Human selenium binding protein hSP56, a putative chemopreventive protein, binds selenium by a novel mechanism

4054 Human selenium binding protein hSP56, a putative chemopreventive protein, binds selenium by a novel mechanism. Cumulative and comprehensive data from both clinical and epidemiological studies have demonstrated the beneficial effect of selenium as a nutritional supplement in reducing the risk of developing several forms of cancer including prostate cancer. A variety of selenium compounds have been shown to inhibit cancer cell growth or induce apoptosis in culture. However, the molecular targets and mechanisms of this anti-tumor activity of selenium compounds remain largely unknown. We have previously identified genes expressed differentially in human prostate cancer cells with different phenotypes in term of metastatic potential and hormone sensitivity. One of the genes, which encodes human selenium binding protein (hSP56), the human homologue of a rodent gene implicated in chemoresistance, is highly expressed in androgen-sensitive LNCaP cells but not in androgen-insensitive PC-3 cells. This expression in LNCaP cells is reversibly down-regulated by androgen in vitro (Yang, M. and A. J. Sytkowski, Cancer Research, 1998. 58:3150). hSP56 has been proposed to mediate the growth inhibitory action of selenium toward prostate cancer cells (Venkateswaran, V., L.H. Klotz, and N.E. Fleshner Cancer Research, 2002. 62:540). In the present study, we show that human selenium binding protein is synthesized by LNCaP cells as a 56 kDa polypeptide. This hSP56 protein contains selenium as covalently attached component. We demonstrated this by culturing LNCaP cells in the presence of radioactive 75Se and subsequently immunoprecipitating the labeled hSP56 protein with antibodies against hSP56 followed by SDS-PAGE and autoradiography. However, hSP56 is not a selenocysteine-containing protein. Although there is nucleotide sequence in the 3’ untranslated region of the hSP56 gene with suggestive of a selenocysteine insertion sequence (SECIS) characteristic of selenoproteins, we determined that it is not functional in directing selenocysteine insertion. In addition, we have developed an effective method combining isoelectric focusing and immunoblotting and have demonstrated the existence of two isoforms of hSP56 protein isolated from LNCaP cells. The data indicate that hSP56 protein employs a novel mechanism to bind selenium. Elucidation of the molecular mechanism of selenium binding and of the function of hSP56 protein will provide important insights into the chemopreventive action of selenium.