The growth of plants, algae, and cyanobacteria relies on the catalytic activity of the oxygen-evolving PSII complex, which uses solar energy to extract electrons from water to feed into the photosynthetic electron transport chain. PSII is proving to be an excellent system to study how large multi-subunit membrane-protein complexes are assembled in the thylakoid membrane and subsequently repaired in response to photooxidative damage. Here we summarize recent developments in understanding the biogenesis of PSII, with an emphasis on recent insights obtained from biochemical and structural analysis of cyanobacterial PSII assembly/repair intermediates. We also discuss how chlorophyll synthesis is synchronized with protein synthesis and suggest a possible role for PSI in PSII assembly. Special attention is paid to unresolved and controversial issues that could be addressed in future research.

• MeSH
• chlorofyl metabolismus   MeSH
• fotosyntéza MeSH
• fotosystém II - proteinový komplex * metabolismus   MeSH
• sinice * metabolismus   MeSH
• tylakoidy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• chlorofyl MeSH
• fotosystém II - proteinový komplex * MeSH

FtsH proteases are membrane-embedded proteolytic complexes important for protein quality control and regulation of various physiological processes in bacteria, mitochondria, and chloroplasts. Like most cyanobacteria, the model species Synechocystis sp. PCC 6803 contains four FtsH homologs, FtsH1-FtsH4. FtsH1-FtsH3 form two hetero-oligomeric complexes, FtsH1/3 and FtsH2/3, which play a pivotal role in acclimation to nutrient deficiency and photosystem II quality control, respectively. FtsH4 differs from the other three homologs by the formation of a homo-oligomeric complex, and together with Arabidopsis thaliana AtFtsH7/9 orthologs, it has been assigned to another phylogenetic group of unknown function. Our results exclude the possibility that Synechocystis FtsH4 structurally or functionally substitutes for the missing or non-functional FtsH2 subunit in the FtsH2/3 complex. Instead, we demonstrate that FtsH4 is involved in the biogenesis of photosystem II by dual regulation of high light-inducible proteins (Hlips). FtsH4 positively regulates expression of Hlips shortly after high light exposure but is also responsible for Hlip removal under conditions when their elevated levels are no longer needed. We provide experimental support for Hlips as proteolytic substrates of FtsH4. Fluorescent labeling of FtsH4 enabled us to assess its localization using advanced microscopic techniques. Results show that FtsH4 complexes are concentrated in well-defined membrane regions at the inner and outer periphery of the thylakoid system. Based on the identification of proteins that co-purified with the tagged FtsH4, we speculate that FtsH4 concentrates in special compartments in which the biogenesis of photosynthetic complexes takes place.

• Klíčová slova
• FtsH4, high light-inducible protein, photosystem II biogenesis, proteolysis, thylakoid,
• MeSH
• Arabidopsis * genetika   metabolismus   MeSH
• chloroplasty metabolismus   MeSH
• fotosystém II - proteinový komplex genetika   metabolismus   MeSH
• fylogeneze MeSH
• metaloproteasy genetika   metabolismus   MeSH
• proteasy MeSH
• proteiny huseníčku * genetika   metabolismus   MeSH
• Synechocystis * genetika   metabolismus   MeSH
• tylakoidy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fotosystém II - proteinový komplex MeSH
• FtsH4 protein, Arabidopsis MeSH Prohlížeč
• metaloproteasy MeSH
• proteasy MeSH
• proteiny huseníčku * MeSH

The repair of photosystem II is a key mechanism that keeps the light reactions of oxygenic photosynthesis functional. During this process, the PSII central subunit D1 is replaced with a newly synthesized copy while the neighbouring CP43 antenna with adjacent small subunits (CP43 module) is transiently detached. When the D2 protein is also damaged, it is degraded together with D1 leaving both the CP43 module and the second PSII antenna module CP47 unassembled. In the cyanobacterium Synechocystis sp. PCC 6803, the released CP43 and CP47 modules have been recently suggested to form a so-called no reaction centre complex (NRC). However, the data supporting the presence of NRC can also be interpreted as a co-migration of CP43 and CP47 modules during electrophoresis and ultracentrifugation without forming a mutual complex. To address the existence of NRC, we analysed Synechocystis PSII mutants accumulating one or both unassembled antenna modules as well as Synechocystis wild-type cells stressed with high light. The obtained results were not compatible with the existence of a stable NRC since each unassembled module was present as a separate protein complex with a mutually similar electrophoretic mobility regardless of the presence of the second module. The non-existence of NRC was further supported by isolation of the His-tagged CP43 and CP47 modules from strains lacking either D1 or D2 and their migration patterns on native gels.

