Cyanobacterial psbA gene family: optimization of oxygenic photosynthesis

The D1 protein of Photosystem II (PSII), encoded by the psbA genes, is an indispensable component of oxygenic photosynthesis. Due to strongly oxidative chemistry of PSII water splitting, the D1 protein is prone to constant photodamage requiring its replacement, whereas most of the other PSII subunits remain ordinarily undamaged. In cyanobacteria, the D1 protein is encoded by a psbA gene family, whose members are differentially expressed according to environmental cues. Here, the regulation of the psbA gene expression is first discussed with emphasis on the model organisms Synechococcus sp. and Synechocystis sp. Then, a general classification of cyanobacterial D1 isoforms in various cyanobacterial species into D1m, D1:1, D1:2, and D1′ forms depending on their expression pattern under acclimated growth conditions and upon stress is discussed, taking into consideration the phototolerance of different D1 forms and the expression conditions of respective members of the psbA gene family

Regulatory subunit B'gamma of protein phosphatase 2A prevents unnecessary defense reactions under low light in Arabidopsis

Peer reviewe

Post-translational Modifications in Regulation of Chloroplast Function: Recent Advances

Post-translational modifications (PTMs) of proteins enable fast modulation of protein function in response to metabolic and environmental changes. Phosphorylation is known to play a major role in regulating distribution of light energy between the Photosystems (PS) I and II (state transitions) and in PSII repair cycle. In addition, thioredoxin-mediated redox regulation of Calvin cycle enzymes has been shown to determine the efficiency of carbon assimilation. Besides these well characterized modifications, recent methodological progress has enabled identification of numerous other types of PTMs in various plant compartments, including chloroplasts. To date, at least N-terminal and Lys acetylation, Lys methylation, Tyr nitration and S-nitrosylation, glutathionylation, sumoylation and glycosylation of chloroplast proteins have been described. These modifications impact DNA replication, control transcriptional efficiency, regulate translational machinery and affect metabolic activities within the chloroplast. Moreover, light reactions of photosynthesis as well as carbon assimilation are regulated at multiple levels by a number of PTMs. It is likely that future studies will reveal new metabolic pathways to be regulated by PTMs as well as detailed molecular mechanisms of PTM-mediated regulation

Gel-based proteomic map of Arabidopsis thaliana root plastids and mitochondria

Background Non-photosynthetic plastids of plants are known to be involved in a range of metabolic and biosynthetic reactions, even if they have been difficult to study due to their small size and lack of color. The morphology of root plastids is heterogeneous and also the plastid size, density and subcellular distribution varies depending on the cell type and developmental stage, and therefore the functional features have remained obscure. Although the root plastid proteome is likely to reveal specific functional features,Arabidopsis thalianaroot plastid proteome has not been studied to date. Results In the present study, we separated Arabidopsis root protein fraction enriched with plastids and mitochondria by 2D-PAGE and identified 84 plastid-targeted and 77 mitochondrion-targeted proteins using LC-MS/MS. The most prevalent root plastid protein categories represented amino acid biosynthesis, carbohydrate metabolism and lipid biosynthesis pathways, while the enzymes involved in starch and sucrose metabolism were not detected. Mitochondrion-targeted proteins were classified mainly into the energetics category. Conclusions This is the first study presenting gel-based map ofArabidopsis thalianaroot plastid and mitochondrial proteome. Our findings suggest that Arabidopsis root plastids have broad biosynthetic capacity, and that they do not play a major role in a long-term storage of carbohydrates. The proteomic map provides a tool for further studies to compare changes in the proteome, e.g. in response to environmental cues, and emphasizes the role of root plastids in nitrogen and sulfur metabolism as well as in amino acid and fatty acid biosynthesis. The results enable taking a first step towards an integrated view of root plastid/mitochondrial proteome and metabolic functions inArabidopsis thalianaroots

Chloroplast Acetyltransferase GNAT2 is Involved in the Organization and Dynamics of Thylakoid Structure

