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1.
Cell ; 184(14): 3643-3659.e23, 2021 07 08.
Artículo en Inglés | MEDLINE | ID: mdl-34166613

RESUMEN

Vesicle-inducing protein in plastids 1 (VIPP1) is essential for the biogenesis and maintenance of thylakoid membranes, which transform light into life. However, it is unknown how VIPP1 performs its vital membrane-remodeling functions. Here, we use cryo-electron microscopy to determine structures of cyanobacterial VIPP1 rings, revealing how VIPP1 monomers flex and interweave to form basket-like assemblies of different symmetries. Three VIPP1 monomers together coordinate a non-canonical nucleotide binding pocket on one end of the ring. Inside the ring's lumen, amphipathic helices from each monomer align to form large hydrophobic columns, enabling VIPP1 to bind and curve membranes. In vivo mutations in these hydrophobic surfaces cause extreme thylakoid swelling under high light, indicating an essential role of VIPP1 lipid binding in resisting stress-induced damage. Using cryo-correlative light and electron microscopy (cryo-CLEM), we observe oligomeric VIPP1 coats encapsulating membrane tubules within the Chlamydomonas chloroplast. Our work provides a structural foundation for understanding how VIPP1 directs thylakoid biogenesis and maintenance.


Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Chlamydomonas/metabolismo , Multimerización de Proteína , Synechocystis/metabolismo , Tilacoides/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/ultraestructura , Sitios de Unión , Membrana Celular/metabolismo , Chlamydomonas/ultraestructura , Microscopía por Crioelectrón , Proteínas Fluorescentes Verdes/metabolismo , Interacciones Hidrofóbicas e Hidrofílicas , Luz , Lípidos/química , Modelos Moleculares , Nucleótidos/metabolismo , Unión Proteica , Estructura Secundaria de Proteína , Estrés Fisiológico/efectos de la radiación , Synechocystis/ultraestructura , Tilacoides/ultraestructura
2.
J Biol Chem ; : 107716, 2024 Aug 22.
Artículo en Inglés | MEDLINE | ID: mdl-39181331

RESUMEN

The low CO2-inducible NDH complex (NDH-1MS) plays a crucial role in the cyanobacterial CO2-concentrating mechanism (CCM). However, the components in this complex and the regulation mechanism are still not completely understood. Using a mutant only with NDH-1MS as active Ci sequestration system, we identified a functional gene sll1736 named as cupAR (CupA Regulator). The cupAR deletion mutant, ΔcupAR, grew faster than the wild type (WT) under high CO2 (HC) condition, more evidently at low pH. The activities of O2 evolution, CO2 uptake,NDH-1 and the building up of a trans-thylakoid proton were stimulated in this mutant under HC conditions. The cupAR gene is co-transcribed with the NDH-1S operon (ndhF3-ndhD3-cupA) and encoded protein which specifically suppresses the transcription level of this operon under HC conditions. Mutation of cupAR significantly up-regulated the accumulation of CupA, the key protein of NDH-1MS, under HC condition. CupAR interacted with NdhD3 and NdhF3, the membrane components of NDH-1MS, while accumulation of CupAR was reduced in the ΔndhD3 mutant. Furthermore, CupAR was co-located with CupA in both NDH-1MS complex and NDH-1S subcomplex. On the other hand, deletion of ndhR, a negative regulator of the NDH-1S operon increased the accumulation of CupAR, while deletion of cupAR significantly lowered NdhR. Based on these results, we concluded that CupAR is a novel subunit of NDH-1MS, negatively regulating the activities of CupA and CO2 uptake dependent on NDH-1MS by positive regulation of NdhR under enriched CO2 conditions.

3.
Plant J ; 118(4): 1207-1217, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38319793

RESUMEN

CpcL-phycobilisomes (CpcL-PBSs) are a reduced type of phycobilisome (PBS) found in several cyanobacteria. They lack the traditional PBS terminal energy emitters, but still show the characteristic red-shifted fluorescence at ~670 nm. We established a method of assembling in vitro a rod-membrane linker protein, CpcL, with phycocyanin, generating complexes with the red-shifted spectral features of CpcL-PBSs. The red-shift arises from the interaction of a conserved key glutamine, Q57 of CpcL in Synechocystis sp. PCC 6803, with a single phycocyanobilin chromophore of trimeric phycocyanin at one of the three ß82-sites. This chromophore is the terminal energy acceptor of CpcL-PBSs and donor to the photosystem(s). This mechanism also operates in PBSs from Acaryochloris marina MBIC11017. We then generated multichromic complexes harvesting light over nearly the complete visible range via the replacement of phycocyanobilin chromophores at sites α84 and ß153 of phycocyanins by phycoerythrobilin and/or phycourobilin. The results demonstrate the rational design of biliprotein-based light-harvesting elements by engineering CpcL and phycocyanins, which broadens the light-harvesting range and accordingly improves the light-harvesting capacity and may be potentially applied in solar energy harvesting.