• Klíčová slova
• CP43, CP47, No reaction centre complex, Photosynthesis, Photosystem II,
• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• fotosystém II - proteinový komplex metabolismus   MeSH
• kyslík metabolismus   MeSH
• Synechocystis * genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• fotosystém II - proteinový komplex MeSH
• kyslík MeSH

Photosystem II (PSII) is the multi-subunit light-driven oxidoreductase that drives photosynthetic electron transport using electrons extracted from water. To investigate the initial steps of PSII assembly, we used strains of the cyanobacterium Synechocystis sp. PCC 6803 arrested at early stages of PSII biogenesis and expressing affinity-tagged PSII subunits to isolate PSII reaction center assembly (RCII) complexes and their precursor D1 and D2 modules (D1mod and D2mod). RCII preparations isolated using either a His-tagged D2 or a FLAG-tagged PsbI subunit contained the previously described RCIIa and RCII* complexes that differ with respect to the presence of the Ycf39 assembly factor and high light-inducible proteins (Hlips) and a larger complex consisting of RCIIa bound to monomeric PSI. All RCII complexes contained the PSII subunits D1, D2, PsbI, PsbE, and PsbF and the assembly factors rubredoxin A and Ycf48, but we also detected PsbN, Slr1470, and the Slr0575 proteins, which all have plant homologs. The RCII preparations also contained prohibitins/stomatins (Phbs) of unknown function and FtsH protease subunits. RCII complexes were active in light-induced primary charge separation and bound chlorophylls (Chls), pheophytins, beta-carotenes, and heme. The isolated D1mod consisted of D1/PsbI/Ycf48 with some Ycf39 and Phb3, while D2mod contained D2/cytochrome b559 with co-purifying PsbY, Phb1, Phb3, FtsH2/FtsH3, CyanoP, and Slr1470. As stably bound, Chl was detected in D1mod but not D2mod, formation of RCII appears to be important for stable binding of most of the Chls and both pheophytins. We suggest that Chl can be delivered to RCII from either monomeric Photosystem I or Ycf39/Hlips complexes.

• MeSH
• chlorofyl metabolismus   MeSH
• feofytiny metabolismus   MeSH
• fotosystém I - proteinový komplex metabolismus   MeSH
• fotosystém II - proteinový komplex * metabolismus   MeSH
• Synechocystis * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chlorofyl MeSH
• feofytiny MeSH
• fotosystém I - proteinový komplex MeSH
• fotosystém II - proteinový komplex * MeSH

Assembly of photosystem II (PSII), a water-splitting catalyst in chloroplasts and cyanobacteria, requires numerous auxiliary proteins which promote individual steps of this sequential process and transiently associate with one or more assembly intermediate complexes. In this study, we focussed on the role of a PSII-associated protein encoded by the ssl1498 gene in the cyanobacterium Synechocystis sp. PCC 6803. The N-terminal domain of this protein, which is here called Psb34, is very similar to the N-terminus of HliA/B proteins belonging to a family of high-light-inducible proteins (Hlips). Psb34 was identified in both dimeric and monomeric PSII, as well as in a PSII monomer lacking CP43 and containing Psb28. When FLAG-tagged, the protein is co-purified with these three complexes and with the PSII auxiliary proteins Psb27 and Psb28. However, the preparation also contained the oxygen-evolving enhancers PsbO and PsbV and lacked HliA/B proteins even when isolated from high-light-treated cells. The data suggest that Psb34 competes with HliA/B for the same binding site and that it is one of the components involved in the final conversion of late PSII assembly intermediates into functional PSII complexes, possibly keeping them free of Hlips. Unlike HliA/B, Psb34 does bind to the CP47 assembly module before its incorporation into PSII. Analysis of strains lacking Psb34 indicates that Psb34 mediates the optimal equilibrium of HliA/B binding among individual PSII assembly intermediates containing CP47, allowing Hlip-mediated photoprotection at all stages of PSII assembly.

