IdiA. In addition, evidence was obtained for a localization of IdiA on the cytoplasmic side of the thylakoid membrane. All available data support a function of IdiA as an Mn-binding protein that facilitates transport of Mn via the thylakoid membrane into the lumen to provide photosystem II with Mn. A possible explanation for the observation that IdiA was not only expressed under Mn deficiency but also under Fe deficiency is given in the discussion. Received: 28 July 1997 / Accepted: 26 November 1997     

Quantitative analysis of 77K fluorescence emission spectra in Synechocystis sp. PCC 6714 and Chlamydomonas reinhardtii with variable PS I/PS II stoichiometries

Low-temperature (77 K) fluorescence emission spectra of intact cells of a cyanobacterium, Synechocystis sp. PCC 6714, and a green alga, Chlamydomonas reinhardtii, were quantitatively analyzed to examine differences in PS I/PS II stoichiometries. Cells cultured under different spectral conditions had various PS I/PS II molar ratios when estimated by oxidation-reduction difference absorption spectra of P700 (for PS I) and Cyt b-559 (for PS II) with thylakoid membranes. The fluorescence emission spectra under the Chl a excitation at 435 nm were resolved into several component bands using curve-fitting methods and the relative band area between PS II (F685 and F695) and PS I (F710 or F720) emissions was compared with the PS I/PS II stoichiometries of the various cell types. The results indicated that the PS I/PS II fluorescence ratios correlated closely with photosystem stoichiometries both in Synechocystis sp. PCC 6714 and in C. reinhardtii grown under different light regimes. Furthermore, the correlation between the PS I/PS II fluorescence ratios and the photosystem stoichiometries is also applicable to vascular plants.     

Photochemical Apparatus Organization in the Diatom Cylindrotheca fusiformis: Photosystem Stoichiometry and Excitation Distribution in Cells Grown under High and Low Irradiance

The photochemical apparatus organization in the thylakoid membraneof the diatom Cylindrotheca fusiformis was investigated in cellsgrown under high and low irradiance. High light (HL, 200µE.m^–2.s^–1)grown cells displayed a relatively low fucoxanthin to chlorophyll(Chl) ratio, a low photosystem (PS) stoichiometry (PSII/PS I=1.3/1.0)and a smaller photosynthetic unit size in both PS I and PS II.Low light (LL, 30µE.m^–2.s^–1) grown cells displayeda 30% elevated fucoxanthin content, elevated PS II/PS I=3.9/1.0and larger photosynthetic unit size for PS II (a change of about100%) and for PS I (by about 30%). In agreement, SDS polyacrylamidegel electrophoresis of thylakoid membrane polypeptides showedgreater abundance of PS I, RuBP carboxylase and ATP synthasepolypeptides in HL cells. In contrast, LL grown cells exhibitedgreater abundance of light-harvesting complex polypeptides.Assuming an efficiency of red (670 nm) light utilization of1.0, the measured efficiency of blue (481 nm) light utilizationwas 0.64 (HL cells) and 0.72 (LL cells). The lower efficiencyof blue versus red light utilization is attributed to the quenchingof absorbed energy by non-fucoxanthin carotenoids. Differencesin the efficiency of blue light utilization between HL and LLgrown cells are attributed to the variable content of fucoxanthin.The results support the hypothesis of a variable Chl a-Chl c-fucoxanthinlight-harvesting antenna associated with PS II and PS I in Cylindrotheca. (Received February 10, 1988; Accepted April 6, 1988)     

Changes in composition of membrane proteins accompanying the regulation of PS I/PS II stoichiometry observed with Synechocystis PCC 6803

