NADPH oxidase homologs are required for normal cell differentiation and morphogenesis in Dictyostelium discoideum

Membrane-associated NADPH oxidase complexes catalyse the production of the superoxide anion radical from oxygen and NADPH. In mammalian systems, NADPH oxidases form a family of at least seven isoforms that participate in host defence and signalling pathways. We report here the cloning and the characterisation of slime mould Dictyostelium discoideum homologs of the mammalian heme-containing subunit of flavocytochrome b (gp91phox) (NoxA, NoxB and NoxC), of the small subunit of flavocytochrome b (p22phox) and of the cytosolic factor p67phox. Null-mutants of either noxA, noxB, noxC or p22phox show aberrant starvation-induced development and are unable to produce spores. The overexpression of NoxAmyc2 in noxA null strain restores spore formation. Remarkably, the gene alg-2B, coding for one of the two penta EF-hand proteins in Dictyostelium, acts as a suppressor in noxA, noxB, and p22phox null-mutant strains. Knockout of alg-2B allows noxA, noxB or p22phox null-mutants to return to normal development. However, the knockout of gene encoding NoxC, which contains two penta EF-hands, is not rescued by the invalidation of alg-2B. These data are consistent with a hypothesis connecting superoxide and calcium signalling during Dictyostelium development.