H-069. Evidence for Involvement of an ECF Sigma Factor in Cyanobacterial Cellular Differentiation
Nostoc punctiforme is a filamentous cyanobacterium capable of cellular differentiation into nitrogen-fixing heterocysts, motile hormogonia, or spore-like akinetes. Akinetes are induced in response to environmental stress and are capable of surviving cold and desiccation. A zwf mutant strain that synchronously differentiates into akinetes upon induction was used in DNA microarray analysis to identify putative genes regulated during akinete differentiation. Two up-regulated genes were homologous to an ECF Sigma Factor (Sig G), and an immediate downstream gene encoding a putative trans-membrane anti-sigma factor. Based on the function of these proteins in other systems, it is hypothesized that together they regulate aspects of cellular differentiation by affecting transcription of target genes in response to an external signal. Two transcriptional start sites were identified upstream from sigG and one upstream from the anti-sigma gene using RACE analysis. Alignments of the promoter region of each gene with the intergenic region of orthologous genes from other akinete-forming cyanobacteria revealed a highly conserved putative regulatory sequence located in the -1 to -37 region of the anti-sigma. Epifluorescence microscopy of promoter GFP reporter strains following akinete induction confirmed akinete-specific gene induction of both genes as indicated from array analysis. Separate reporters for each of the sigG promoters have been used to find the contribution of each promoter in cell-type specific gene expression. PCR mediated ligation was used to create in-frame deletional mutation constructs for each gene. Phenotypic analysis of the resulting mutants will allow determination of their role in akinete formation or function. This is the first evidence for ECF sigma factor involvement in akinete development.