A point mutation in nucleoside diphosphate kinase results in a deficient light response for perithecial polarity in...

In Neurospora crassa, the phosphorylation of nucleoside diphosphate kinase (NDK)-1 is rapidly enhanced after blue light irradiation. We have investigated the function of NDK-1 in the blue light signal transduction pathway. A mutant called psp (phosphorylation of small protein) shows undetectable phosphorylation of NDK-1 and is defective in light-responsive regulation of perithecial polarity. Sequencing analysis of ndk-1 cDNA by reverse transcription-polymerase chain reaction revealed that proline 72 of ndk-1 was replaced with histidine in psp. The mutation ndk-1(P72H) resulted in accumulation of normal levels of mRNA and of about 25% of NDK-1(P72H) protein compared with that of wild type as determined by Western blot analysis. The ectopic expression of cDNA and introduction of genomic DNA of wild type ndk-1 in psp (ndk-1(P72H)) suppressed the reduction in accumulation and phosphorylation of NDK-1 and the light-insensitive phenotype. These findings demonstrated that the phenotype of psp was caused by the ndk-1(P72H) mutation. Biochemical analysis using recombinant NDK-1 and NDK-1(P72H) indicated that the P72H substitution in NDK-1 was responsible for the decrease in phosphotransfer activities, 5% of autophosphorylation activity, and 2% of V(max) for protein kinase activity phosphorylating myelin basic protein, compared with those of wild type NDK-1, respectively.