The Neurospora crassa Nuclear Dbf2-related (NDR) Ser/Thr kinase- COT1 is important for polar growth and normal hyphal development. COT1 regulates elongation and branching in an independent manner which is determined by its phosphorylation state. PP2A is a heterotrimeric Ser/Thr phosphatase protein that consists of scaffolding subunit A, catalytic subunit C and variable regulatory subunits B. PP2A plays an important role in regulation of growth and development in N. crassa and other fungi. pph-1, which encodes PP2Ac, is essential in N. crassa and deletion of the PP2A B regulatory subunits rgb-1 and b-56 impairs hyphal growth, branching and conidiation. On the basis of co-immunoprecipitation experiments we determined that the PP2A catalytic subunit and COT1 can physically interact. We also determined the presence of a genetic interaction between the PP2A regulatory subunit rgb-1 and cot-1(S189E), cot-1(S417E) and cot-1(T589E), all of which are alleles that mimic constitutive phosphorylation on the respective residues. In all cases, the presence of the phosphomimetic allele suppressed the slow growth phenotype of rgb-1RIP, indicating a functional link between the two gene products, on the basis of COT1 phosphorylation state. In contrast, none of the cot-1 phosphomimetic mutants suppressed the slow growth of the b-56 regulatory subunit mutant. We conclude that PP2A catalytic subunit as well as the regulatory subunit RGB1, but not B56, interact with COT1. Based on the differential interactions with cot-1 phosphomimetic mutants, the regulatory subunits have distinct functions in regulation of growth and development in N. crassa.
