Ectopic expression of hetP can partially bypass the need for hetR in heterocyst differentiation by Anabaena sp. strain PCC 7120

Abstract

SummaryDifferentiation of nitrogen-fixing cells, called heterocysts, by the cyanobacterium Anabaena sp. strain PCC 7120 requires HetR, which is considered the master regulator of heterocyst differentiation. In this study, ectopic expression of hetP from an inducible promoter was shown to partially bypass the need for hetR in heterocyst differentiation. A hetR-deletion strain with hetP expressed ectopically produced cells morphologically similar to heterocysts that produced exopolysaccharide and glycolipids specific to heterocysts, and nitrogenase activity was present under anaerobic conditions, as indicated by an acetylene reduction assay. Five potential transcription start points or transcript processing sites located at positions -727, -545, -208, -177 and -12 bp relative to the putative translational start site were identified in the promoter region of hetP. Relative levels of transcription and their location in filaments resembled that previously reported for HetR protein in various genetic backgrounds, and an inverted repeat sequence in the promoter of hetP was necessary for binding of HetR. Two of the three identified homologues of hetP found in PCC 7120 partially complemented a hetP-null mutant. Taken together, these results suggest that HetP functions directly downstream of HetR in the regulatory network responsible for heterocyst differentiation.

The HetR-binding site that activates expression of patA in vegetative cells is required for normal heterocyst patterning in Anabaena sp. PCC 7120

“…HetR has been shown to directly up-regulate hetP and hetZ, two genes that are involved early in the differentiation of heterocysts [10,17], and hepA, a gene involved in formation of the heterocyst envelope polysaccharide layer [19].…”

“…Like P hetP -gfp [17] and P hetZ -gfp [10], P patA -gfp exhibited intensified expression in vegetative cells in a patA-null mutant and a gradient of GFP fluorescence near the terminal heterocysts. The GFP gradient should be due to the HetR-dependent transcription in P patA .…”

“…So far, four modes of regulation via HetR-binding sites have been identified: (1) up-regulating gene expression in heterocysts [10,17]; (2) supporting gene expression in vegetative cells (such as patA); (3) suppressing gene expression in heterocysts (such as alr0202); and (4) suppressing gene expression in both types of cells (such as alr3234; [16,19] ). In some cases, deletion of HetRbinding sites affects the expression of downstream genes similarly as inactivation of hetR.…”

“…[15,16]. HetR binds to consensus recognition sites upstream of hetP [17] and hetZ [10], two genes required for heterocyst differentiation and up-regulated in developing heterocysts. The crystal structure of the HetR-DNA complex has been reported, showing the importance of GGG(n5)CCC and several flanking base pairs in the interaction [18].…”

The pknH gene restrictively expressed in heterocysts is required for diazotrophic growth in the cyanobacterium Anabaena sp. strain PCC 7120

“…HetR is a master regulator of heterocyst development, and plays a key role in differentiation and pattern formation (Buikema & Haselkorn, 1991;Buikema & Haselkorn, 2001). HetR is thought to activate expression of many genes involved in heterocyst development (Higa & Callahan, 2010;Huang et al, 2004;Saha & Golden, 2011). been recognized in the last decade.…”