Asunto(s)
Proteínas Bacterianas , Ficobilinas , Ficobilisomas , Ficocianina , Synechocystis , Ficobilisomas/metabolismo , Ficocianina/metabolismo , Ficocianina/química , Synechocystis/metabolismo , Proteínas Bacterianas/metabolismo , Ficobilinas/metabolismo , Ficobilinas/química , Cianobacterias/metabolismo
4.
Mol Cell Proteomics ; 22(7): 100582, 2023 07.
Artículo en Inglés | MEDLINE | ID: mdl-37225018

RESUMEN

Carbon metabolism is central to photosynthetic organisms and involves the coordinated operation and regulation of numerous proteins. In cyanobacteria, proteins involved in carbon metabolism are regulated by multiple regulators including the RNA polymerase sigma factor SigE, the histidine kinases Hik8, Hik31 and its plasmid-borne paralog Slr6041, and the response regulator Rre37. To understand the specificity and the cross-talk of such regulations, we simultaneously and quantitatively compared the proteomes of the gene knockout mutants for the regulators. A number of proteins showing differential expression in one or more mutants were identified, including four proteins that are unanimously upregulated or downregulated in all five mutants. These represent the important nodes of the intricate and elegant regulatory network for carbon metabolism. Moreover, serine phosphorylation of PII, a key signaling protein sensing and regulating in vivo carbon/nitrogen (C/N) homeostasis through reversible phosphorylation, is massively increased with a concomitant significant decrease in glycogen content only in the hik8-knockout mutant, which also displays impaired dark viability. An unphosphorylatable PII S49A substitution restored the glycogen content and rescued the dark viability of the mutant. Together, our study not only establishes the quantitative relationship between the targets and the corresponding regulators and elucidated their specificity and cross-talk but also unveils that Hik8 regulates glycogen accumulation through negative regulation of PII phosphorylation, providing the first line of evidence that links the two-component system with PII-mediated signal transduction and implicates them in the regulation of carbon metabolism.


Asunto(s)
Carbono , Synechocystis , Fosforilación , Carbono/metabolismo , Proteómica , Synechocystis/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Glucógeno/metabolismo , Nitrógeno , Regulación Bacteriana de la Expresión Génica
5.
Mol Cell Proteomics ; 22(4): 100521, 2023 04.
Artículo en Inglés | MEDLINE | ID: mdl-36858286

RESUMEN

Lysine methylation is a conserved and dynamic regulatory posttranslational modification performed by lysine methyltransferases (KMTs). KMTs catalyze the transfer of mono-, di-, or tri-methyl groups to substrate proteins and play a critical regulatory role in all domains of life. To date, only one KMT has been identified in cyanobacteria. Here, we tested all of the predicted KMTs in the cyanobacterium Synechocystis sp. PCC 6803 (Synechocystis), and we biochemically characterized sll1526 that we termed cKMT1 (cyanobacterial lysine methyltransferase 1) and determined that it can catalyze lysine methylation both in vivo and in vitro. Loss of cKMT1 alters photosynthetic electron transfer in Synechocystis. We analyzed cKMT1-regulated methylation sites in Synechocystis using a timsTOF Pro instrument. We identified 305 class I lysine methylation sites within 232 proteins, and of these, 80 methylation sites in 58 proteins were hypomethylated in ΔcKMT1 cells. We further demonstrated that cKMT1 could methylate ferredoxin-NADP(+) oxidoreductase (FNR) and its potential sites of action on FNR were identified. Amino acid residues H118 and Y219 were identified as key residues in the putative active site of cKMT1 as indicated by structure simulation, site-directed mutagenesis, and KMT activity measurement. Using mutations that mimic the unmethylated forms of FNR, we demonstrated that the inability to methylate K139 residues results in a decrease in the redox activity of FNR and affects energy transfer in Synechocystis. Together, our study identified a new KMT in Synechocystis and elucidated a methylation-mediated molecular mechanism catalyzed by cKMT1 for the regulation of energy transfer in cyanobacteria.


