E4orf4, a Novel Adenovirus Death Factor That Induces p53-independent Apoptosis by a Pathway That Is Not Inhibited by zVAD-fmk

Abstract

In the absence of E1B, the 289-amino acid product of human adenovirus type 5 13S E1A induces p53-independent apoptosis by a mechanism that requires viral E4 gene products (Marcellus, R.C., J.C. Teodoro, T. Wu, D.E. Brough, G. Ketner, G.C. Shore, and P.E. Branton. 1996. J. Virol. 70:6207–6215) and involves a mechanism that includes activation of caspases (Boulakia, C.A., G. Chen, F.W. Ng, J.G. Teodoro, P.E. Branton, D.W. Nicholson, G.G. Poirier, and G.C. Shore. 1996. Oncogene. 12:529–535). Here, we show that one of the E4 products, E4orf4, is highly toxic upon expression in rodent cells regardless of the p53 status, and that this cytotoxicity is significantly overcome by coexpression with either Bcl-2 or Bcl-Xl. Conditional expression of E4orf4 induces a cell death process that is characterized by apoptotic hallmark features, such as externalization of phosphatidylserine, loss of mitochondrial membrane potential, cytoplasmic vacuolation, condensation of chromatin, and internucleosomal DNA degradation. However, the wide-spectrum inhibitor of caspases, tetrapeptide zVAD-fmk, does not affect any of these apoptogenic manifestations, and does not alter the kinetics of E4orf4-induced cell death. Moreover, E4orf4 expression does not result in activation of the downstream effector caspase common to most apoptosis-inducing events, caspase-3 (CPP32). We conclude, therefore, that in the absence of E1A, E4orf4 is sufficient by itself to trigger a p53-independent apoptosis pathway that may operate independently of the known zVAD-inhibitable caspases, and that may involve an as yet uncharacterized mechanism.

Death-associated protein kinase 2 is a new calcium/calmodulin-dependent protein kinase that signals apoptosis through its catalytic activity

“…In these cells, however, treatment with this peptide did not a ect the ability of DAPK2 to induce apoptosis (our unpublished data), suggesting that DAPK2-induced apotosis is independent of the caspase activation or DAPK2 functions downstream to the caspases. Recently, it has been reported that several types of apoptosis are not inhibited by the caspase inhibitors (Xiang et al, 1996;Miller et al, 1997;Lavoie et al, 1998;Quignon et al, 1998). DAPK2-induced cell death may contribute with these pathways.…”

The permeability transition pore in cell death

“…In fact, the breakdown of this proton gradient seems to be a better cell death predicator than the activation of caspases, since these cystein proteases can be activated without the subsequent demise of the cells provided that m is kept intact [4,5]. Also, in some models of apoptosis caspase or nuclease activation does not occur but m dissipation is nevertheless observed [6,7]. However, as a cautionary note we have to stress that this does not suggest that in each case the PT-pore is responsible for m dissipation as it is possible that factors other than the PT-pore are involved [8].…”

Subcellular and submitochondrial mode of action of Bcl-2-like oncoproteins

“…After dissipation of the DC m and/or release of cytochrome c (which disrupts the transfer of electrons between respiratory chain complexes III and IV), mitochondria lose their anti-oxidant function, produce superoxide anion, and provoke a bioenergetic catastrophe, culminating in cell death irrespective of the activation of caspases and endonucleases (DeMaria et al, 1997;Hirsch et al, 1997;McCarthy et al, 1997;Xiang et al, 1996). In some models of receptor-mediated apoptosis, caspase and nuclease activation does not occur, although cell death is preceded by DC m disruption (Lavoie et al, 1998;Lesage et al, 1997). In general terms, it appears that DC m disruption has a higher predictive value for cell death than caspase or endonuclease activation.…”