Higher plants acclimate to changes in light conditions by adjusting the thylakoid membrane ultrastructure. Additionally, excitation energy transfer between photosystem II (PSII) and photosystem I (PSI) is balanced in a process known as state transition. These modifications are mediated by reversible phosphorylation of Lhcb1 and Lhcb2 proteins in different pools of light-harvesting complex (LHCII) trimers. Our recent study demonstrated that chloroplast acetyltransferase NUCLEAR SHUTTLE INTERACTING (NSI)/GNAT2 (general control non-repressible 5 (GCN5)-related N-acetyltransferase 2) is also needed for the regulation of light harvesting, evidenced by the inability of the gnat2 mutant to perform state transitions although there are no defects in LHCII phosphorylation. Here, we show that despite contrasting phosphorylation states of LHCII, grana packing in the gnat2 and state transition 7 (stn7) mutants possesses similar features, as the thylakoid structure of the mutants does not respond to the shift from darkness to light, which is in striking contrast to wild type (Wt). Circular dichroism and native polyacrylamide gel electrophoresis analyses further revealed that the thylakoid protein complex organization of gnat2 and stn7 resembles each other, but differ from that of Wt. Also, the location of the phosphorylated Lhcb2 as well as the LHCII antenna within the thylakoid network in gnat2 mutant is different from that of Wt. In gnat2, the LHCII antenna remains largely in grana stacks, where the phosphorylated Lhcb2 is found in all LHCII trimer pools, including those associated with PSII. These results indicate that in addition to phosphorylation-mediated regulation through STN7, the GNAT2 enzyme is involved in the organization and dynamics of thylakoid structure, probably through the regulation of chloroplast protein acetylation.</p

Root-type ferredoxin-NADP(+) oxidoreductase isoforms in Arabidopsis thaliana : Expression patterns, location and stress responses

In Arabidopsis, two leaf-type ferredoxin-NADP(+) oxidoreductase (LFNR) isoforms function in photosynthetic electron flow in reduction of NADP(+), while two root-type FNR (RFNR) isoforms catalyse reduction of ferredoxin in non-photosynthetic plastids. As the key to understanding, the function of RFNRs might lie in their spatial and temporal distribution in different plant tissues and cell types, we examined expression of RFNR1 and RFNR2 genes using beta-glucuronidase (GUS) reporter lines and investigated accumulation of distinct RFNR isoforms using a GFP approach and Western blotting upon various stresses. We show that while RFNR1 promoter is active in leaf veins, root tips and in the stele of roots, RFNR2 promoter activity is present in leaf tips and root stele, epidermis and cortex. RFNR1 protein accumulates as a soluble protein within the plastids of root stele cells, while RFNR2 is mainly present in the outer root layers. Ozone treatment of plants enhanced accumulation of RFNR1, whereas low temperature treatment specifically affected RFNR2 accumulation in roots. We further discuss the physiological roles of RFNR1 and RFNR2 based on characterization of rfnr1 and rfnr2 knock-out plants and show that although the function of these proteins is partly redundant, the RFNR proteins are essential for plant development and survival.Peer reviewe

Gene expression and organization of thylakoid protein complexes in the PSII-less mutant of Synechocystis sp. PCC 6803

Photosystems I and II (PSI arid PSII) are the integral components of the photosynthetic electron transport chain that utilize light to provide chemical energy for CO2 fixation. In this study, we investigated how the deficiency of PSII affects the gene expression, accumulation, and organization of thylakoid protein complexes as well as physiological characteristics of Synechocystis sp. PCC 6803 by combining biochemical, biophysical, and transcriptomic approaches. RNA-seq analysis showed upregulated expression of genes encoding the PSII core proteins, and downregulation of genes associated with interaction between light-harvesting phycobilisomes and PSI. Two-dimensional separation of thylakoid protein complexes confirmed the lack of PSII complexes, yet unassembled PSII subunits were detected. The content of PsaB representing PSI was lower, while the content of cytochrome b(6)f complexes was higher in the PSII-less strain as compared with control (CS). Application of oxygraph measurements revealed higher rates of dark respiration and lower PSI activity in the mutant. The latter likely resulted from the detected decrease in the accumulation of PSI, PSI monomerization, increased proportion of energetically decoupled phycobilisomes in PSII-less cultures, and low abundance of phycocyanin. Merging the functional consequences of PSII depletion with differential protein and transcript accumulation in the mutant, in comparison to CS, identified signal transduction from the photosynthetic apparatus to the genome level.</p