Asunto(s)
Cianobacterias , Ferredoxinas , Synechocystis , Transferencia de Energía , Ferredoxina-NADP Reductasa/química , Ferredoxina-NADP Reductasa/genética , Ferredoxina-NADP Reductasa/metabolismo , Ferredoxinas/química , Ferredoxinas/metabolismo , Lisina , Metiltransferasas/metabolismo , NADP/metabolismo , Synechocystis/metabolismo , Cianobacterias/metabolismo
6.
Proteomics ; : e2300222, 2024 Apr 05.
Artículo en Inglés | MEDLINE | ID: mdl-38581091

RESUMEN

The group 2 σ factor for RNA polymerase SigE plays important role in regulating central carbon metabolism in cyanobacteria. However, the regulation of SigE for these pathways at a proteome level remains unknown. Using a sigE-deficient strain (ΔsigE) of Synechocystis sp. PCC 6803 and quantitative proteomics, we found that SigE depletion induces differential protein expression for sugar catabolic pathways including glycolysis, oxidative pentose phosphate (OPP) pathway, and glycogen catabolism. Two glycogen debranching enzyme homologues Slr1857 and Slr0237 are found differentially expressed in ΔsigE. Glycogen determination indicated that Δslr0237 accumulated glycogen under photomixotrophic condition but was unable to utilize these reserves in the dark, whereas Δslr1857 accumulates and utilizes glycogen in a similar way as the WT strain does in the same condition. These results suggest that Slr0237 plays the major role as the glycogen debranching enzyme in Synechocystis.

7.
Plant J ; 116(3): 706-716, 2023 11.
Artículo en Inglés | MEDLINE | ID: mdl-37493543

RESUMEN

Cyclic electron transport (CET) around photosystem I (PSI) is crucial for photosynthesis to perform photoprotection and sustain the balance of ATP and NADPH. However, the critical component of CET, cyt b6 f complex (cyt b6 f), functions in CET has yet to be understood entirely. In this study, we found that NdhS, a subunit of NADPH dehydrogenase-like (NDH) complex, interacted with cyt b6 f to form a complex in Arabidopsis. This interaction depended on the N-terminal extension of NdhS, which was conserved in eukaryotic plants but defective in prokaryotic algae. The migration of NdhS was much more in cyt b6 f than in PSI-NDH super-complex. Based on these results, we suggested that NdhS and NADP+ oxidoreductase provide a docking domain for the mobile electron carrier ferredoxin to transfer electrons to the plastoquinone pool via cyt b6 f in eukaryotic photosynthesis.


Asunto(s)
Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Complejo de Citocromo b6f/metabolismo , Citocromos b , Transporte de Electrón , Ferredoxinas/metabolismo , Fotosíntesis , Complejo de Proteína del Fotosistema I/metabolismo
8.
Plant Mol Biol ; 114(2): 27, 2024 Mar 13.
Artículo en Inglés | MEDLINE | ID: mdl-38478146

RESUMEN

Cyanobacteria are oxygen-evolving photosynthetic prokaryotes that affect the global carbon and nitrogen turnover. Synechocystis sp. PCC 6803 (Synechocystis 6803) is a model cyanobacterium that has been widely studied and can utilize and uptake various nitrogen sources and amino acids from the outer environment and media. l-arginine is a nitrogen-rich amino acid used as a nitrogen reservoir in Synechocystis 6803, and its biosynthesis is strictly regulated by feedback inhibition. Argininosuccinate synthetase (ArgG; EC 6.3.4.5) is the rate-limiting enzyme in arginine biosynthesis and catalyzes the condensation of citrulline and aspartate using ATP to produce argininosuccinate, which is converted to l-arginine and fumarate through argininosuccinate lyase (ArgH). We performed a biochemical analysis of Synechocystis 6803 ArgG (SyArgG) and obtained a Synechocystis 6803 mutant overexpressing SyArgG and ArgH of Synechocystis 6803 (SyArgH). The specific activity of SyArgG was lower than that of other arginine biosynthesis enzymes and SyArgG was inhibited by arginine, especially among amino acids and organic acids. Both arginine biosynthesis enzyme-overexpressing strains grew faster than the wild-type Synechocystis 6803. Based on previous reports and our results, we suggest that SyArgG is the rate-limiting enzyme in the arginine biosynthesis pathway in cyanobacteria and that arginine biosynthesis enzymes are similarly regulated by arginine in this cyanobacterium. Our results contribute to elucidating the regulation of arginine biosynthesis during nitrogen metabolism.


