scholarly journals Anabaena sp. strain PCC 7120 conR contains a LytR-CpsA-Psr domain, is developmentally regulated, and is essential for diazotrophic growth and heterocyst morphogenesis

Microbiology ◽  
2011 ◽  
Vol 157 (3) ◽  
pp. 617-626 ◽  
Author(s):  
Rodrigo A. Mella-Herrera ◽  
M. Ramona Neunuebel ◽  
James W. Golden
Keyword(s):  
Nitrogenase Activity ◽  
Growth Defect ◽  
Filamentous Cyanobacterium ◽  
Developmentally Regulated ◽  
Vegetative Cells ◽  
Septum Formation ◽  
Insertional Inactivation ◽  
Step Down ◽  
Pcc 7120 ◽  
Diazotrophic Growth

The conR (all0187) gene of the filamentous cyanobacterium Anabaena (Nostoc) sp. strain PCC 7120 is predicted to be part of a family of proteins that contain the LytR-CpsA-Psr domain associated with septum formation and cell wall maintenance. The conR gene was originally misannotated as a transcription regulator. Northern RNA blot analysis showed that conR expression was upregulated 8 h after nitrogen step-down. Fluorescence microscopy of a P conR –gfp reporter strain revealed increased GFP fluorescence in proheterocysts and heterocysts beginning 9 h after nitrogen step-down. Insertional inactivation of conR caused a septum-formation defect of vegetative cells grown in nitrate-containing medium. In nitrate-free medium, mutant filaments formed abnormally long heterocysts and were defective for diazotrophic growth. Septum formation between heterocysts and adjacent vegetative cells was abnormal, often with one or both poles of the heterocysts appearing partially open. In a conR mutant, expression of nifH was delayed after nitrogen step-down and nitrogenase activity was approximately 70 % of wild-type activity, indicating that heterocysts of the conR mutant strain are partially functional. We hypothesize that the diazotrophic growth defect is caused by an inability of the heterocysts to transport fixed nitrogen to the neighbouring vegetative cells.

scholarly journals Inactivation of a Heterocyst-Specific Invertase Indicates a Principal Role of Sucrose Catabolism in Heterocysts of Anabaena sp.

2010 ◽  
Vol 192 (20) ◽  
pp. 5526-5533 ◽  
Author(s):  
Rocío López-Igual ◽  
Enrique Flores ◽  
Antonia Herrero
Keyword(s):  
Fluorescent Protein ◽  
Northern Analysis ◽  
Filamentous Cyanobacterium ◽  
Nitrogen Deprivation ◽  
Vegetative Cells ◽  
Anabaena Sp ◽  
Green Fluorescent ◽  
Pcc 7120 ◽  
Diazotrophic Growth

ABSTRACT Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that carries out N2 fixation in specialized cells called heterocysts, which exchange nutrients and regulators with the filament's vegetative cells that perform the photosynthetic fixation of CO2. The Anabaena genome carries two genes coding for alkaline/neutral invertases, invA and invB. As shown by Northern analysis, both genes were expressed monocistronically and induced under nitrogen deprivation, although induction was stronger for invB than for invA. Whereas expression of an InvA-N-GFP fusion (green fluorescent protein [GFP] fused to the N terminus of the InvA protein [InvA-N]) was homogeneous along the cyanobacterial filament, consistent with the lack of dependence on HetR, expression of an InvB-N-GFP fusion upon combined nitrogen deprivation took place mainly in differentiating and mature heterocysts. In an hetR genetic background, the InvB-N-GFP fusion was strongly expressed all along the filament. An insertional mutant of invA could grow diazotrophically but was impaired in nifHDK induction and exhibited an increased frequency of heterocysts, suggesting a regulatory role of the invertase-mediated carbon flux in vegetative cells. In contrast, an invB mutant was strongly impaired in diazotrophic growth, showing a crucial role of sucrose catabolism mediated by the InvB invertase in the heterocysts.

scholarly journals PatS and Products of Nitrogen Fixation Control Heterocyst Pattern

2001 ◽  
Vol 183 (8) ◽  
pp. 2605-2613 ◽  
Author(s):  
Ho-Sung Yoon ◽  
James W. Golden
Keyword(s):  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Early Stage ◽  
Critical Role ◽  
Filamentous Cyanobacterium ◽  
Vegetative Cells ◽  
Heterocyst Differentiation ◽  
Partial Restoration ◽  
Fixation Control ◽  
Pcc 7120

