The identification of transcriptional regulation related gene of laccase poxc through yeast one-hybrid screening from Pleurotus ostreatus.

Pleurotus ostreatus laccase gene poxc is transcriptionally induced by copper, and several putative MREs have been found and confirmed in its promoter region. However, the related transcription factors mediating copper response via MREs have not been reported. To isolate MRE binding proteins, we carried out yeast one-hybrid screens. Reporter genes containing two trimers of the cMRE2 and cMRE3 element were prepared and introduced into a yeast strain. The yeast was transformed with library cDNA that represents RNA isolated from CuSO4-treated fungi of P. ostreatus. From the screen of yeast transformants, we isolated ltf4 which encoded protein with HTH DNA binding domain. Electrophoretic mobility shift assay suggested direct and specific interaction of ltf4 with the MRE2 of poxc. Quantitative RT-PCR showed that the transcription of ltf4 was significantly up-regulated after the copper addition, and the expression trend was consistent with the poxc after copper introduction. The results indicated that ltf4 could interact with cMRE2 and thus participated in the regulation of copper mediated poxc transcription in P. ostreatus.