Inhibition of Apoptosis by the African Swine Fever Virus Bcl-2 Homologue: Role of the BH1 Domain

Abstract

The function of the African swine fever virus (ASFV) bcl-2 homologue, gene A179L, in the regulation of apoptosis was investigated using as a model system the human myeloid leukemia cell line K562 induced to die by apoptosis with inhibitors of macromolecular synthesis, a process that is prevented by overexpression of human bcl-2. It is shown that transfection of K562 cells with the ASFV A179L gene protects these cells from apoptotic cell death induced by a combination of cycloheximide and actinomycin D or by treatment with cytosine arabinoside. To test the functional role of the highly conserved BH1 domain present in the A179L protein, the Gly residue at position 85 was mutated to Ala, since it has been shown that substitution of the corresponding Gly in human Bcl-2 abrogates its death-repressor activity. It was found that the Gly-to-Ala mutation in the BH1 domain of the viral protein abolished its capacity to protect the K562 cells from apoptosis, indicating that this Gly is essential for A179L action. This finding stresses the functional similarity of the BH1 domains of the viral protein and cellular Bcl-2.

Novel Swine Virulence Determinant in the Left Variable Region of the African Swine Fever Virus Genome

“…Two additional ASFV genes, 5HL and 4CL (also known as A224L) (1,9,44,50), have also been shown to have roles in the maintenance of infected-cell survival. Thus, ASFV encodes multiple genes involved in aspects of maintaining infected-cell survival.…”

“…Virus isolation and titration were done in primary swine macrophage cultures. Virus titers were calculated using the method of Spearman-Karber and expressed as TCID 50 (22). To select rescued virulent viruses, viremic blood (2 ml) was taken from pigs infected with infection-transfection lysates at 7 days postinfection and inoculated into a second group of naïve pigs.…”

Modulation of p53 Cellular Function and Cell Death by African Swine Fever Virus

“…It has been shown that ASFV induces apoptosis after 13 h in the cell, a time at which viral morphogenesis is well under way (7). It was also shown that several ASFV genes are involved in the inhibition of apoptosis using different mechanisms (34,43,44), thus demonstrating that programmed cell death during ASFV infection is a tightly regulated process in which the action of inducers is balanced by the expression of antiapoptotic genes. However, less information is available on the process of ASFV-induced apoptosis in terms of the cellular pathways and specific proteins involved.…”

“…The analysis of the DNA sequence of the virus has revealed the presence of several genes able to modulate host-virus interactions (59). Among these, A179L, which encodes a 19-kDa protein structurally similar to the members of the Bcl-2/Bax family, and A224L, which encodes a 27-kDa protein homologous to IAP family members, have been shown to inhibit apoptosis (1,34,43,44). It has also been reported that ASFV induces apoptosis in the cell in a postbinding step, during or after virus uncoating, through the activation of caspases (7).…”

African Swine Fever Virus IAP-Like Protein Induces the Activation of Nuclear Factor Kappa B

“…As A224L induces, whereas A238L blocks, NF-B activation, this may suggest that the virus requires a low NF-B activity at early times of the viral cycle to avoid immune responses but a high activity at late times, probably to prevent apoptosis. The virus also encodes another apoptosis inhibitor, a Bcl-2-like molecule (25), which is an early protein in the viral cycle that could also act to inhibit programmed cell death at early times after infection.…”