ABSTRACT The filamentous cyanobacterium Anabaena sp. strain PCC 7120 forms a developmental pattern of single heterocysts separated by approximately 10 vegetative cells. Heterocysts differentiate from vegetative cells and are specialized for nitrogen fixation. ThepatS gene, which encodes a small peptide that inhibits heterocyst differentiation, is expressed in proheterocysts and plays a critical role in establishing the heterocyst pattern. Here we present further analysis of patS expression and heterocyst pattern formation. A patS-gfp reporter strain revealed clusters of patS-expressing cells during the early stage of heterocyst differentiation. PatS signaling is likely to be involved in the resolution of these clusters. Differentiating cells were inhibited by PatS during the time period 6 to 12 h after heterocyst induction, when groups of differentiating cells were being resolved to a single proheterocyst. Increased transcription ofpatS during development coincided with expression from a new transcription start site. In vegetative cells grown on nitrate, the 5′ end of a transcript for patS was localized 314 bases upstream from the first translation initiation codon. After heterocyst induction, a new transcript with a 5′ end at −39 bases replaced the vegetative cell transcript. A patS mutant grown for several days under nitrogen-fixing conditions showed partial restoration of the normal heterocyst pattern, presumably because of a gradient of nitrogen compounds supplied by the heterocysts. ThepatS mutant formed heterocysts when grown in the presence of nitrate but showed no nitrogenase activity and no obvious heterocyst pattern. We conclude that PatS and products of nitrogen fixation are the main signals determining the heterocyst pattern.

scholarly journals Catabolic Function of Compartmentalized Alanine Dehydrogenase in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120

2010 ◽  
Vol 192 (19) ◽  
pp. 5165-5172 ◽  
Author(s):  
Rafael Pernil ◽  
Antonia Herrero ◽  
Enrique Flores
Keyword(s):  
Fluorescent Protein ◽  
Nitrogenase Activity ◽  
Vegetative Cells ◽  
Reading Frame ◽  
Alanine Dehydrogenase ◽  
Cyanobacterium Anabaena ◽  
Green Fluorescent ◽  
Pcc 7120 ◽  
Diazotrophic Growth

ABSTRACT In the diazotrophic filaments of heterocyst-forming cyanobacteria, an exchange of metabolites takes place between vegetative cells and heterocysts that results in a net transfer of reduced carbon to the heterocysts and of fixed nitrogen to the vegetative cells. Open reading frame alr2355 of the genome of Anabaena sp. strain PCC 7120 is the ald gene encoding alanine dehydrogenase. A strain carrying a green fluorescent protein (GFP) fusion to the N terminus of Ald (Ald-N-GFP) showed that the ald gene is expressed in differentiating and mature heterocysts. Inactivation of ald resulted in a lack of alanine dehydrogenase activity, a substantially decreased nitrogenase activity, and a 50% reduction in the rate of diazotrophic growth. Whereas production of alanine was not affected in the ald mutant, in vivo labeling with [14C]alanine (in whole filaments and isolated heterocysts) or [14C]pyruvate (in whole filaments) showed that alanine catabolism was hampered. Thus, alanine catabolism in the heterocysts is needed for normal diazotrophic growth. Our results extend the significance of a previous work that suggested that alanine is transported from vegetative cells into heterocysts in the diazotrophic Anabaena filament.

scholarly journals Anabaena sp. Strain PCC 7120 Gene devH Is Required for Synthesis of the Heterocyst Glycolipid Layer

2005 ◽  
Vol 187 (7) ◽  
pp. 2326-2331 ◽  
Author(s):  
Martha E. Ramírez ◽  
Pratibha B. Hebbar ◽  
Ruanbao Zhou ◽  
C. Peter Wolk ◽  
Stephanie E. Curtis
Keyword(s):  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Regulatory Protein ◽  
Protein A ◽  
Filamentous Cyanobacterium ◽  
Fixed Nitrogen ◽  
Ultrastructural Studies ◽  
Anabaena Sp ◽  
Pcc 7120 ◽  
Mutant Forms

ABSTRACT In response to deprivation for fixed nitrogen, the filamentous cyanobacterium Anabaena sp. strain PCC 7120 provides a microoxic intracellular environment for nitrogen fixation through the differentiation of semiregularly spaced vegetative cells into specialized cells called heterocysts. The devH gene is induced during heterocyst development and encodes a product with characteristics of a trans-acting regulatory protein. A devH mutant forms morphologically distinguishable heterocysts but is Fox−, incapable of nitrogen fixation in the presence of oxygen. We demonstrate that rearrangements of nitrogen fixation genes take place normally in the devH mutant and that it is Fix+, i.e., has nitrogenase activity under anoxic conditions. The Fox− phenotype was shown by ultrastructural studies to be associated with the absence of the glycolipid layer of the heterocyst envelope. The expression of glycolipid biosynthetic genes in the mutant is greatly reduced, and heterocyst glycolipids are undetectable.

