Role of regucalcin in liver nuclear function: Binding of regucalcin to nuclear protein or DNA and modulation of tumor-related gene expression

Abstract

Regucalcin is a regulatory protein in the intracellular signaling pathway which is related to regulation of nuclear function. In this study the binding of regucalcin to nuclear proteins or DNA in vitro was examined. The results of the Far-Western analysis showed the existence of protein components which bind to regucalcin in the nucleus isolated from rat liver. Whether regucalcin binds deoxyribonucleic acid (DNA) was analyzed using DNA cellulose in vitro. Regucalcin was incubated in reaction mixture containing DNA cellulose, and DNA-binding regucalcin was detected using Western blot analysis for regucalcin. The results showed that regucalcin binds DNA in vitro. Moreover, the expression of c-src, p53, and Rb mRNAs was examined in the cloned rat hepatoma H4-II-E cells cultured for 24 or 48 h in the presence of fetal bovine serum (10%), using reverse transcription-polymerase chain reaction (RT-PCR). The expression of oncogene c-src mRNA was significantly suppressed in the hepatoma cells (transfectants), overexpressing regucalcin. Meanwhile, the expression of the tumor suppressor gene p53 or Rb mRNA was significantly enhanced in transfectants. This study may support the view that regucalcin modulates the transcriptional process by binding to protein and DNA in the nucleus.