Changes in composition of membrane proteins in Synechocystis PCC 6803 induced by the shift of light regime for photosynthetic growth were studied in relation to the regulation of PS I/PS II stoichiometry. Special attention was paid to the changes in abundance of proteins of PS I and PS II complexes. Composition was examined using a LDS-PAGE and a quantitative enzyme immunoassay. Abundance of PsaA/B polypeptides and the PsaC polypeptide of the PS I complex, on a per cell basis, increased under the light regime exciting preferentially PS II and decreased under the light regime exciting mainly PS I. Similar changes were observed with polypeptides of 18.5, 10 and 8.5 kDa. The abundance of other proteins associated with membranes, including PsbA polypeptide of the PS II complex, was fairly constant irrespective of light regime. These results are consistent with our previous observations with other strains of cyanophytes (Anabaena variabilis M2 and Synechocystis PCC 6714) that PS I is the variable component in changes in PS I/PS II stoichiometry in response to changing light regimes for photosynthesis.Abbreviations CBB Coomassie brilliant blue - Chl chlorophyll - EIA enzyme immunoassay - LDS lithium dodecyl sulfate - PAGE polyacrylamide gel electrophoresis - PS photosystem - PVDF polyvinylidene difluoride     

Regulation of Photosystem Composition in Cyanobacterial Photosynthetic System: Evidence Indicating that Photosystem I Formation is Controlled in Response to the Electron Transport State

The quantitative composition of thylakoid components such asthe photosystem (PS) I/PS II ratio in the cyanobacterial photosyntheticsystem is regulated in response to the light regime. The regulationoccurs as changes in PS I content due to control of either PSI formation or decomposition. In order to determine which ofthese two is controlled in this regulation, experiments wereperformed to determine the light-induced PS I decrease in cellsof Synechocystis PCC 6714 under conditions where protein synthesiswas suppressed, i.e. the incubation without a nitrogen sourcefor cell growth or with chloramphenicol. The results revealedthat light-induced PS I decrease did not occur when synthesisof the thylakoid system was suppressed by incubation withouta nitrogen source or by the addition of chloramphenicol, indicatingthat (1) the thylakoid composition is regulted in the processof thylakoid formation and (2) the regulation is achieved bythe control of PS I formation. (Received November 6, 1987; Accepted March 2, 1988)     

<Emphasis Type="Italic">PsbS</Emphasis> genotype in relation to coordinated function of PS II and PS I in <Emphasis Type="Italic">Arabidopsis</Emphasis> leaves

Application of multiple probes to systems that carry specific mutations provides a powerful means for studying how known regulators of light utilization interact in vivo. Two lines of Arabidopsis thaliana were studied, each carrying a unique lesion in the nuclear psbS gene encoding a 22-kDa pigment-binding protein (PS II-S) essential for full expression of photoprotective, rapid-phase, nonphotochemical quenching of chlorophyll fluorescence (NPQ). The PS II-S protein is absent in line npq4-1 due to deletion of psbS. Line npq4-9 expresses normal levels of PS II-S but carries a single amino acid substitution that lowers NPQ capacity by about 50%. A prior report [Peterson RB and Havir EA (2001) Planta 214: 142–152] described an altered pattern of redox states of the acceptor side of Photosystem II (PS II) and donor side of Photosystem I (PS I) for npq4-9 suggesting that interphotosystem electron transport may be restricted by a higher transthylakoid ΔpH in this line. In vivo steady state fluorescence and absorbance measurements (820 nm) confirmed these earlier observations for line npq4-9 but not for npq4-1. Thus, the prior results cannot be correlated simply to a loss of NPQ capacity. Likewise, the kinetics of the 820-nm absorbance change did not indicate a substantial effect of psbS genotype on electron flow from plastoquinol to PS I. A simple model is proposed to relate linear electron transport rate (measured gasometrically) to a parameter (based on fluorescence) that provides a relative measure of the density of excitation available for photochemistry in PS II. Surprisingly, analyses using this model suggested that the in vivo midpoint potential of the primary quinone acceptor in PS II (Q_A) is lowered in both psbS mutant lines. This heretofore-unsuspected role for PS II-S is discussed with regard to: (1) numerous prior reports indicating plasticity of the redox potential of Q_A and (2) the basis for the contrasting regulation of quantum yields of PS I and II in npq4-1 and npq4-9.     