KEY MESSAGE: This study revealed the catalytic efficiency and inhibition of cyanobacterial argininosuccinate synthetase by arginine and demonstrated that a strain overexpressing this enzyme grew faster than the wild-type strain.


Asunto(s)
Synechocystis , Synechocystis/genética , Synechocystis/metabolismo , Ácido Aspártico/metabolismo , Arginina/metabolismo , Fotosíntesis , Nitrógeno/metabolismo
9.
Plant Mol Biol ; 114(3): 60, 2024 May 17.
Artículo en Inglés | MEDLINE | ID: mdl-38758412

RESUMEN

Pyruvate kinase (Pyk, EC 2.7.1.40) is a glycolytic enzyme that generates pyruvate and adenosine triphosphate (ATP) from phosphoenolpyruvate (PEP) and adenosine diphosphate (ADP), respectively. Pyk couples pyruvate and tricarboxylic acid metabolisms. Synechocystis sp. PCC 6803 possesses two pyk genes (encoded pyk1, sll0587 and pyk2, sll1275). A previous study suggested that pyk2 and not pyk1 is essential for cell viability; however, its biochemical analysis is yet to be performed. Herein, we biochemically analyzed Synechocystis Pyk2 (hereafter, SyPyk2). The optimum pH and temperature of SyPyk2 were 7.0 and 55 °C, respectively, and the Km values for PEP and ADP under optimal conditions were 1.5 and 0.053 mM, respectively. SyPyk2 is activated in the presence of glucose-6-phosphate (G6P) and ribose-5-phosphate (R5P); however, it remains unaltered in the presence of adenosine monophosphate (AMP) or fructose-1,6-bisphosphate. These results indicate that SyPyk2 is classified as PykA type rather than PykF, stimulated by sugar monophosphates, such as G6P and R5P, but not by AMP. SyPyk2, considering substrate affinity and effectors, can play pivotal roles in sugar catabolism under nonphotosynthetic conditions.


Asunto(s)
Glucosa-6-Fosfato , Fosfoenolpiruvato , Piruvato Quinasa , Ribosamonofosfatos , Synechocystis , Synechocystis/metabolismo , Synechocystis/genética , Piruvato Quinasa/metabolismo , Piruvato Quinasa/genética , Fosfoenolpiruvato/metabolismo , Glucosa-6-Fosfato/metabolismo , Ribosamonofosfatos/metabolismo , Especificidad por Sustrato , Concentración de Iones de Hidrógeno , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Cinética , Temperatura
10.
Plant Cell Physiol ; 65(6): 975-985, 2024 Jun 27.
Artículo en Inglés | MEDLINE | ID: mdl-38147500

RESUMEN

DesC1 and DesC2, which are fatty acid desaturases found in cyanobacteria, are responsible for introducing a double bond at the Δ9 position of fatty-acyl chains, which are subsequently esterified to the sn-1 and sn-2 positions of the glycerol moiety, respectively. However, since the discovery of these two desaturases in the Antarctic cyanobacterium Nostoc sp. SO-36, no further research has been reported. This study presents a comprehensive characterization of DesC1 and DesC2 through targeted mutagenesis and transformation using two cyanobacteria strains: Anabaena sp. PCC 7120, comprising both desaturases, and Synechocystis sp. PCC 6803, containing a single Δ9 desaturase (hereafter referred to as DesCs) sharing similarity with DesC1 in amino acid sequence. The results suggested that both DesC1 and DesC2 were essential in Anabaena sp. PCC 7120 and that DesC1, but not DesC2, complemented DesCs in Synechocystis sp. PCC 6803. In addition, DesC2 from Anabaena sp. PCC 7120 desaturated fatty acids esterified to the sn-2 position of the glycerol moiety in Synechocystis sp. PCC 6803.