• Klíčová slova
• CP47, High-light-inducible protein, Photosynthesis, Photosystem II,
• MeSH
• bakteriální proteiny metabolismus   MeSH
• fotosyntéza MeSH
• fotosystém II - proteinový komplex metabolismus   MeSH
• protein TNFSF14 metabolismus   MeSH
• Synechocystis * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• fotosystém II - proteinový komplex MeSH
• protein TNFSF14 MeSH

Cytochrome (Cyt) b559 is a key component of the photosystem II complex (PSII) that is essential for its proper functioning and assembly. Site-directed mutants of the model cyanobacterium Synechocystis sp. PCC6803 with mutated heme axial ligands of Cyt b559 have little PSII and are therefore unable to grow photoautotrophically. Here we describe two types of Synechocystis autotrophic transformants that retained the same mutations in Cyt b559 but are able to accumulate PSII and grow photoautotrophically. Whole-genome sequencing revealed that all of these autotrophic transformants carried a variable number of tandem repeats (from 5 to 15) of chromosomal segments containing the psbEFLJ operon. RNA-seq analysis showed greatly increased transcript levels of the psbEFLJ operon in these autotrophic transformants. Multiple copies of the psbEFLJ operon in these transformants were only maintained during autotrophic growth, while its copy numbers gradually decreased under photoheterotrophic conditions. Two-dimensional PAGE analysis of membrane proteins revealed a strong deficiency in PSII complexes in the Cyt b559 mutants that was reversed in the autotrophic transformants. These results illustrate how tandem gene amplification restores PSII accumulation and photoautotrophic growth in Cyt b559 mutants of cyanobacteria, and may serve as an important adaptive mechanism for cyanobacterial survival.

• Klíčová slova
• cyanobacterium, cytochrome b559, photosynthesis, photosystem II (PSII), tandem gene amplification,
• MeSH
• amplifikace genu MeSH
• cytochromy b genetika   metabolismus   MeSH
• cytochromy typu b genetika   metabolismus   MeSH
• fotosystém II - proteinový komplex * genetika   metabolismus   MeSH
• Synechocystis * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytochromy b MeSH
• cytochromy typu b MeSH
• fotosystém II - proteinový komplex * MeSH

Photochemical energy conversion during oxygenic photosynthesis is performed by membrane-embedded chlorophyll-binding protein complexes. The biogenesis and maintenance of these complexes requires auxiliary protein factors that optimize the assembly process and protect nascent complexes from photodamage. In cyanobacteria, several lipoproteins contribute to the biogenesis and function of the photosystem II (PSII) complex. They include CyanoP, CyanoQ, and Psb27, which are all attached to the lumenal side of PSII complexes. Here, we show that the lumenal Ycf48 assembly factor found in the cyanobacterium Synechocystis sp. PCC 6803 is also a lipoprotein. Detailed mass spectrometric analysis of the isolated protein supported by site-directed mutagenesis experiments indicates lipidation of the N-terminal C29 residue of Ycf48 and removal of three amino acids from the C-terminus. The lipobox sequence in Ycf48 contains a cysteine residue at the -3 position compared to Leu/Val/Ile residues found in the canonical lipobox sequence. The atypical Ycf48 lipobox sequence is present in most cyanobacteria but is absent in eukaryotes. A possible role for lipoproteins in the coordinated assembly of cyanobacterial PSII is discussed.

• Klíčová slova
• chlorophyll-binding proteins, photosynthesis, photosystem II,
• MeSH
• bakteriální proteiny metabolismus   MeSH
• fotosystém II - proteinový komplex metabolismus   MeSH
• metabolismus lipidů * MeSH
• Synechocystis metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• fotosystém II - proteinový komplex MeSH

Biological membranes were originally described as a fluid mosaic with uniform distribution of proteins and lipids. Later, heterogeneous membrane areas were found in many membrane systems including cyanobacterial thylakoids. In fact, cyanobacterial pigment-protein complexes (photosystems, phycobilisomes) form a heterogeneous mosaic of thylakoid membrane microdomains (MDs) restricting protein mobility. The trafficking of membrane proteins is one of the key factors for long-term survival under stress conditions, for instance during exposure to photoinhibitory light conditions. However, the mobility of unbound 'free' proteins in thylakoid membrane is poorly characterized. In this work, we assessed the maximal diffusional ability of a small, unbound thylakoid membrane protein by semi-single molecule FCS (fluorescence correlation spectroscopy) method in the cyanobacterium Synechocystis sp. PCC6803. We utilized a GFP-tagged variant of the cytochrome b6f subunit PetC1 (PetC1-GFP), which was not assembled in the b6f complex due to the presence of the tag. Subsequent FCS measurements have identified a very fast diffusion of the PetC1-GFP protein in the thylakoid membrane (D = 0.14 - 2.95 µm2s-1). This means that the mobility of PetC1-GFP was comparable with that of free lipids and was 50-500 times higher in comparison to the mobility of proteins (e.g., IsiA, LHCII-light-harvesting complexes of PSII) naturally associated with larger thylakoid membrane complexes like photosystems. Our results thus demonstrate the ability of free thylakoid-membrane proteins to move very fast, revealing the crucial role of protein-protein interactions in the mobility restrictions for large thylakoid protein complexes.

