Cloning and characterization of the HSP90 beta gene from Tanichthys albonubes Lin (Cyprinidae): effect of copper and cadmium exposure

Abstract

Heat shock protein 90 (HSP90) is a highly conserved molecular chaperone important in the maturation of a broad spectrum of proteins. In order to evaluate the effect of copper (Cu(2+)) and cadmium (Cd(2+)) on the expression of HSP90 from Tanichthys albonubes (designated TaHSP90), the full-length complementary DNA (cDNA) of TaHSP90 was cloned using reverse transcription PCR and rapid amplification of cDNA ends (RACE) techniques. A 2,687-bp sequence was sequenced and consisted of an open reading frame (ORF) of 2,181bp encoding a polypeptide of 727 amino acids with five HSP90 family signatures. Homologous analysis revealed that TaHSP90 gene shared high similarity with other known HSP90 genes and belonged to HSP90β subtype. Fluorescent real-time quantitative PCR was used to examine the expression pattern of TaHSP90β mRNA in different tissues (liver, muscle, gill, fin, eye, ovary, intestine and brain), and the result indicated that TaHSP90β was widely expressed in all examined tissues at different levels. Sensitivity of TaHSP90β to copper and cadmium was examined by exposing fish to different concentrations of Cu(2+) (0, 13.50 and 27.00μg/L) and Cd(2+) (0, 1.15, 2.31mg/L) for 24, 48, 72 and 96h, respectively. The copper treatment induced TaHSP90β expression slight increase only at 24 and 48h, while cadmium treatment caused slight down-regulation of TaHSP90β only 72 and 96h. Our data suggest that the cloning and expression analysis of T. albonubes HSP90β gene provided useful molecular information of T. albonubes responses in stress conditions and potential ways to monitor the chronic stressors in T. albonubes culture environments.