title: The sigE gene is required for normal expression of heterocyst-specific genes in Anabaena sp. strain PCC 7120.
abstract: The filamentous cyanobacterium Anabaena (Nostoc) sp. strain PCC 7120 produces specialized cells for nitrogen fixation called heterocysts. Previous work showed that the group 2 sigma factor sigE (alr4249; previously called sigF) is upregulated in differentiating heterocysts 16 h after nitrogen step-down. We now show that the sigE gene is required for normal heterocyst development and normal expression levels of several heterocyst-specific genes. Mobility shift assays showed that the transcription factor NtcA binds to sites in the upstream region of sigE and that this binding is enhanced by 2-oxoglutarate (2-OG). Deletions of the region containing the NtcA binding sites in P(sigE)-gfp reporter plasmids showed that the sites contribute to normal developmental regulation but are not essential for upregulation in heterocysts. Northern RNA blot analysis of nifH mRNA revealed delayed and reduced transcript levels during heterocyst differentiation in a sigE mutant background. Quantitative reverse transcription-PCR (qRT-PCR) analyses of the sigE mutant showed lower levels of transcripts for nifH, fdxH, and hglE2 but normal levels for hupL. We developed a P(nifHD)-gfp reporter construct that showed strong heterocyst-specific expression. Time-lapse microscopy of the P(nifHD)-gfp reporter in a sigE mutant background showed delayed development and undetectable green fluorescent protein (GFP) fluorescence. Overexpression of sigE caused accelerated heterocyst development, an increased heterocyst frequency, and premature expression of GFP fluorescence from the P(nifHD)-gfp reporter.
authors.affiliation: Department of Biology, Texas A&M University, College Station, Texas 77843-3258, USA.
authors.forename: Rodrigo A
authors.initials: RA
authors.lastname: Mella-Herrera
authors.forename: M Ramona
authors.initials: MR
authors.lastname: Neunuebel
authors.forename: Krithika
authors.initials: K
authors.lastname: Kumar
authors.forename: Sushanta K
authors.initials: SK
authors.lastname: Saha
authors.forename: James W
authors.initials: JW
authors.lastname: Golden
mesh_terms: D017033:Anabaena / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015152:Blotting, Northern; D004269:DNA, Bacterial / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D017353:Gene Deletion; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018715:Microscopy, Video; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D012808:Sigma Factor; C111259:sigE protein, Bacteria; D049452:Green Fluorescent Proteins