scholarly journals Identification and Inactivation of Three Group 2 Sigma Factor Genes in Anabaena sp. Strain PCC 7120

2001 ◽  
Vol 183 (22) ◽  
pp. 6667-6675 ◽  
Author(s):  
Ivan Y. Khudyakov ◽  
James W. Golden
Keyword(s):  
Sigma Factor ◽  
Double Mutant ◽  
Developmental Regulation ◽  
Single Gene ◽  
Sigma Factors ◽  
Nitrogen Deprivation ◽  
Insertional Inactivation ◽  
Group 2 ◽  
Pcc 7120 ◽  
Diazotrophic Growth

ABSTRACT Three new Anabaena sp. strain PCC 7120 genes encoding group 2 alternative sigma factors have been cloned and characterized. Insertional inactivation of sigD,sigE, and sigF genes did not affect growth on nitrate under standard laboratory conditions but did transiently impair the abilities of sigD andsigE mutant strains to establish diazotrophic growth. AsigD sigE double mutant, though proficient in growth on nitrate and still able to differentiate into distinct proheterocysts, was unable to grow diazotrophically due to extensive fragmentation of filaments upon nitrogen deprivation. This double mutant could be complemented by wild-type copies of sigDor sigE, indicating some degree of functional redundancy that can partially mask phenotypes of single gene mutants. However, thesigE gene was required for lysogenic development of the temperate cyanophage A-4L. Several other combinations of double mutations, especially sigE sigF, caused a transient defect in establishing diazotrophic growth, manifested as a strong and prolonged bleaching response to nitrogen deprivation. We found no evidence for developmental regulation of the sigma factor genes.luxAB reporter fusions with sigD,sigE, and sigF all showed slightly reduced expression after induction of heterocyst development by nitrogen stepdown. Phylogenetic analysis of cyanobacterial group 2 sigma factor sequences revealed that they fall into several subgroups. Three morphologically and physiologically distant strains,Anabaena sp. strain PCC 7120,Synechococcus sp. strain PCC 7002, andSynechocystis sp. strain PCC 6803 each contain representatives of four subgroups. Unlike unicellular strains,Anabaena sp. strain PCC 7120 has three additional group 2 sigma factors that cluster in subgroup 2.5b, which is perhaps specific for filamentous or heterocystous cyanobacteria.

scholarly journals Posttranscriptional Regulation of Glutamine Synthetase in the Filamentous Cyanobacterium Anabaena sp. PCC 7120: Differential Expression between Vegetative Cells and Heterocysts

2010 ◽  
Vol 192 (18) ◽  
pp. 4701-4711 ◽  
Author(s):  
Carla V. Galmozzi ◽  
Lorena Saelices ◽  
Francisco J. Florencio ◽  
M. Isabel Muro-Pastor
Keyword(s):  
Glutamine Synthetase ◽  
Differential Expression ◽  
Posttranscriptional Regulation ◽  
Filamentous Cyanobacterium ◽  
Vegetative Cells ◽  
Amino Terminal ◽  
Cyanobacterium Anabaena ◽  
In The Wild ◽  
Pcc 7120

ABSTRACT Genes homologous to those implicated in glutamine synthetase (GS) regulation by protein-protein interaction in the cyanobacterium Synechocystis sp. strain PCC 6803 are conserved in several cyanobacterial sequenced genomes. We investigated this GS regulatory mechanism in Anabaena sp. strain PCC 7120. In this strain the system operates with only one GS inactivation factor (inactivation factor 7A [IF7A]), encoded by open reading frame (ORF) asl2329 (gifA). Following addition of ammonium, expression of gifA is derepressed, leading to the synthesis of IF7A, and consequently, GS is inactivated. Upon ammonium removal, the GS activity returns to the initial level and IF7A becomes undetectable. The global nitrogen control protein NtcA binds to the gifA promoter. Constitutive high expression levels of gifA were found in an Anabaena ntcA mutant (CSE2), indicating a repressor role for NtcA. In vitro studies demonstrate that Anabaena GS is not inactivated by Synechocystis IFs (IF7 and IF17), indicating the specificity of the system. We constructed an Anabaena strain expressing a second inactivating factor, containing the amino-terminal part of IF17 from Synechocystis fused to IF7A. GS inactivation in this strain is more effective than that in the wild type (WT) and resembles that observed in Synechocystis. Finally we found differential expression of the IF system between heterocysts and vegetative cells of Anabaena.

