Anabaena sp. strain PCC 7120 ntcA gene required for growth on nitrate and heterocyst development

Wei, Tingcun; Ramasubramanian, T.; Golden, James W.

doi:10.1128/jb.176.15.4473-4482.1994

Cited by 195 publications

(182 citation statements)

References 40 publications

(67 reference statements)

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“…strain PCC 7120 requires ammonium for growth and is unable to induce hetR, develop heterocysts, or synthesize nitrogenase and enzymes of the nitrate reduction system (Frı! as et al, 1994 ;Wei et al, 1994).…”

Section: Nitrogen Regulation and Heterocyst Developmentmentioning

confidence: 94%

Tansley Review No. 107. Heterocyst and akinete differentiation in cyanobacteria

1999

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Section: Nitrogen Regulation and Heterocyst Developmentmentioning

confidence: 94%

Tansley Review No. 107. Heterocyst and akinete differentiation in cyanobacteria

1999

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“…For all constructs, the ribosome-binding site and start codon were derived from the promoters used. In all constructs, the inserts were downstream of a transcription terminator on shuttle vectors pAM504, pAM505, and pAM2770 (23,32,36). All plasmid constructs were confirmed by sequencing.…”

Section: Methodsmentioning

confidence: 99%

patS Minigenes Inhibit Heterocyst Development of Anabaena sp. Strain PCC 7120

Duan

Lee

et al. 2004

J Bacteriol

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The patS gene encodes a small peptide that is required for normal heterocyst pattern formation in the cyanobacterium Anabaena sp. strain PCC 7120. PatS is proposed to control the heterocyst pattern by lateral inhibition. patS minigenes were constructed and expressed by different developmentally regulated promoters to gain further insight into PatS signaling. patS minigenes patS4 to patS8 encode PatS C-terminal 4 (GSGR) to 8 (CDERGSGR) oligopeptides. When expressed by P petE , P patS , or P rbcL promoters, patS5 to patS8 inhibited heterocyst formation but patS4 did not. In contrast to the full-length patS gene, P hepA -patS5 failed to restore a wild-type pattern in a patS null mutant, indicating that PatS-5 cannot function in cell-to-cell signaling if it is expressed in proheterocysts. To establish the location of the PatS receptor, PatS-5 was confined within the cytoplasm as a gfp-patS5 fusion. The green fluorescent protein GFP-PatS-5 fusion protein inhibited heterocyst formation. Similarly, full-length PatS with a C-terminal hexahistidine tag inhibited heterocyst formation. These data indicate that the PatS receptor is located in the cytoplasm, which is consistent with recently published data indicating that HetR is a PatS target. We speculated that overexpression of other Anabaena strain PCC 7120 RGSGR-encoding genes might show heterocyst inhibition activity. In addition to patS and hetN, open reading frame (ORF) all3290 and an unannotated ORF, orf77, encode an RGSGR motif. Overexpression of all3290 and orf77 under the control of the petE promoter inhibited heterocyst formation, indicating that the RGSGR motif can inhibit heterocyst development in a variety of contexts.

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“…Many genes are specifically involved in heterocyst differentiation. The genes that are required for initiation of cell differentiation include ntcA (15,16) and hetR (17). HetR is a serine-type protease that plays an important role in heterocyst formation (18).…”

mentioning

confidence: 99%

CcbP, a calcium-binding protein from Anabaena sp. PCC 7120, provides evidence that calcium ions regulate heterocyst differentiation

Zhao

Shi

Zhao

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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Although it is known that calcium is a very important messenger involved in many eukaryotic cellular processes, much less is known about calcium's role in bacteria. CcbP, a Ca 2؉ -binding protein, was isolated from the heterocystous cyanobacterium Anabaena sp. PCC 7120, and the ccbP gene was cloned and inactivated. In the absence of combined nitrogen, inactivation of ccbP resulted in multiple contiguous heterocysts, whereas overexpression of ccbP suppressed heterocyst formation. Calmodulin, which is not present in Anabaena species, could also suppress heterocyst formation in both Anabaena sp. PCC 7120 and Anabaena variabilis. HetR induction upon nitrogen step-down was slow in the strain overexpressing ccbP. It has been demonstrated that [Ca 2ϩ ] i is tightly regulated in some bacteria (6), and there is evidence indicating that calcium is critical to some bacterial cellular processes such as sporulation of Bacillus (7), chemotaxis of Escherichia coli (8), and heterocyst differentiation of cyanobacteria (9). Heterocyst development may represent one of the earliest examples of pattern formation in evolution (10). Heterocysts are specialized cells for nitrogen fixation of some filamentous cyanobacteria (11)(12)(13)(14). When heterocystous cyanobacteria are subjected to nitrogen step-down, some vegetative cells differentiate and become heterocysts. Many genes are specifically involved in heterocyst differentiation. The genes that are required for initiation of cell differentiation include ntcA (15, 16) and hetR (17). HetR is a serine-type protease that plays an important role in heterocyst formation (18).Evidence obtained by manipulating extracellular calcium concentration and by using inhibitors of Ca 2ϩ -binding proteins suggested that Ca 2ϩ could be involved in heterocyst differentiation (9). Here we describe a Ca 2ϩ -binding protein, CcbP, from Anabaena sp. PCC 7120. Our study shows that free calcium accumulates in differentiating cells and mature heterocysts, correlated with a drop in the level of the Ca 2ϩ -binding protein.The free Ca 2ϩ concentration appears to be critical for the differentiation process. Materials and MethodsThe following protocols can be found in Supporting Text, which is published as supporting information on the PNAS web site: growth of the strains of Anabaena; isolation of the Ca 2ϩ -binding protein CcbP; isolation of the ccbP gene from total genomic DNA; expression of recombinant ccbP in Escherichia coli; construction of plasmids for inactivation of the ccbP gene, for copper-regulated expression of ccbP, for copper-regulated expression of rat calmodulin (CaM) gene, for ccbP promoter regulation of GFP expression, and for expression of obelin, a reporter for the level of free Ca 2ϩ .Assays for Ca 2؉ -Binding Proteins. 45 Ca 2ϩ overlay assay was performed as follows: Proteins were first separated by SDS͞PAGE and then transferred to a poly(vinylidene difluoride) membrane. It was then washed three times with buffer C (20 mM Tris⅐HCl, pH 7.2͞5 mM MgCl 2 ͞60 mM KCl͞10 mM imidazole). The memb...

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Anabaena sp. strain PCC 7120 ntcA gene required for growth on nitrate and heterocyst development

Cited by 195 publications

References 40 publications

Tansley Review No. 107. Heterocyst and akinete differentiation in cyanobacteria

Tansley Review No. 107. Heterocyst and akinete differentiation in cyanobacteria

patS Minigenes Inhibit Heterocyst Development of Anabaena sp. Strain PCC 7120

CcbP, a calcium-binding protein from Anabaena sp. PCC 7120, provides evidence that calcium ions regulate heterocyst differentiation

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