E2F1 mediated apoptosis induced by the DNA damage response is blocked by EBV nuclear antigen 3C in lymphoblastoid cells

• Published in: PLoS pathogens Vol. 8; no. 3; p. e1002573
• Main Authors: Saha, Abhik, Lu, Jie, Morizur, Lise, Upadhyay, Santosh K, Aj, Mahadesh Prasad, Robertson, Erle S
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01-03-2012; Public Library of Science (PLoS)
• Subjects: Antigens; Antigens, Viral - genetics; Antigens, Viral - metabolism; Apoptosis; Apoptosis - genetics; Biology; Cell division; Cell Line; Cell Line, Tumor; Cell lines; Cell Proliferation; Colleges & universities; DNA Damage; E2F1 Transcription Factor - antagonists & inhibitors; E2F1 Transcription Factor - metabolism; Enzymes; Epstein-Barr virus; Epstein-Barr Virus Infections - genetics; Epstein-Barr Virus Infections - metabolism; Epstein-Barr Virus Infections - virology; Epstein-Barr Virus Nuclear Antigens; Gene expression; Gene Expression Regulation, Viral; Genetic aspects; Growth rate; Health aspects; HEK293 Cells; Humans; Lymphocytes - metabolism; Lymphocytes - pathology; Lymphocytes - virology; Lymphoma; Lymphoma, B-Cell - immunology; Lymphoma, B-Cell - virology; Osteoblasts - immunology; Osteoblasts - virology; Phosphorylation; Physiological aspects; Proteins; Transfection; United States
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1002573

EBV latent antigen EBNA3C is indispensible for in vitro B-cell immortalization resulting in continuously proliferating lymphoblastoid cell lines (LCLs). EBNA3C was previously shown to target pRb for ubiquitin-proteasome mediated degradation, which facilitates G1 to S transition controlled by the major transcriptional activator E2F1. E2F1 also plays a pivotal role in regulating DNA damage induced apoptosis through both p53-dependent and -independent pathways. In this study, we demonstrate that in response to DNA damage LCLs knocked down for EBNA3C undergo a drastic induction of apoptosis, as a possible consequence of both p53- and E2F1-mediated activities. Importantly, EBNA3C was previously shown to suppress p53-induced apoptosis. Now, we also show that EBNA3C efficiently blocks E2F1-mediated apoptosis, as well as its anti-proliferative effects in a p53-independent manner, in response to DNA damage. The N- and C-terminal domains of EBNA3C form a stable pRb independent complex with the N-terminal DNA-binding region of E2F1 responsible for inducing apoptosis. Mechanistically, we show that EBNA3C represses E2F1 transcriptional activity via blocking its DNA-binding activity at the responsive promoters of p73 and Apaf-1 apoptosis induced genes, and also facilitates E2F1 degradation in an ubiquitin-proteasome dependent fashion. Moreover, in response to DNA damage, E2F1 knockdown LCLs exhibited a significant reduction in apoptosis with higher cell-viability. In the presence of normal mitogenic stimuli the growth rate of LCLs knockdown for E2F1 was markedly impaired; indicating that E2F1 plays a dual role in EBV positive cells and that active engagement of the EBNA3C-E2F1 complex is crucial for inhibition of DNA damage induced E2F1-mediated apoptosis. This study offers novel insights into our current understanding of EBV biology and enhances the potential for development of effective therapies against EBV associated B-cell lymphomas.