Epstein-Barr virus LMP1 blocks p16INK4a-RB pathway by promoting nuclear export of E2F4/5

Authors:
Ohtani N, Brennan P, Gaubatz S, Sanij E, Hertzog P, Wolvetang E, Ghysdael J, Rowe M and Hara E
In:
J Cell Biol (2003) 162(2): 173-183
Research Area:
Dermatology/Tissue Engineering
Cells used in publication:
Fibroblast, dermal, human
Species: human
Tissue Origin: dermal
Experiment
The p16INK4a-RB pathway plays a critical role in preventing inappropriate cell proliferation and is often targeted by viral oncogenes during immortalization. Ets2 is an important transcription factor of p16INK4a. E2F4/5 are essential downstream mediators of the p16INK4a growth arrest pathway. The authors studied how the latent membrane protein (LMP1) of Eppstein-Barr virus represses the p16INK4a gene expression. Primary fibroblasts were co-transfected with an Ets2 expression vector and an inducible GFP-tagged LMP1 expression vector. Induction of LMP1 lead to a translocation of Ets2 to the cytoplasm. To evaluate the impact of the LMP1-induced intracellular redistribution of E2F4 on cell growth fibroblasts were nucleofected with expression vectors encoding WT E2F4 or E2F4 fused to a NLS (nuclear localization sequence). LMP1 expression alone significantly increased cell number whereas co-expression of NLS-E2F4 with LMP1 blocked this effect indicating the relevance of LMP1-induced intracellular redistribution of E2F4 to LMP1-dependent cell proliferation.
Abstract
The p16INK4a-RB pathway plays a critical role in preventing inappropriate cell proliferation and is often targeted by viral oncoproteins during immortalization. Latent membrane protein 1 (LMP1) of Epstein-Barr virus (EBV) is often present in EBV-associated proliferative diseases and is critical for the immortalizing and transforming activity of EBV. Unlike other DNA tumor virus oncoproteins, which possess immortalizing activity, LMP1 does not bind to retinoblastoma tumor suppressor protein, but instead blocks the expression of p16INK4a tumor suppressor gene. However, it has been unclear how LMP1 represses the p16INK4a gene expression. Here, we report that LMP1 promotes the CRM1-dependent nuclear export of Ets2, which is an important transcription factor for p16INK4a gene expression, thereby reducing the level of p16INK4a expression. We further demonstrate that LMP1 also blocks the function of E2F4 and E2F5 (E2F4/5) transcription factors through promoting their nuclear export in a CRM1-dependent manner. As E2F4/5 are essential downstream mediators for a p16INK4a-induced cell cycle arrest, these results indicate that the action of LMP1 on nuclear export has two effects on the p16INK4a-RB pathway: (1) repression of p16INK4a expression and (2) blocking the downstream mediator of the p16INK4a-RB pathway. These results reveal a novel activity of LMP1 and increase an understanding of how viral oncoproteins perturb the p16INK4a-RB pathway.