The nuclear chaperone nucleophosmin escorts an Epstein-Barr Virus nuclear antigen to establish transcriptional cascades for latent infection in human B cells

• Published in: PLoS pathogens Vol. 8; no. 12; p. e1003084
• Main Authors: Liu, Cheng-Der, Chen, Ya-Lin, Min, Yi-Li, Zhao, Bo, Cheng, Chi-Ping, Kang, Myung-Soo, Chiu, Shu-Jun, Kieff, Elliott, Peng, Chih-Wen
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01.12.2012; Public Library of Science (PLoS)
• Subjects: B-Lymphocytes - metabolism; B-Lymphocytes - virology; Biology; Cell Line, Tumor; Epstein-Barr virus; Epstein-Barr Virus Nuclear Antigens - genetics; Epstein-Barr Virus Nuclear Antigens - metabolism; Gene expression; Herpesvirus 4, Human - physiology; Humans; Infections; Lymphocytes; Medical research; Medicine; Molecular Chaperones - genetics; Molecular Chaperones - metabolism; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Protein Structure, Tertiary; Proteins; Proto-Oncogene Proteins c-myc; Transcription, Genetic; Viral infections; Viral Matrix Proteins - biosynthesis; Viral Matrix Proteins - genetics; Viral Proteins - genetics; Viral Proteins - metabolism; Virus Assembly - physiology; Virus Latency - physiology
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1003084

Epstein-Barr Virus (EBV) is an oncogenic γ-herpesvirus that capably establishes both latent and lytic modes of infection in host cells and causes malignant diseases in humans. Nuclear antigen 2 (EBNA2)-mediated transcription of both cellular and viral genes is essential for the establishment and maintenance of the EBV latency program in B lymphocytes. Here, we employed a protein affinity pull-down and LC-MS/MS analysis to identify nucleophosmin (NPM1) as one of the cellular proteins bound to EBNA2. Additionally, the specific domains that are responsible for protein-protein interactions were characterized as EBNA2 residues 300 to 360 and the oligomerization domain (OD) of NPM1. As in c-MYC, dramatic NPM1 expression was induced in EBV positively infected B cells after three days of viral infection, and both EBNA2 and EBNALP were implicated in the transactivation of the NPM1 promoter. Depletion of NPM1 with the lentivirus-expressed short-hairpin RNAs (shRNAs) effectively abrogated EBNA2-dependent transcription and transformation outgrowth of lymphoblastoid cells. Notably, the ATP-bound state of NPM1 was required to induce assembly of a protein complex containing EBNA2, RBP-Jκ, and NPM1 by stabilizing the interaction of EBNA2 with RBP-Jκ. In a NPM1-knockdown cell line, we demonstrated that an EBNA2-mediated transcription defect was fully restored by the ectopic expression of NPM1. Our findings highlight the essential role of NPM1 in chaperoning EBNA2 onto the latency-associated membrane protein 1 (LMP1) promoters, which is coordinated with the subsequent activation of transcriptional cascades through RBP-Jκ during EBV infection. These data advance our understanding of EBV pathology and further imply that NPM1 can be exploited as a therapeutic target for EBV-associated diseases.