Cloning and sequence analysis of the basidiomycete Lentinus edodes ribonucleotide reductase small subunit cDNA and expression of a corresponding gene in L. edodes.

We have previously isolated the uck1 gene encoding UMP-CMP kinase from the basidiomycete Lentinus edodes (Kaneko et al., 1998). It was shown to be most actively transcribed in hymenophores of mature fruiting bodies of L. edodes. The reduction of NDPs produced by the nucleoside monophosphate kinase to dNDPs has been known to be catalyzed by ribonucleotide reductase (RNR) which consists of a heterodimer of large and small subunits. So we attempted to isolate the L. edodes cDNA(s) of RNR and study the expression in L. edodes of the corresponding gene(s), resulting in an isolation of the small subunit cDNA from a mature fruiting-body cDNA library of the fungus. This cDNA, named Le.rnr2c, was shown to encode a 418 amino acids (aa) protein, named Le.RNR2, of which the deduced aa sequence shows an overall identity of 71.9% to that of Schizosaccharomyces pombe RNR small subunit. The Le.rnr2 gene was found to be most actively transcribed in hymenophores of mature fruiting body of L. edodes. The in situ RNA-RNA hybridization analysis showed the presence of markedly large amount of the Le.rnr2 transcript in both hymenium and outer region of trama in the hymenophore. The same experiment was done for the uck1 gene, obtaining a similar result. The hymenium contains many basidia in which fusion of two nuclei, meiosis, replication, etc. essential for production of basidiospores occur. The outer region of trama is the region branching out into subhymenium. These imply that Le.rnr2 gene (and uck1 gene) play a role mainly in the nucleotide biosynthesis essential both for production of basidiospores and for divergence of trama cells into subhymenium cells in the hymenophore.