Functional Overlap of hetP and hetZ in Regulation of Heterocyst Differentiation in Anabaena sp. Strain PCC 7120

Abstract

HetR plays a key role in regulation of heterocyst differentiation and patterning in Anabaena It directly regulates genes involved in heterocyst differentiation (such as hetP and hetZ), genes involved in pattern formation (patA), and many others. In this study, we investigated the functional relationship of hetP and hetZ and their role in HetR-dependent cell differentiation. Coexpression of hetP and hetZ from the promoter of ntcA, which encodes the global nitrogen regulator, enabled a hetR mutant to form heterocysts with low aerobic nitrogenase activity. Overexpression of hetZ restored heterocyst differentiation in a hetP mutant and vice versa. Overexpression of hetR restored heterocyst formation in either a hetP or a hetZ mutant but not in a hetZ hetP double mutant. The functional overlap of hetP and hetZ was further confirmed by reverse transcription-quantitative PCR (RT-qPCR) and transcriptomic analyses of their effects on gene expression. In addition, yeast two-hybrid and pulldown assays showed the interaction of HetZ with HetR. HetP and HetZ are proposed as the two major factors that control heterocyst formation in response to upregulation of hetRIMPORTANCE Heterocyst-forming cyanobacteria contribute significantly to N₂ fixation in marine, freshwater, and terrestrial ecosystems. Formation of heterocysts enables this group of cyanobacteria to fix N₂ efficiently under aerobic conditions. HetR, HetP, and HetZ are among the most important factors involved in heterocyst differentiation. We present evidence for the functional overlap of hetP and hetZ and for the key role of the HetR-HetP/HetZ circuit in regulation of heterocyst differentiation. The regulatory mechanism based on HetR, HetP, and HetZ is probably conserved in all heterocyst-forming cyanobacteria.

Keywords: Anabaena; heterocyst differentiation; regulatory circuit.