• Klíčová slova
• FCS, cyanobacteria, photosynthesis, proteins mobility, thylakoids,
• Publikační typ
• časopisecké články MeSH

Oxygenic photosynthesis relies on accessory factors to promote the assembly and maintenance of the photosynthetic apparatus in the thylakoid membranes. The highly conserved membrane-bound rubredoxin-like protein RubA has previously been implicated in the accumulation of both PSI and PSII, but its mode of action remains unclear. Here, we show that RubA in the cyanobacterium Synechocystis sp PCC 6803 is required for photoautotrophic growth in fluctuating light and acts early in PSII biogenesis by promoting the formation of the heterodimeric D1/D2 reaction center complex, the site of primary photochemistry. We find that RubA, like the accessory factor Ycf48, is a component of the initial D1 assembly module as well as larger PSII assembly intermediates and that the redox-responsive rubredoxin-like domain is located on the cytoplasmic surface of PSII complexes. Fusion of RubA to Ycf48 still permits normal PSII assembly, suggesting a spatiotemporal proximity of both proteins during their action. RubA is also important for the accumulation of PSI, but this is an indirect effect stemming from the downregulation of light-dependent chlorophyll biosynthesis induced by PSII deficiency. Overall, our data support the involvement of RubA in the redox control of PSII biogenesis.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• biologické pigmenty izolace a purifikace   MeSH
• chlorofyl biosyntéza   MeSH
• fotosyntéza fyziologie   MeSH
• fotosystém I - proteinový komplex metabolismus   MeSH
• fotosystém II - proteinový komplex metabolismus   MeSH
• mutace MeSH
• rubredoxiny chemie   genetika   metabolismus   MeSH
• Synechocystis genetika   růst a vývoj   metabolismus   MeSH
• tylakoidy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• biologické pigmenty MeSH
• chlorofyl MeSH
• fotosystém I - proteinový komplex MeSH
• fotosystém II - proteinový komplex MeSH
• rubredoxiny MeSH

Robust photosynthesis in chloroplasts and cyanobacteria requires the participation of accessory proteins to facilitate the assembly and maintenance of the photosynthetic apparatus located within the thylakoid membranes. The highly conserved Ycf48 protein acts early in the biogenesis of the oxygen-evolving photosystem II (PSII) complex by binding to newly synthesized precursor D1 subunit and by promoting efficient association with the D2 protein to form a PSII reaction center (PSII RC) assembly intermediate. Ycf48 is also required for efficient replacement of damaged D1 during the repair of PSII. However, the structural features underpinning Ycf48 function remain unclear. Here we show that Ycf48 proteins encoded by the thermophilic cyanobacterium Thermosynechococcus elongatus and the red alga Cyanidioschyzon merolae form seven-bladed beta-propellers with the 19-aa insertion characteristic of eukaryotic Ycf48 located at the junction of blades 3 and 4. Knowledge of these structures has allowed us to identify a conserved "Arg patch" on the surface of Ycf48 that is important for binding of Ycf48 to PSII RCs but also to larger complexes, including trimeric photosystem I (PSI). Reduced accumulation of chlorophyll in the absence of Ycf48 and the association of Ycf48 with PSI provide evidence of a more wide-ranging role for Ycf48 in the biogenesis of the photosynthetic apparatus than previously thought. Copurification of Ycf48 with the cyanobacterial YidC protein insertase supports the involvement of Ycf48 during the cotranslational insertion of chlorophyll-binding apopolypeptides into the membrane.

• Klíčová slova
• chlorophyll-binding proteins, photosynthesis, photosystem II,
• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• fotosystém I - proteinový komplex biosyntéza   genetika   MeSH
• fotosystém II - proteinový komplex biosyntéza   genetika   MeSH
• sinice genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• fotosystém I - proteinový komplex MeSH
• fotosystém II - proteinový komplex MeSH