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© The Rockefeller University Press,
0021-9525/2000//1037 $5.00
The Journal of Cell Biology, Volume 150, Number 5,
, 2000 1037-1056
Original Article |
Adenovirus E4 Open Reading Frame 4–Induced Apoptosis Involves Dysregulation of Src Family Kinases
The adenoviral early region 4 open reading frame 4 (E4orf4) death factor induces p53-independent apoptosis in many cell types and appears to kill selectively transformed cells. Here we show that expression of E4orf4 in transformed epithelial cells results in early caspase-independent membrane blebbing, associated with changes in the organization of focal adhesions and actin cytoskeleton. Evidence that E4orf4 can associate with and modulate Src family kinase activity, inhibiting Src-dependent phosphorylation of focal adhesion kinase (FAK) and paxillin while increasing phosphorylation of cortactin and some other cellular proteins, is presented. Furthermore, E4orf4 dramatically inhibited the ability of FAK and c-src to cooperate in induction of tyrosine phosphorylation of cellular substrates, suggesting that E4orf4 can interfere with the formation of a signaling complex at focal adhesion sites. Consistent with a functional role for E4orf4–Src interaction, overexpression of activated c-src dramatically potentiated E4orf4-induced membrane blebbing and apoptosis, whereas kinase dead c-src constructs inhibited E4orf4 effects on cell morphology and death. Moreover treatment of E4orf4-expressing cells with PP2, a selective Src kinase inhibitor, led to inhibition of E4orf4-dependent membrane blebbing and later to a marked decrease in E4orf4-induced nuclear condensation. Taken together, these observations indicate that expression of adenovirus 2 E4orf4 can initiate caspase-independent extranuclear manifestations of apoptosis through a modulation of Src family kinases and that these are involved in signaling E4orf4-dependent apoptosis. This study also suggests that Src family kinases are likely to play a role in the cytoplasmic execution of apoptotic programs.
Key Words: actin • blebbing • Src family kinase • focal adhesion kinase • cortactin
© 2000 The Rockefeller University Press
Abbreviations used in this paper: Ad, adenovirus; ECM, extracellular matrix; E4orf4, early region 4 open reading frame 4; ERK, extracellular-regulated kinase; EV, vector alone; FAK, focal adhesion kinase; GFP, green fluorescent protein; GST, glutathione S-transferase; HA, a hemagglutinin; IP, immune complex; PP2A, protein phosphatase 2A.
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