HDAC 1 and 6 modulate cell invasion and migration in clear cell renal cell carcinoma

Abstract

Background: Class I histone deacetylases (HDACs) have been reported to be overexpressed in clear cell renal cell carcinoma (ccRCC), whereas the expression of class II HDACs is unknown. Methods: Four isogenic cell lines C2/C2VHL and 786-O/786-OVHL with differential VHL expression are used in our studies. Cobalt chloride is used to mimic hypoxia in vitro. HIF-2α knockdowns in C2 and 786-O cells is used to evaluate the effect on HDAC 1 expression and activity. Invasion and migration assays are used to investigate the role of HDAC 1 and HDAC 6 expression in ccRCC cells. Comparisons are made between experimental groups using the paired T-test, the two-sample Student's T-test or one-way ANOVA, as appropriate. ccRCC and the TCGA dataset are used to observe the clinical correlation between HDAC 1 and HDAC 6 overexpression and overall and progression free survival. Results: Our analysis of tumor and matched non-tumor tissues from radical nephrectomies showed overexpression of class I and II HDACs (HDAC6 only in a subset of patients). In vitro, both HDAC1 and HDAC6 over-expression increased cell invasion and motility, respectively, in ccRCC cells. HDAC1 regulated invasiveness by increasing matrix metalloproteinase (MMP) expression. Furthermore, hypoxia stimulation in VHL-reconstituted cell lines increased HIF isoforms and HDAC1 expression. Presence of hypoxia response elements in the HDAC1 promoter along with chromatin immunoprecipitation data suggests that HIF-2α is a transcriptional regulator of HDAC1 gene. Conversely, HDAC6 and estrogen receptor alpha (ERa) were co-localized in cytoplasm of ccRCC cells and HDAC6 enhanced cell motility by decreasing acetylated a-tubulin expression, and this biological effect was attenuated by either biochemical or pharmacological inhibition. Finally, analysis of human ccRCC specimens revealed positive correlation between HIF isoforms and HDAC. HDAC1 mRNA upregulation was associated with worse overall survival in the TCGA dataset. Conclusions: Taking together, these results suggest that HDAC1 and HDAC6 may play a role in ccRCC biology and could represent rational therapeutic targets.