Light adaptation of cyclic electron transport through Photosystem I in the cyanobacterium Synechococcus sp. PCC 7942

Photosystem I-driven cyclic electron transport was measured in intact cells of Synechococcus sp PCC 7942 grown under different light intensities using photoacoustic and spectroscopic methods. The light-saturated capacity for PS I cyclic electron transport increased relative to chlorophyll concentration, PS I concentration, and linear electron transport capacity as growth light intensity was raised. In cells grown under moderate to high light intensity, PS I cyclic electron transport was nearly insensitive to methyl viologen, indicating that the cyclic electron supply to PS I derived almost exclusively from a thylakoid dehydrogenase. In cells grown under low light intensity, PS I cyclic electron transport was partially inhibited by methyl viologen, indicating that part of the cyclic electron supply to PS I derived directly from ferredoxin. It is proposed that the increased PSI cyclic electron transport observed in cells grown under high light intensity is a response to chronic photoinhibition.Abbreviations DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - ES energy storage - MV methyl viologen - PA_m photoacoustic thermal signal with strong non-modulated background light added - PA_s photoacoustic thermal signal without background light added CIW/DPB Publication No. 1205.     

Photoinhibition and Recovery of Photosynthesis in Canker-susceptible and Resistant Needles of Cypress (Cupressus sempervirens L.)

The aim of the present investigation was to test the hypothesis that the cypress canker caused by a fungus (Seiridium cardinale) infection induced effects on photosynthesis which could be related to photoinhibition and the process of recovery in susceptible and resistant needles. Photoinhibition of photosynthesis and recovery was studied in canker‐infected susceptible and resistant needles of cypress (Cupressus sempervirens L.) under controlled conditions (irradiation of detached needles to approximately 1900 μmol/m²/s). The degree of photoinhibition was determined by means of the ratio of variable to maximum chlorophyll (Chl) fluorescence (F_v/F_m) and electron transport measurements. The potential efficiency of photosystem (PS) II, F_v/F_m declined, and F_o increased significantly in canker‐susceptible needles, while F_o did not change in resistant needles. In isolated thylakoids, high light (HL) decreased the rate of whole chain and PS II activity markedly more in susceptible than in resistant needles. A smaller reduction of PS I activity was noticed only in susceptible needles. Upon subsequent dark incubation, fast recovery was noticed in both needle types and reached maximum rates of PS II efficiency similar to those noticed in non‐photoinhibited needles. The artificial exogenous electron donors such as diphenyl carbazide (DPC), NH₂OH and Mn²⁺ failed to restore the HL induced loss of PS II activity in susceptible needles, while DPC and NH₂OH significantly restored it in resistant needles. The results suggest that HL inactivates the donor side of PS II in resistant and the acceptor side of PS II in susceptible needles. The results on the quantification of the PS II reaction centre protein D1 and 33 kDa protein of water‐splitting complex following HL exposure showed pronounced differences between susceptible and resistant needles. The marked loss of PS II activity in HL‐irradiated needles was due to the marked loss of D1 protein in susceptible and 33 kDa protein in resistant needles, respectively.     

Phosphorylation of PS II polypeptides inhibits D1 protein-degradation and increases PS II stability

