Tmac1, a transcription factor which regulated high affinity copper transport in Trichoderma reesei

Abstract

The Mac1 protein was a transcriptional activator that sensed very low concentration of copper and regulated the copper transport in Saccharomyces cerevisiae. Here, we cloned a gene from Trichoderma reesei named Tmac1, whose deduced amino acid sequence showed 29% identical to Mac1p. Furthermore, two Cys-His repeats metal binding motifs of Tmac1p, one in the 354-369C terminus and one in the 475-490C terminus were also present in Mac1p. A deletion mutant of Tmac1 was hypersensitive to the copper starvation and showed poor growth. Subsequently, the function was recovered by the gene complementation experiment. Furthermore, the Tmac1 gene fully complemented growth defects of yeast ΔMac1 mutant. The expression of Tmac1p was activated at low concentration of copper and depressed when the concentration of copper excess 1mM. Furthermore, the fluorescence intensity enhanced at copper starvation and decreased under copper excess by fusion the eGFP to the Tmac1p. It proved that the expression of Mac1p was exactly regulated by copper concentration, because eGFP and Mac1p were expressed under the control of the same one promoter. We also cloned a gene named Tctr3 with bioinformatics. With a series of experiments, we proved it was the target gene of Tmac1. To sum up, Tmac1 may encode a transcriptional activator regulated high-affinity copper transport in T. reesei.