Proangiogenic Effect of El A Oncoprotein

摘要

The viral oncoprotein E1A interacts with members of the retinoblastoma and p300/CBP gene families, altering cell phenotype and reactivating proliferation of terminally differentiated cells. Angiogenesis is a growth factor-dependent process sustained by the reactivation of proliferation of quiescent endothelial cells and, at least in part, it mimics the effect of El A in differentiated cells. Therefore the aim of the present study was to examine the effect of El A on endothelial cell's function and to investigate the mechanisms of angiogenesis at molecular level. Bovine aortic endothelial cells (ECs) were transfected either with wild type E1A (wtElA) or with different E1A mutants. We found that transient overexpression of wtElA proteins greatly accelerated ECs' ability to form capillary-like structures on reconstituted basement membrane proteins (Matrigel). In contrast, the E1A mutant dl646N, lacking the CR1 domain of El A, did not modulate the development of capillary-like structures. ECs' invasion, examined in a modified Boyden chamber assay, revealed that wtElA-transfected ECs exhibited an enhanced ability to invade, compared to both dl646N and mock transfectants. Zymograms and RT-PCRs showed that matrix metalloproteinase-9 (MMP-9) protein and m-RNA were induced in wtElA, but not in dl646N- transfected cells. ELISA tests for bFGF revealed an increase of this growth factor in the conditioned media from wtElA-transfected ECs but not in those collected from E1A dl646N-transfected cells. The results demonstrate a role for E1A oncoprotein as inducer of in vitro angiogenesis in adult ECs. This observation, associated with the enhanced MMP-9 activity and bFGF production, is apparently dependent on the integrity of the CR1 domain and suggests a potential role for p300 and Rb family proteins as regulators of nuclear events leading to the angiogenic switch.