Modulation of Histone Acetyltransferase Activity through Interaction of Epstein-Barr Nuclear Antigen 3C with Prothymosin Alpha

Abstract

The Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA3C) is essential for EBV-dependent immortalization of human primary B lymphocytes. Genetic analysis indicated that amino acids 365 to 992 are important for EBV-mediated immortalization of B lymphocytes. We demonstrate that this region of EBNA3C critical for immortalization interacts with prothymosin alpha (ProTα), a cellular protein previously identified to be important for cell division and proliferation. This interaction maps to a region downstream of amino acid 365 known to be involved in transcription regulation and critical for EBV-mediated transformation of primary B lymphocytes. Additionally, we show that EBNA3C also interacts with p300, a cellular acetyltransferase. This interaction suggests a possible role in regulation of histone acetylation and chromatin remodeling. An increase in histone acetylation was observed in EBV-transformed lymphoblastoid cell lines, which is consistent with increased cellular gene expression. These cells express the entire repertoire of latent nuclear antigens, including EBNA3C. Expression of EBNA3C in cells with increased acetyltransferase activity mediated by the EBV transactivator EBNA2 results in down-modulation of this activity in a dose-responsive manner. The interactions of EBNA3C with ProTα and p300 provide new evidence implicating this essential EBV protein EBNA3C in modulating the acetylation of cellular factors, including histones. Hence, EBNA3C plays a critical role in balancing cellular transcriptional events by linking the biological property of mediating inhibition of EBNA2 transcription activation and the observed histone acetyltransferase activity, thereby orchestrating immortalization of EBV-infected cells.

ACKNOWLEDGMENTS

We are grateful to S. Elledge for the GAL4 cDNA activation library and the reagents necessary for use in the yeast two-hybrid assay. We thank Elliott Kieff for the EBNA3C reagents and Andrew Cooper, Kenneth Izumi, Jeffrey Lin, and George Mosialos for helpful advice. We also thank Gary Nabel for critical comments during the preparation of the manuscript and the p300 reagents. Jennifer Callahan provided technical assistance on this project. Fernando Dominguez provided the ProTα construct and polyclonal antibody, for which we are grateful. We also extend special thanks to Vojo Deretic and Eric Fearon for use of their fluorescence microscope and digital imaging camera system.

This work was supported by grant AHA9650467N and by a grant from the National Cancer Institute (CA072150-01) (to E.S.R.). E.S.R. is a Scholar of the Leukemia and Lymphoma Society of America. M.A.C. is supported by funds from the Medical Scientist Training Program of the National Institute of General Medical Sciences (NIH5 T32 GM07863) and is a fellow of the Lady Tata Memorial Trust.