A two-component system mediates developmental regulation of biosynthesis of a heterocyst polysaccharide

Some cyanobacteria couple oxygenic photosynthesis in vegetative cells with O-2-sensitive N-2 fixation in differentiated cells called heterocysts. Heterocyst differentiation involves extensive biochemical and structural changes that collectively permit heterocysts to assimilate N-2 aerobically and supply the products of N-2 fixation to vegetative cells. HepK and DevR are required for the development of functional heterocysts in Anabaena and Nostoc, respectively. We show that HepK is an autokinase and that Anabaena DevR(A) is its cognate response regulator, together comprising part or all of a two-component system that mediates developmental regulation of biosynthesis of a heterocyst envelope polysaccharide. Recombinant N-hexahistidine-tagged HepK (H(6)HepK), the cytoplasmic portion H-6'HepK of H(6)HepK, H(6)DevR, and H(6)DevR(A) were overexpressed in Escherichia coli and purified to homogeneity. H-6'HepK, but not H(6)HepK, autophosphorylates with [gamma-P-32]ATP. ADP, specifically, elicits dephosphorylation of phosphorylated H-6'HepK. The phosphoryl group of H-6'HepK is transferred rapidly and efficiently to both H(6)DevR and H(6)DevRA but not to His-tagged OmpR, whose cognate sensor kinase is EnvZ. Sequence comparisons, the results of site-specific mutagenesis, and tests of chemical stability support identification of HepK-His(348) and DevR-Asp(53) as the phosphorylated residues. The mutation HepK-H348A abolishes both in vitro autokinase activity and in vivo functionality of HepK. Heterocysts of both hepK Anabaena and devR(A) Anabaena lack an envelope polysaccharide layer and are nonfunctional. Consistent with the normal site of deposition of that polysaccharide, a hepK::gfp transcriptional fusion is expressed principally in proheterocysts. HepK/DevR(A) is the first two-component system identified that regulates the biosynthesis of a polysaccharide as part of a patterned differentiation process.