EBNA3C attenuates the function of p53 through interaction with inhibitor of growth family proteins 4 and 5.

Epstein-Barr virus (EBV)-encoded EBNA3C is one of the latent proteins essential for the efficient transformation of human primary B lymphocytes into continuously proliferating lymphoblastoid cell lines (LCLs) in vitro through manipulation of a number of major cellular pathways. Although it does not have direct DNA-binding activity, EBNA3C plays a central role in the transcriptional modulation of a wide range of both viral and cellular genes during latent infection. Recently, we showed that EBNA3C can directly bind to the tumor suppressor protein p53 and repress its functions, in part by blocking its transcriptional activity as well as facilitating its degradation through stabilization of its negative regulator, Mdm2. In this study, we further showed that EBNA3C can negatively regulate p53-mediated functions by interacting with its regulatory proteins, the inhibitor of growth family proteins ING4 and ING5, shown to be frequently deregulated in different cancers. Functional mapping revealed that both ING4 and ING5 bound to N-terminal domain residues 129 to 200 of EBNA3C, which was previously demonstrated to associate with p53 and is also essential for LCL growth. In addition, we showed that a conserved domain of either ING4 or ING5 bound to both p53 and EBNA3C in a competitive manner, suggesting a potential role for EBNA3C whereby the ING4 or -5/p53 pathway is modulated in EBV-infected cells. Subsequently, we demonstrated that EBNA3C significantly suppresses both the ING4- and ING5-mediated regulation of p53 transcriptional activity in a dose-dependent manner. A colony formation assay as well as an apoptosis assay showed that EBNA3C nullified the negative regulatory effects on cell proliferation induced by coupled expression of p53 in the presence of either ING4 or ING5 in Saos-2 (p53(-/-)) cells. This report demonstrates a possible role for the candidate tumor suppressor ING genes in the biology of EBV-associated cancers.