scholarly journals The Cyanotoxin BMAA Induces Heterocyst Specific Gene Expression in Anabaena sp. PCC 7120 under Repressive Conditions

Toxins ◽  
2018 ◽  
Vol 10 (11) ◽  
pp. 478 ◽  
Author(s):  
Alexandra Popova ◽  
Tatiana Semashko ◽  
Natalia Kostina ◽  
Ulla Rasmussen ◽  
Vadim Govorun ◽  
...  
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Specific Gene ◽  
Filamentous Cyanobacteria ◽  
Protein Amino Acid ◽  
Vegetative Cells ◽  
Differentiated Cells ◽  
Anabaena Sp ◽  
Polymerase Chain ◽  
Pcc 7120 ◽  
Diazotrophic Growth

Cyanobacteria synthesize neurotoxic β-N-methylamino-l-alanine (BMAA). The roles of this non-protein amino acid in cyanobacterial cells are insufficiently studied. During diazotrophic growth, filamentous cyanobacteria form single differentiated cells, called heterocysts, which are separated by approximately 12–15 vegetative cells. When combined nitrogen is available, heterocyst formation is blocked and cyanobacterial filaments contain only vegetative cells. In the present study, we discovered that exogenous BMAA induces the process of heterocyst formation in filamentous cyanobacteria under nitrogen-replete conditions that normally repress cell differentiation. BMAA treated cyanobacteria form heterocyst-like dark non-fluorescent non-functional cells. It was found that glutamate eliminates the BMAA mediated derepression. Quantitative polymerase chain reaction (qPCR) permitted to detect the BMAA impact on the transcriptional activity of several genes that are implicated in nitrogen assimilation and heterocyst formation in Anabaena sp. PCC 7120. We demonstrated that the expression of several essential genes increases in the BMAA presence under repressive conditions.

Cytochrome c oxidase genes required for nitrogenase activity and diazotrophic growth in Anabaena sp. PCC 7120

2003 ◽  
Vol 47 (5) ◽  
pp. 1239-1249 ◽  
Author(s):  
Ana Valladares ◽  
Antonia Herrero ◽  
Dietmar Pils ◽  
Georg Schmetterer ◽  
Enrique Flores
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Nitrogenase Activity ◽  
Anabaena Sp ◽  
Pcc 7120 ◽  
Diazotrophic Growth

scholarly journals A TolC-Like Protein Is Required for Heterocyst Development in Anabaena sp. Strain PCC 7120

2007 ◽  
Vol 189 (21) ◽  
pp. 7887-7895 ◽  
Author(s):  
Suncana Moslavac ◽  
Kerstin Nicolaisen ◽  
Oliver Mirus ◽  
Fadi Al Dehni ◽  
Rafael Pernil ◽  
...  
Keyword(s):  
Morphological Changes ◽  
Protein Export ◽  
Filamentous Cyanobacterium ◽  
Sole Nitrogen Source ◽  
Wild Type ◽  
Anabaena Sp ◽  
Pcc 7120 ◽  
Export System ◽  
Diazotrophic Growth ◽  
Mutant Cells

ABSTRACT The filamentous cyanobacterium Anabaena sp. strain PCC 7120 forms heterocysts in a semiregular pattern when it is grown on N2 as the sole nitrogen source. The transition from vegetative cells to heterocysts requires marked metabolic and morphological changes. We show that a trimeric pore-forming outer membrane β-barrel protein belonging to the TolC family, Alr2887, is up-regulated in developing heterocysts and is essential for diazotrophic growth. Mutants defective in Alr2887 did not form the specific glycolipid layer of the heterocyst cell wall, which is necessary to protect nitrogenase from external oxygen. Comparison of the glycolipid contents of wild-type and mutant cells indicated that the protein is not involved in the synthesis of glycolipids but might instead serve as an exporter for the glycolipid moieties or enzymes involved in glycolipid attachment. We propose that Alr2887, together with an ABC transporter like DevBCA, is part of a protein export system essential for assembly of the heterocyst glycolipid layer. We designate the alr2887 gene hgdD (heterocyst glycolipid deposition protein).

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