To study the significance of Photosystem (PS) II phosphorylation for the turnover of the D1 protein, phosphorylation was compared with the synthesis and content of the D1 protein in intact chloroplasts. As shown by radioactive labelling with [³²P_i] phosphorylation of PS II polypeptides was saturated at light intensities of 125 mol m^-2 s^-1. Under steady state conditions, in intact chloroplasts D1 protein, once it was phosphorylated, was neither dephosphorylated nor degraded in the light. D1 protein-synthesis was measured as incorporation of [¹⁴C] leucine. As shown by non-denaturing gel-electrophoresis followed by SDS-PAGE newly synthesised D1 protein was assembled to intact PS II-centres and no free D1 protein could be detected. D1 protein-synthesis was saturated at light intensities of 500 mol m^-2 s^-1. The content of D1 protein stayed stable even after illumination with 5000 mol m^-2 s^-1 showing that D1 protein-degradation was saturated at the same light intensities. The difference in the light saturation points of phosphorylation and of D1 protein-turnover indicates a complex regulation of D1 protein-turnover by phosphorylation. Separation of the phosphorylated and dephosphorylated D1 protein by LiDS-gelelectrophoresis combined with radioactive pulse-labelling with [¹⁴C] leucine and [³²P_i] revealed that D1 protein, synthesised under steady state conditions in the light, did not become phosphorylated but instead was rapidly degraded whereas the phosphorylated form of the D1 protein was not a good substrate for degradation. According to these observations phosphorylation of the D1 protein creates a pool of PS II centres which is not involved in D1 to these observations phosphorylation of the D1 protein creates a pool of PS II centres which is not involved in D1 protein-turnover. Fractionation of thylakoid membranes confirms that the phosphorylated, non-turning over pool of PS II-centres was located in the central regions of the grana, whereas PS II-centres involved in D1 protein-turnover were found exclusively in the stroma-lamellae and in the grana-margins.Abbreviations chl chlorophyll - F_v yield of variable fluorescence, difference between F_m, the maximal fluorescence yield at saturating light, when all reaction-centres are closed, and F_o, the fluorescence yield in the dark, when all reaction-centres are open - LHC light harvesting complex - PFD photon flux density - PS photosystem     

Changes in the antenna size of Photosystem I and Photosystem II in Synechococcus sp. strain PCC 7942 grown in the presence of SANDOZ 9785 — a Photosystem II inhibitor

SANDOZ 9785, also known as BASF 13.338, is a pyridazinone derivative that inhibits Photosystem II (PS II) activity leading to an imbalance in the rate of electron transport through the photosystems. Synechococcus sp. strain PCC 7942 cells grown in the presence of sublethal concentration of SANDOZ 9785 (SAN 9785) for 48 hours exhibited a 20% decrease in Chl a per cell. However, no changes were observed in the content of phycocyanin per cell, the size of the phycobilisomes or in the PS II:PS I ratio. From an estimate of PS II electron transport rate under varying light intensities and spectral qualities and analysis of room temperature Chl a fluorescence induction, it was deduced that growth of Synechococcus PCC 7942 in the presence of SAN 9785 leads to a redistribution of excitation energy in favour of PS II. Though the redistribution appears to be primarily caused by changes affecting the Chl a antenna of PS II, the extent of energetic coupling between phycobilisomes and PS II is also enhanced in SAN 9785 grown Synechococcus PCC 7942 cells. There was a reduction in the effective size of PS I antenna based on measurement of P700 photooxidation kinetics. These results indicate that when PS II is partially inhibited, the structure of photosynthetic apparatus alters to redistribute the excitation energy in favour of PS II so that the efficiency of utilization of light energy by the two photosystems is optimized. Our results suggest that under the conditions used, drastic structural changes are not essential for redistribution of excitation energy between the photosystems.Abbreviations APC Allophycocyanin - Chl a chlorophyll a - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCMU 3-(3,4-dichlorophyenyl)-1,1-dimethyl urea - DCIP 2,6-dichlorophenolindophenol - F_o fluorescence when all the reaction centres are open - f_m fluorescence yield when all the reaction centres are closed - F_v variable chlorophyll fluorescence - HEPES N-2-Hydroxyethylpiperazine-N-2-ethanesulphonic Acid - I₅₀ concentration that causes 50% inhibition in activity - MV methyl viologen - pBQ para benzoquinone - PBS phycobilisome - PC phycocyanin - PS I, PS II Photosystem I, Photosystem II - P700 reaction centre Chl a of PS I - SAN 9785 SANDOZ 9785 i.e. 4-chloro-5-dimethylamino-2-phenyl-3 (2H) pyridazinone, also known as BASF 13.338     

Genetic engineering of thylakoid protein complexes by chloroplast transformation in Chlamydomonas reinhardtii

