The Hybrid Histidine Kinase HrmK Is an Early-Acting Factor in the Hormogonium Gene Regulatory Network.

Filamentous, heterocyst-forming cyanobacteria belonging to taxonomic subsections IV and V are developmentally complex multicellular organisms capable of differentiating an array of cell and filament types, including motile hormogonia. Hormogonia exhibit gliding motility that facilitates dispersal, phototaxis, and the establishment of nitrogen-fixing symbioses. The gene regulatory network (GRN) governing hormogonium development involves a hierarchical sigma factor cascade, but the factors governing the activation of this cascade are currently undefined. Here, using a forward genetic approach, we identified hrmK, a gene encoding a putative hybrid histidine kinase that functions upstream of the sigma factor cascade. The deletion of hrmK produced nonmotile filaments that failed to display hormogonium morphology or accumulate hormogonium-specific proteins or polysaccharide. Targeted transcriptional analyses using reverse transcription-quantitative PCR (RT-qPCR) demonstrated that hormogonium-specific genes both within and outside the sigma factor cascade are drastically downregulated in the absence of hrmK and that hrmK may be subject to indirect, positive autoregulation via sigJ and sigC Orthologs of HrmK are ubiquitous among, and exclusive to, heterocyst-forming cyanobacteria. Collectively, these results indicate that hrmK functions upstream of the sigma factor cascade to initiate hormogonium development, likely by modulating the phosphorylation state of an unknown protein that may serve as the master regulator of hormogonium development in heterocyst-forming cyanobacteria.IMPORTANCE Filamentous cyanobacteria are morphologically complex, with several representative species amenable to routine genetic manipulation, making them excellent model organisms for the study of development. Furthermore, two of the developmental alternatives, nitrogen-fixing heterocysts and motile hormogonia, are essential to establish nitrogen-fixing symbioses with plant partners. These symbioses are integral to global nitrogen cycles and could be artificially recreated with crop plants to serve as biofertilizers, but to achieve this goal, detailed understanding and manipulation of the hormogonium and heterocyst gene regulatory networks may be necessary. Here, using the model organism Nostoc punctiforme, we identify a previously uncharacterized hybrid histidine kinase that is confined to heterocyst-forming cyanobacteria as the earliest known participant in hormogonium development.