Absract: BACKGROUND: Tumor metastasis is the major reason for poor prognosis of hepatocellular carcinoma (HCC) patients after hepatic resection. SMYD3 has been demonstrated to promote liver tumor metastasis in mice. However, the detailed molecular mechanism is still largely unknown.
Methods: The effect of SMYD3 on invasiveness and metastasis of HCC was analyzed by immunohistochemistry, migration assay, invasion assay, wound healing assay and in vivo lung metastasis assay. Mass spectrometry analysis was conducted using proteins pulled down by H3K4me3 antibody in SMYD3-overexpressing cells. Luciferase reporter, chromatin immunoprecipitation, Electrophoretic mobility shift assay were used to measure the regulation of SLUG transcription by SMYD3-ANKHD1. In addition, the role of SMYD3-ANKHD1 in determining clinical outcomes for HCC patients was investigated by immunohistochemistry in 243 HCC tissues.
Results: SMYD3 was an independent prognostic factor of HCC and promoted migration and invasion of human HCC cells. ANKHD1 was identified by mass spectrometry as a co-regulator with SMYD3. ANKHD1 interacted with H3K4me3 when cells were overexpressing SMYD3. The pro-migratory and pro-invasive effects of SMYD3 were attenuated when ANKHD1 was knocked down by siRNA. Furthermore, we found that SMYD3 bound and activated the SLUG gene promoter in a manner associated with elevating H3K4me3, H3K9Ac and H3K14Ac. Knockdown of ANKHD1 could attenuate the SMYD3-dependent activation of Slug expression. We further detected the expression of SMYD3 and ANKHD1 in 243 HCC patients and found that patients with positive coexpression of SMYD3 and ANKHD1 (SMYD3+ANKHD1+) had the shortest overall and recurrence-free survival.
Conclusion: Our findings provide a novel molecular mechanism for the SMYD3-regulated HCC migration and metastasis, and indicates that SMYD3-ANKHD1 may be a potential target for treating HCC.
Keywords: ANKHD1; Hepatocellular carcinoma; Metastasis; SMYD3; Slug.