[Show abstract][Hide abstract] ABSTRACT: The ubiquitin-like protein FAT10 and the homeobox protein HOXB9 each promote metastatic progression in hepatocellular carcinoma (HCC). In this study, we investigated the clinicopathologic significance of FAT10 and HOXB9 in HCC and investigated a mechanistic role for FAT10 in HOXB9-mediated invasiveness and metastasis. Relative to adjacent normal tissues, FAT10 and HOXB9 were markedly overexpressed in HCC, where a positive correlation in their expression and associated malignant characteristics were found. RNAi-mediated silencing of FAT10 decreased HOXB9 expression and inhibited HCC invasion and metastasis in vitro and in vivo. The effects of FAT10 silencing were reversed by HOXB9 overexpression, whereas RNAi-mediated silencing of HOXB9 decreased HCC invasion and metastasis driven by FAT10 overexpression. Mechanistically, FAT10 regulated HOXB9 expression by modulating the β-catenin/TCF4 pathway, directly binding to β-catenin and preventing its ubiquitination and degradation. Together, our results identiﬁed a novel HCC regulatory circuit involving FAT10, β-catenin/TCF4 and HOXB9, the dysfunction of which drives invasive and metastatic character in HCC.
Cancer Research 07/2014; 74(18). DOI:10.1158/0008-5472.CAN-14-0284 · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: FAT10 is known to execute its functions mainly through conjugation to different substrates, and these known functions include cytokine responses, apoptosis, mitosis, and tumorigenesis. Nonetheless, the known binding proteins of FAT10 cannot explain all its known functions. As such, the aim of this study was to identify unidentified conjugation proteins of FAT10.
The yeast two-hybrid system was employed in this study. FAT10 was used as the bait protein for screening of a cDNA library from a human hepatocellular carcinoma cell line, Hep3B. Protein interactions were confirmed based on localization studies and co-immunoprecipitation assays. The expression of mRNA and protein was determined using real-time polymerase chain reaction and western blot analyses, respectively.
In this study, we identified eukaryotic elongation factor 1A1 (eEF1A1) as a FAT10-specific binding protein. The binding between FAT10 and eEF1A1 was confirmed both in vivo and in vitro. We also found that, when the expression of FAT10 was reduced by siRNA knockdown, this resulted in downregulation of eEF1A1 expression at both the mRNA and protein levels in human hepatocellular carcinoma cells.
We propose a model in which eEF1A1 serves as a substrate of FAT10 to accomplish, in part, its functions in regulating the biological behavior of tumor cells. Since both eEF1A1 and FAT10 are important for tumorigenesis and development, comprehending the mechanisms of this interaction can provide clues for identification of novel strategic targets for drug screening and molecular typing, and possibly in the development of new effective therapeutic strategies against hepatocellular carcinoma.