Asunto(s)
Anabaena , Proteínas Bacterianas , Ácido Graso Desaturasas , Synechocystis , Ácido Graso Desaturasas/metabolismo , Ácido Graso Desaturasas/genética , Synechocystis/enzimología , Synechocystis/genética , Anabaena/enzimología , Anabaena/genética , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Ácidos Grasos/metabolismo , Cianobacterias/enzimología , Cianobacterias/genética , Secuencia de Aminoácidos
11.
Plant Cell Physiol ; 65(1): 95-106, 2024 Jan 19.
Artículo en Inglés | MEDLINE | ID: mdl-37874689

RESUMEN

The spatial separation of photosystems I and II (PSI and PSII) is thought to be essential for efficient photosynthesis by maintaining a balanced flow of excitation energy between them. Unlike the thylakoid membranes of plant chloroplasts, cyanobacterial thylakoids do not form tightly appressed grana stacks that enforce strict lateral separation. The coexistence of the two photosystems provides a ground for spillover-excitation energy transfer from PSII to PSI. Spillover has been considered as a pathway of energy transfer from the phycobilisomes to PSI and may also play a role in state transitions as means to avoid overexcitation of PSII. Here, we demonstrate a significant degree of energy spillover from PSII to PSI in reconstituted membranes and isolated thylakoid membranes of Thermosynechococcus (Thermostichus) vulcanus and Synechocystis sp. PCC 6803 by steady-state and time-resolved fluorescence spectroscopy. The quantum yield of spillover in these systems was determined to be up to 40%. Spillover was also found in intact cells but to a considerably lower degree (20%) than in isolated thylakoid membranes. The findings support a model of coexistence of laterally separated microdomains of PSI and PSII in the cyanobacterial cells as well as domains where the two photosystems are energetically connected. The methodology presented here can be applied to probe spillover in other photosynthetic organisms.


Asunto(s)
Synechocystis , Tilacoides , Tilacoides/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Fotosíntesis , Complejo de Proteína del Fotosistema I/metabolismo , Synechocystis/metabolismo
12.
Plant Cell Physiol ; 65(5): 790-797, 2024 May 30.
Artículo en Inglés | MEDLINE | ID: mdl-38441322

RESUMEN

Cyanobacteria inhabit areas with a broad range of light, temperature and nutrient conditions. The robustness of cyanobacterial cells, which can survive under different conditions, may depend on the resilience of photosynthetic activity. Cyanothece sp. PCC 8801 (Cyanothece), a freshwater cyanobacterium isolated from a Taiwanese rice field, had a higher repair activity of photodamaged photosystem II (PSII) under intense light than Synechocystis sp. PCC 6803 (Synechocystis), another freshwater cyanobacterium. Cyanothece contains myristic acid (14:0) as the major fatty acid at the sn-2 position of the glycerolipids. To investigate the role of 14:0 in the repair of photodamaged PSII, we used a Synechocystis transformant expressing a T-1274 encoding a lysophosphatidic acid acyltransferase (LPAAT) from Cyanothece. The wild-type and transformant cells contained 0.2 and 20.1 mol% of 14:0 in glycerolipids, respectively. The higher content of 14:0 in the transformants increased the fluidity of the thylakoid membrane. In the transformants, PSII repair was accelerated due to an enhancement in the de novo synthesis of D1 protein, and the production of singlet oxygen (1O2), which inhibited protein synthesis, was suppressed. The high content of 14:0 increased transfer of light energy received by phycobilisomes to PSI and CP47 in PSII and the content of carotenoids. These results indicated that an increase in 14:0 reduced 1O2 formation and enhanced PSII repair. The higher content of 14:0 in the glycerolipids may be required as a survival strategy for Cyanothece inhabiting a rice field under direct sunlight.


