Molecular cloning, modeling and differential expression of a gene encoding a silent information regulator-like protein from Sporothrix schenckii.

Abstract

Sporothrix schenckii (S. schenckii) is a dimorphic fungus that produces lymphocutaneous lesions. The signature characteristic of S. schenckii is a temperature-induced phase transition. Silent information regulator (Sir) has been proven to be involved in phenotypic switching in Saccharomyces cerevisiae (S. cerevisiae) and Candida albicans (C. albicans) by organizing chromatin structure. In this study, we isolated and characterized a Sir homologue gene, designated as SsSir2, from the yeast form of S. schenckii. The full-length SsSir2 cDNA sequence is 1753 bp in size and contains an open reading frame of 1329 bp encoding 442 amino acids. The predicted molecular mass of SsSir2 is 48.1 kDa with an estimated theoretical isoelectric point of 4.6. The SsSir2 kinase domain shows a 78% identity with that of Hst2, a Sir2 Ib gene from S. cerevisiae. Three exons and two introns were identified within the 1472-bp SsSir2 genomic DNA sequence of S. schenckii. A three-dimensional model of SsSir2 was constructed using a homology modeling method, and its reliability was evaluated. The active site of SsSir2 was identified by docking simulation, which indicated that several important residues, such as Asn127 and Asp129, play an important role in the histone deacetylase activity of Sir2 family proteins. The differential expression of the SsSir2 in two stages was demonstrated by real-time RT-PCR. The expression of SsSir2 was higher in the yeast stage compared with that in the mycelial one, which indicated that SsSir2 may be involved in the phenotypic switching and morphogenesis of the yeast phase in S. schenckii.