Nitric oxide synthase is induced in sporulation of Physarum polycephalum

The myxomycete Physarum polycephalum expresses a calcium-independent nitric oxide (NO) synthase (NOS) resembling the inducible NOS isoenzyme in mammal s. We have now cloned and sequenced this, the first nonanimal NOS to be ide ntified, showing that it shares < 39% amino acid identity with known NOSs b ut contains conserved binding motifs for all NOS cofactors. It lacks the se quence insert responsible for calcium dependence in the calcium-dependent N OS isoenzymes. NOS expression was strongly up-regulated in Physarum macropl asmodia during the 5-day starvation period needed to induce sporulation com petence. Induction of both NOS and sporulation competence were inhibited by glucose, a growth signal and known repressor of sporulation, and by L-N6-( 1-iminoethyl)-lysine (NIL), an inhibitor of inducible NOS. Sporulation, whi ch is triggered after the starvation period by light exposure, was also pre vented by 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1 (ODQ), an inhibitor of N O-sensitive guanylate cyclase. In addition, also expression of lig1, a spor ulation-specific gene, was strongly attenuated by NIL or ODQ. 8-Bromo-cGMP, added 2 h before the light exposure, restored the capacity of NIL-treated macroplasmodia to express lig1 and to sporulate. This indicates that the se cond messenger used for NO signaling in sporulation of Physarum is cGMP and links this signaling pathway to expression of lig1.