Asunto(s)
Luz , Ácido Mirístico , Complejo de Proteína del Fotosistema II , Synechocystis , Tilacoides , Complejo de Proteína del Fotosistema II/metabolismo , Synechocystis/metabolismo , Synechocystis/genética , Ácido Mirístico/metabolismo , Tilacoides/metabolismo , Fotosíntesis , Aciltransferasas/metabolismo , Aciltransferasas/genética , Oxígeno Singlete/metabolismo
13.
Plant Cell Physiol ; 2024 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-39034452

RESUMEN

Phycobilisomes play a crucial role in the light-harvesting mechanisms of cyanobacteria, red algae, and glaucophytes, but the molecular mechanism of their regulation is largely unknown. In the cyanobacterium, Synechocystis sp. PCC 6803, we identified a gene, slr0244, as a phycobilisome-related gene using phylogenetic profiling analysis, a method to predict gene function based on comparative genomics. To investigate the physiological function of the slr0244 gene, we characterize the slr0244 mutants spectroscopically. The disruption of the slr0244 gene impaired state transition, a process by which the distribution of light energy absorbed by the phycobilisomes between two photosystems was regulated in response to the changes in light conditions. The Slr0244 protein seems to act somewhere at or downstream of the sensing step of the redox state of the plastoquinone pool in the process of state transition. These findings, together with the past report of the interaction of this gene product with thioredoxin or glutaredoxin, suggest that the slr0244 gene is a novel state-transition regulator that integrates the redox signal of plastoquinone pools with that of photosystem I-reducing side. The protein has two USP (universal stress protein) motifs in tandem. The second motif has two conserved cysteine residues found in USPs of other cyanobacteria and land plants. These redox-type USPs with conserved cysteines may function as redox regulators in various photosynthetic organisms. Our study also showed the efficacy of the phylogenetic profiling analysis in predicting the function of cyanobacterial genes that have not been annotated so far.

14.
Biochem Biophys Res Commun ; 702: 149595, 2024 Apr 02.
Artículo en Inglés | MEDLINE | ID: mdl-38340653

RESUMEN

The Photosystem II water-plastoquinone oxidoreductase is a multi-subunit complex which catalyses the light-driven oxidation of water to molecular oxygen in oxygenic photosynthesis. The D1 reaction centre protein exists in multiple forms in cyanobacteria, including D1FR which is expressed under far-red light. We investigated the role of Phe184 that is found in the lumenal cd-loop of D1FR but is typically an isoleucine in other D1 isoforms. The I184F mutant in Synechocystis sp. PCC 6803 was similar to the control strain but accumulated a spontaneous mutation that introduced a Gln residue in place of His252 located on the opposite side of the thylakoid membrane. His252 participates in the protonation of the secondary plastoquinone electron acceptor QB. The I184F:H252Q double mutant exhibited reduced high-light-induced photodamage and an altered QB-binding site that impaired herbicide binding. Additionally, the H252Q mutant had a large increase in the variable fluorescence yield although the number of photochemically active PS II centres was unchanged. In the I184F:H252Q mutant the extent of the increased fluorescence yield decreased. Our data indicates substitution of Ile184 to Phe modulates PS II-specific variable fluorescence in cells with the His252 to Gln substitution by modifying the QB-binding site.


Asunto(s)
Complejo de Proteína del Fotosistema II , Synechocystis , Complejo de Proteína del Fotosistema II/química , Synechocystis/genética , Synechocystis/metabolismo , Plastoquinona/química , Plastoquinona/metabolismo , Mutagénesis , Oxígeno/metabolismo , Mutación , Agua/metabolismo
15.
New Phytol ; 241(3): 1236-1249, 2024 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-37986097

RESUMEN

Biogenesis of the photosynthetic apparatus requires complicated molecular machinery, individual components of which are either poorly characterized or unknown. The BtpA protein has been described as a factor required for the stability of photosystem I (PSI) in cyanobacteria; however, how the BtpA stabilized PSI remains unexplained. To clarify the role of BtpA, we constructed and characterized the btpA-null mutant (ΔbtpA) in the cyanobacterium Synechocystis sp. PCC 6803. The mutant contained only c. 1% of chlorophyll and nearly no thylakoid membranes. However, this strain, growing only in the presence of glucose, was genetically unstable and readily generated suppressor mutations that restore the photoautotrophy. Two suppressor mutations were mapped into the hemA gene encoding glutamyl-tRNA reductase (GluTR) - the first enzyme of tetrapyrrole biosynthesis. Indeed, the GluTR was not detectable in the ΔbtpA mutant and the suppressor mutations restored biosynthesis of tetrapyrroles and photoautotrophy by increased GluTR expression or by improved GluTR stability/processivity. We further demonstrated that GluTR associates with a large BtpA oligomer and that BtpA is required for the stability of GluTR. Our results show that the BtpA protein is involved in the biogenesis of photosystems at the level of regulation of tetrapyrrole biosynthesis.