Chloroplast transformation of Chlamydomonas reinhardtii has developed into a powerful tool for studying the structure, function and assembly of thylakoid protein complexes in a eukaryotic organism. In this article we review the progress that is being made in the development of procedures for efficient chloroplast transformation. This focuses on the development of selectable markers and the use of Chlamydomonas mutants, individually lacking thylakoid protein complexes, as recipients. Chloroplast transformation has now been used to engineer all four major thylakoid protein complexes, photosystem II, photosystem I, cytochrome b ₆/f and ATP synthase. These results are discussed with an emphasis on new insights into assembly and function of these complexes in chloroplasts as compared with their prokaryotic counterparts.Abbreviations ENDOR electron nuclear double resonance - ESEEM electron spin echo envelope modulation - LHC light harvesting complex - PSI Photosystem I - PS II Photosystem II - P₆₈₀ primary electron donor in PS II - P₇₀₀ primary electron donor in PS I     

State 1-State 2 transitions in the cyanobacterium Synechococcus 6301 are controlled by the redox state of electron carriers between Photosystems I and II

The mechanism by which state 1-state 2 transitions in the cyanobacterium Synechococcus 6301 are controlled was investigated by examining the effects of a variety of chemical and illumination treatments which modify the redox state of the plastoquinone pool. The extent to which these treatments modify excitation energy distribution was determined by 77K fluorescence emission spectroscopy. It was found that treatment which lead to the oxidation of the plastoquinone pool induce a shift towards state 1 whereas treatments which lead to the reduction of the plastoquinone pool induce a shift towards state 2. We therefore propose that state transitions in cyanobacteria are triggered by changes in the redox state of plastoquinone or a closely associated electron carrier. Alternative proposals have included control by the extent of cyclic electron transport around PS I and control by localised electrochemical gradients around PS I and PS II. Neither of these proposals is consistent with the results reported here.Abbreviations DBMIB 2,5-dibromo-3methyl-6-isopropyl-p-benzoquinone - Chl chlorophyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DQH₂ duroquinol (tetramethyl-p-hydroquinone) - LHC II light-harvesting chlorophyll a/b-binding protein of PS II - Light 1 light predominantly exciting PS I - Light 2 light predominantly exciting PS II - M.V. methyl viologen - PS photosystem     

The role of chloroplast-encoded protein biosynthesis on the rate of D1 protein degradation in Dunaliella salina

Mechanistic aspects of the Photosystem II (PS II) damage and repair cycle in Dunaliella salina were investigated. The work addressed the role of chloroplast-encoded protein biosynthesis on the rate of the D1 protein (chloroplast psbA gene product) degradation, following photoinhibition of PS II under in vivo conditions. Cells were grown under different light-intensities and the rate of D1 photodamage and degradation was measured via pulse-chase measurements with (³⁵S)sulfate. It is shown that no detectable difference exists in the rate of D1 degradation in D. salina, measured in the presence or absence of lincomycin, a chloroplast protein biosynthesis inhibitor. The results suggest that de novo D1 biosynthesis does not play a role in the regulation of D1 degradation. In low-light (100 mol photons m^–2 s^–1) grown cells, the rate of photodamage to D1 did not exceed the rate of its degradation and replacement. In high-light (2200 mol photons m^–1 s^–1) grown cells, the rate of D1 photodamage was faster than the rate of its degradation, resulting in a significant accumulation of photoinactivated PS II centers in the chloroplast thylakoids (chronic photoinhibition). The latter was coincident with the appearance of a 160 kD complex that contained photodamaged D1. Electron micrographs of D. salina thylakoids revealed extensive grana stacks in the thylakoid membrane of low-light grown cells. Only rudimentary appressions consisting of simple membrane pairings were found in the high-light grown cells. The results are discussed in terms of the regulation of D1 degradation in chloroplasts under in vivo conditions.Abbreviations Chl chlorophyll - D1 the 32 kD reaction center protein of PS II, encoded by the chloroplast psbA gene - D2 the 34 kD reaction center protein of PS II, encoded by the chloroplast psbD gene - HL high light - LL low light - Linc lincomycin