RETRACTED ARTICLE:The Arabidopsis NOT4A E3 ligase promotes PGR3 expression and regulates chloroplast translation

Chloroplast function requires the coordinated action of nuclear- and chloroplast-derived proteins, including several hundred nuclear-encoded pentatricopeptide repeat (PPR) proteins that regulate plastid mRNA metabolism. Despite their large number and importance, regulatory mechanisms controlling PPR expression are poorly understood. Here we show that the Arabidopsis NOT4A ubiquitin-ligase positively regulates the expression of PROTON GRADIENT REGULATION 3 (PGR3), a PPR protein required for translating several thylakoid-localised photosynthetic components and ribosome subunits within chloroplasts. Loss of NOT4A function leads to a strong depletion of cytochrome b6f and NAD(P)H dehydrogenase (NDH) complexes, as well as plastid 30 S ribosomes, which reduces mRNA translation and photosynthetic capacity, causing pale-yellow and slow-growth phenotypes. Quantitative transcriptome and proteome analysis of the not4a mutant reveal it lacks PGR3 expression, and that its molecular defects resemble those of a pgr3 mutant. Furthermore, we show that normal plastid function is restored to not4a through transgenic PGR3 expression. Our work identifies NOT4A as crucial for ensuring robust photosynthetic function during development and stress-response, through promoting PGR3 production and chloroplast translation.</p

Root-type ferredoxin-NADP(+) oxidoreductase isoforms in Arabidopsis thaliana: Expression patterns, location and stress responses

In Arabidopsis, two leaf-type ferredoxin-NADP(+) oxidoreductase (LFNR) isoforms function in photosynthetic electron flow in reduction of NADP(+), while two root-type FNR (RFNR) isoforms catalyse reduction of ferredoxin in non-photosynthetic plastids. As the key to understanding, the function of RFNRs might lie in their spatial and temporal distribution in different plant tissues and cell types, we examined expression of RFNR1 and RFNR2 genes using beta-glucuronidase (GUS) reporter lines and investigated accumulation of distinct RFNR isoforms using a GFP approach and Western blotting upon various stresses. We show that while RFNR1 promoter is active in leaf veins, root tips and in the stele of roots, RFNR2 promoter activity is present in leaf tips and root stele, epidermis and cortex. RFNR1 protein accumulates as a soluble protein within the plastids of root stele cells, while RFNR2 is mainly present in the outer root layers. Ozone treatment of plants enhanced accumulation of RFNR1, whereas low temperature treatment specifically affected RFNR2 accumulation in roots. We further discuss the physiological roles of RFNR1 and RFNR2 based on characterization of rfnr1 and rfnr2 knock-out plants and show that although the function of these proteins is partly redundant, the RFNR proteins are essential for plant development and survival.</p

Comparative analysis of thylakoid protein complexes in state transition mutants nsi and stn7: focus on PSI and LHCII

The photosynthetic machinery of plants can acclimate to changes in light conditions by balancing light-harvesting between the two photosystems (PS). This acclimation response is induced by the change in the redox state of the plastoquinone pool, which triggers state transitions through activation of the STN7 kinase and subsequent phosphorylation of light-harvesting complex II (LHCII) proteins. Phosphorylation of LHCII results in its association with PSI (state 2), whereas dephosphorylation restores energy allocation to PSII (state 1). In addition to state transition regulation by phosphorylation, we have recently discovered that plants lacking the chloroplast acetyltransferase NSI are also locked in state 1, even though they possess normal LHCII phosphorylation. This defect may result from decreased lysine acetylation of several chloroplast proteins. Here, we compared the composition of wild type (wt), stn7 and nsi thylakoid protein complexes involved in state transitions separated by Blue Native gel electrophoresis. Protein complex composition and relative protein abundances were determined by LC–MS/MS analyses using iBAQ quantification. We show that despite obvious mechanistic differences leading to defects in state transitions, no major differences were detected in the composition of PSI and LHCII between the mutants. Moreover, both stn7 and nsi plants show retarded growth and decreased PSII capacity under fluctuating light as compared to wt, while the induction of non-photochemical quenching under fluctuating light was much lower in both nsi mutants than in stn7.</p