Asunto(s)
Cianobacterias , Tilacoides , Tilacoides/metabolismo , Clorofila/metabolismo , Complejo de Proteína del Fotosistema I/genética , Complejo de Proteína del Fotosistema I/metabolismo , Tetrapirroles/metabolismo , Cianobacterias/metabolismo
16.
New Phytol ; 243(3): 936-950, 2024 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-38831647

RESUMEN

Glycosyltransferases (GTs) are enzymes that transfer sugars to various targets. They play important roles in diverse biological processes, including photosynthesis, cell motility, exopolysaccharide biosynthesis, and lipid metabolism; however, their involvement in regulating carbon metabolism in Synechocystis sp. PCC 6803 has not been reported. We identified a novel GT protein, Slr1064, involved in carbon metabolism. The effect of slr1064 deletion on the growth of Synechocystis cells and functional mechanisms of Slr1064 on carbon metabolism were thoroughly investigated through physiological, biochemistry, proteomic, and metabolic analyses. We found that this GT, which is mainly distributed in the membrane compartment, is essential for the growth of Synechocystis under heterotrophic and mixotrophic conditions, but not under autotrophic conditions. The deletion of slr1064 hampers the turnover rate of Gap2 under mixotrophic conditions and disrupts the assembly of the PRK/GAPDH/CP12 complex under dark culture conditions. Additionally, UDP-GlcNAc, the pivotal metabolite responsible for the O-GlcNAc modification of GAPDH, is downregulated in the Δslr1064. Our work provides new insights into the role of GTs in carbon metabolism in Synechocystis and elucidate the mechanism by which carbon metabolism is regulated in this important model organism.


Asunto(s)
Proteínas Bacterianas , Carbono , Glicosiltransferasas , Synechocystis , Uridina Difosfato N-Acetilglucosamina , Synechocystis/metabolismo , Synechocystis/genética , Synechocystis/crecimiento & desarrollo , Carbono/metabolismo , Glicosiltransferasas/metabolismo , Glicosiltransferasas/genética , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Uridina Difosfato N-Acetilglucosamina/metabolismo , Regulación Bacteriana de la Expresión Génica , Eliminación de Gen
17.
Photosynth Res ; 159(2-3): 241-251, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-37480468

RESUMEN

In this study, the effects of cationic antiseptics such as chlorhexidine, picloxidine, miramistin, and octenidine at concentrations up to 150 µM on fluorescence spectra and its lifetimes, as well as on light-induced electron transfer in protein-pigment complexes of photosystem I (PSI) isolated from cyanobacterium Synechocystis sp. PCC 6803 have been studied. In doing so, octenidine turned out to be the most "effective" in terms of its influence on the spectral and functional characteristics of PSI complexes. It has been shown that the rate of energy migration from short-wavelength forms of light-harvesting chlorophyll to long-wavelength ones slows down upon addition of octenidine to the PSI suspension. After photo-separation of charges between the primary electron donor P700 and the terminal iron-sulfur center(s) FA/FB, the rate of forward electron transfer from (FA/FB)- to the external medium slows down while the rate of charge recombination between reduced FA/FB- and photooxidized P700+ increases. The paper considers the possible causes of the observed action of the antiseptic.


Asunto(s)
Antiinfecciosos Locales , Iminas , Piridinas , Synechocystis , Complejo de Proteína del Fotosistema I , Electrones , Cationes
18.
Photosynth Res ; 2024 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-39037691

RESUMEN

Mg2+, the most abundant divalent cation in living cells, plays a pivotal role in numerous enzymatic reactions and is of particular importance for organisms performing oxygenic photosynthesis. Its significance extends beyond serving as the central ion of the chlorophyll molecule, as it also acts as a counterion during the light reaction to balance the proton gradient across the thylakoid membranes. In this study, we investigated the effects of Mg2+ limitation on the physiology of the well-known model microorganism Synechocystis sp. PCC6803. Our findings reveal that Mg2+ deficiency triggers both morphological and functional changes. As seen in other oxygenic photosynthetic organisms, Mg2+ deficiency led to a decrease in cellular chlorophyll concentration. Moreover, the PSI-to-PSII ratio decreased, impacting the photosynthetic efficiency of the cell. In line with this, Mg2+ deficiency led to a change in the proton gradient built up across the thylakoid membrane upon illumination.

