Identification and characterization of two cofactors (HBP21 and CAB39) in the development and progression of hepatocellular carcinoma

Abstract

Hepatocellular carcinoma (HCC) is one of the most lethal of human malignancies. It is generally believed that hepatocarcinogenesis is a long-term and complicated process accumulating multiple genetic and epigenetic alterations. Unfortunately, HCC is usually diagnosed late and of high aggressiveness. Moreover, even if HCC is diagnosed at early stage, high recurrence rate after surgery and resistance to chemotherapies frequently occurs. Despite intensive studies, the molecular mechanism of progression of HCC remains to be further clarified. A better understanding of the epigenetic and genetic alteration during HCC development and progression is of great importance to early diagnosis and treatment of HCC. In many circumstances, activated oncogenes and inactivated tumor suppressors have clinical significances with acting as prognostic markers and treatment targets for cancer therapeutics. Previously, our laboratory has conducted an integrative transcriptome sequencing analysis on three paired HCC tissues and their corresponding nontumor counterparts to identify differentially expressed genes (Genbank accession no.GSE33294). Cofactors function as interaction scaffolds in assembling multiprotein complexes which regulate various cell processes including gene transcription, cell division, activation of kinases, protein translocation and degradation, etc. Among those differentially expressed genes, two cofactors genes, heat shock binding protein 21 (HBP21) and calcium binding protein 39 (CAB39) were selected for further investigation. The expression levels of two genes were further examined in a larger cohort of HCC clinical samples. It was found that the expression of HBP21 was frequently reduced in primary liver cancer samples (87/120, 72.5%), and this down-regulation was significantly associated with advanced clinical stage (P = 0.049), poor differentiation (P = 0.018) and poor overall survival (P = 0.026). On the contrary, CAB39 was frequently up-regulated in primary HCC samples (65/120, 54.1%), and the up-regulation was significantly associated with metastasis (P=0.049) and poor outcome (P=0.038). Further studies demonstrated that loss of heterozygosity and aberrant promoter methylation are responsible for the reduced expression of HBP21, while expression of CAB39 was regulated by microRNA451 which was commonly down-regulated in HCC samples. Functional study revealed that HBP21 could reduce tumor cell growth rate, decrease the efficiencies and frequencies of foci formation, inhibit colonies formation in soft agar assays and tumor formation in nude mice, in contrast, overexpression of CAB39 in HCC cells has the opposite results in vitro and vivo functional assays. Molecular studies demonstrate that HBP21 can inhibit interaction of HSP70 and Bax, thus increases Bax translocation from cytoplasm to mitochondria, and induces apoptosis. CAB39, a master regulator of STE20 kinases, can activate ERK/c-Jun/Snail/Slug pathway and promote epithelial mesenchymal transition (EMT). In summary, we delineated the novel molecular mechanisms of two cofactors, (HBP21)-(BAX)-(Cyt c) and (CAB39)-(ERK)-(c-JUN)-(SNAIL)-(SLUG) during HCC pathogenesis. Also, the effects of these two cofactors on HCC development and progression were established for the first time in this study. Identification and characterization of their mechanism may shed a light on developing novel biomarkers and targets in HCC diagnosis and treatment.