The hetF Gene Product Is Essential to Heterocyst Differentiation and Affects HetR Function in the Cyanobacterium Nostoc punctiforme

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Abstract
A novel gene, hetF , was identified as essential for heterocyst development in the filamentous cyanobacterium Nostoc punctiforme strain ATCC 29133. In the absence of combined nitrogen, hetF mutants were unable to differentiate heterocysts, whereas extra copies of hetF in trans induced the formation of clusters of heterocysts. Sequences hybridizing to a hetF probe were detected only in heterocyst-forming cyanobacteria. The inactivation and multicopy effects of hetF were similar to those of hetR , which encodes a self-degrading serine protease thought to be a central regulator of heterocyst development. Increased transcription of hetR begins in developing cells 3 to 6 h after deprivation for combined nitrogen (N step-down), and the HetR protein specifically accumulates in heterocysts. In the hetF mutant, this increase in hetR transcription was delayed, and a hetR promoter::green fluorescent protein (GFP) transcriptional reporter indicated that increased transcription of hetR occurred in all cells rather than only in developing heterocysts. When a fully functional HetR-GFP fusion protein was expressed in the hetF mutant from a multicopy plasmid, HetR-GFP accumulated nonspecifically in all cells under nitrogen-replete conditions; when expressed in the wild type, HetR-GFP was observed only in heterocysts after N step-down. HetF therefore appears to cooperate with HetR in a positive regulatory pathway and may be required for the increased transcription of hetR and localization of the HetR protein in differentiating heterocysts.