19.
Photosynth Res ; 161(1-2): 117-125, 2024 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-38546812

RESUMEN

Cyanobacteria are among the most suitable organisms for the capture of excessive amounts of CO2 and can be grown in extreme environments. In our research we use the single-celled freshwater cyanobacteria Synechococcus elongatus PCC7942 PAMCOD strain and Synechocystis sp. PCC6714 for the production of carbohydrates and hydrogen. PAMCOD strain and Synechocystis sp. PCC6714 synthesize sucrose when exposed to salinity stress, as their main compatible osmolyte. We examined the cell proliferation rate and the sucrose accumulation in those two different strains of cyanobacteria under salt (0.4 M NaCl) and heat stress (35 0C) conditions. The intracellular sucrose (mol sucrose content per Chl a) was found to increase by 50% and 108% in PAMCOD strain and Synechocystis sp. PCC6714 cells, respectively. As previously reported, PAMCOD strain has the ability to produce hydrogen through the process of dark anaerobic fermentation (Vayenos D, Romanos GE, Papageorgiou GC, Stamatakis K (2020) Photosynth Res 146, 235-245). In the present study, we demonstrate that Synechocystis sp. PCC6714 has also this ability. We further examined the optimal conditions during the dark fermentation of PAMCOD and Synechocystis sp. PCC6714 regarding H2 formation, increasing the PAMCOD H2 productivity from 2 nmol H2 h- 1 mol Chl a- 1 to 23 nmol H2 h- 1 mol Chl a- 1. Moreover, after the dark fermentation, the cells demonstrated proliferation in both double BG-11 and BG-11 medium enriched in NaNO3, thus showing the sustainability of the procedure.


Asunto(s)
Hidrógeno , Synechococcus , Synechocystis , Hidrógeno/metabolismo , Synechococcus/metabolismo , Synechococcus/fisiología , Synechococcus/efectos de los fármacos , Synechocystis/metabolismo , Synechocystis/fisiología , Respuesta al Choque Térmico/fisiología , Sacarosa/metabolismo , Cloruro de Sodio/farmacología , Fermentación , Fotosíntesis , Calor
20.
Photosynth Res ; 159(2-3): 97-114, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-37093504

RESUMEN

Flavodiiron proteins Flv1/Flv3 accept electrons from photosystem (PS) I. In this work we investigated light adaptation mechanisms of Flv1-deficient mutant of Synechocystis PCC 6803, incapable to form the Flv1/Flv3 heterodimer. First seconds of dark-light transition were studied by parallel measurements of light-induced changes in chlorophyll fluorescence, P700 redox transformations, fluorescence emission at 77 K, and OCP-dependent fluorescence quenching. During the period of Calvin cycle activation upon dark-light transition, the linear electron transport (LET) in wild type is supported by the Flv1/Flv3 heterodimer, whereas in Δflv1 mutant activation of LET upon illumination is preceded by cyclic electron flow that maintains State 2. The State 2-State 1 transition and Orange Carotenoid Protein (OCP)-dependent non-photochemical quenching occur independently of each other, begin in about 10 s after the illumination of the cells and are accompanied by a short-term re-reduction of the PSI reaction center (P700+). ApcD is important for the State 2-State 1 transition in the Δflv1 mutant, but S-M rise in chlorophyll fluorescence was not completely inhibited in Δflv1/ΔapcD mutant. LET in Δflv1 mutant starts earlier than the S-M rise in chlorophyll fluorescence, and the oxidation of plastoquinol (PQH2) pool promotes the activation of PSII, transient re-reduction of P700+ and transition to State 1. An attempt to induce state transition in the wild type under high intensity light using methyl viologen, highly oxidizing P700 and PQH2, was unsuccessful, showing that oxidation of intersystem electron-transport carriers might be insufficient for the induction of State 2-State 1 transition in wild type of Synechocystis under high light.


Asunto(s)
Synechocystis , Transporte de Electrón , Synechocystis/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Mutación , Oxidación-Reducción , Complejo de Proteína del Fotosistema I/genética , Complejo de Proteína del Fotosistema I/metabolismo , Carotenoides/metabolismo , Clorofila/metabolismo , Complejo de Proteína del Fotosistema II/genética , Complejo de Proteína del Fotosistema II/metabolismo
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