Abstract
Apobec-1 complementation factor (A1CF) functions as an RNA-binding cofactor for APO-BEC1-mediated C-to-U conversion during RNA editing and as a hepatocyte-specific regulator in the alternative pre-mRNA splicing of metabolic enzymes. Its role in RNA editing has not been clearly established. Western blot, co-immunoprecipitation (Co-IP), immunofluorescence (IF), methyl thiazolyl tetrazolium (MTT), and 5-ethynyl-2'-deoxyuridine (EdU) assays were used to examine the role of A1CF beyond RNA editing in renal carcinoma cells. We demonstrated that A1CF interacts with NKRF, independent of RNA and DNA, without affecting its expression or nuclear translocation; however, it modulates p65(Ser536) phosphorylation and IFN-β levels. Truncation of A1CF or deletion on NKRF revealed that the RRM1 domain of A1CF and the p65 binding motif of NKRF are required for their interaction. Deletion of RRM1 on A1CF abrogates NKRF binding, and the decrease in IFN-β expression and p65(Ser536) phosphorylation was induced by A1CF. Moreover, full-length A1CF, but not an RRM1 deletion mutant, promoted cell proliferation in renal carcinoma cells. Perturbation of A1CF levels in renal carcinoma cells altered anchorage-independent growth and tumor progression in nude mice. Moreover, p65(Ser536) phosphorylation and IFN-β expression were lower, but ki67 was higher in A1CF-overexpressing tumor tissues of a xenograft mouse model. Notably, primary and metastatic samples from renal cancer patients exhibited high A1CF expression, low p65(Ser536) phosphorylation, and decreased IFN-β levels in renal carcinoma tissues compared with the corresponding paracancerous tissues. Our results indicate that A1CF-decreased p65(Ser536) phosphorylation and IFN-β levels may be caused by A1CF competitive binding to the p65-combined site on NKRF and demonstrate the direct binding of A1CF independent of RNA or DNA in signal pathway regulation and tumor promotion in renal carcinoma cells.