Molluscum contagiosum virus (MCV), a member of the human poxvirus family, encodes the MC159 protein that inhibits Fas-, tumor necrosis factor (TNF)-, and TNF-related apoptosis-inducing ligant (TRAIL)-induced apoptosis. We used site-directed mutagenesis to change charged or hydrophobic amino acid residues to alanines to identify regions of MC159 that are critical for protection from apoptosis and for protein-protein interactions. Surprisingly, while MC159 is thought to block apoptosis by binding to Fas-associated death domain (FADD) or caspase-8, several mutants that lost apoptosis blocking activity still bound to both FADD and caspase-8. Mutations in the predicted hydrophobic patch 1 and ␣2 regions of both death effector domains (DEDs) within MC159 resulted in loss of the ability to bind to FADD or caspase-8 and to block apoptosis. Amino acid substitutions in the RXDL motif located in the ␣6 region of either DED resulted in loss of protection from apoptosis induced by Fas, TNF, and TRAIL and abolished the ability of MC159 to block death effector filament formation. Thus, charged or hydrophobic amino acids in three regions of the MC159 DEDs (hydrophobic patch 1, ␣2, and ␣6) are critical for the protein's ability to interact with cellular proteins and to block apoptosis.Death receptors constitute a subgroup of the tumor necrosis factor (TNF) receptor superfamily, which are defined by the presence of a signaling domain with six ␣-helices in the cytoplasmic region that is termed the death domain (13). These receptors function to maintain homeostasis in the immune system by eliminating autoreactive cells, antigen-reactive T cells following an immune response, and virus-infected or malignant cells. In humans, six death receptors have been identified: Fas (also called CD95 and APO-1), TNF receptor 1 (TNFR1; also called TNFRSF1A and CD120a p55-R), DR3 (also called APO3, Wsl-1, TRAMP. and LARD), DR4 (also called TRAIL-R1 and Apo-2), DR5 (also called TRAIL-R2, KILLER, and TRICK2), and DR6 (17,19).Binding of a death receptor to its ligand initiates a change in the receptor complex, resulting in signaling through a series of protein-protein interactions that culminate in apoptosis (4, 22). Fas binding to its ligand (FasL) leads to the recruitment of the adapter molecule FADD (Fas-associated death domain) through interactions of the death domains in Fas and FADD. FADD contains another region known as the death effector domain (DED). The FADD DED also possesses six ␣-helices in a folded region similar to the death domains, but it forms distinct contacts only with other DED-containing proteins, whereas death domain-containing proteins interact chiefly with other death domain-containing adapter proteins.Once FADD is recruited to Fas, its DED binds to the DEDs in the prodomain of caspase-8 or caspase-10 (1, 28). The complex containing Fas, FADD, and caspase-8 or caspase-10 is termed the death-inducing signaling complex (DISC). Recruitment of caspase-8 or caspase-10 into the DISC results in autocatalytic cleavage of the caspase i...

The murine gammaherpesvirus-68 (MHV-68) M11 gene encodes a protein predicted to have limited homology to the bcl-2 family of proteins. Unlike most of the other viral bcl-2 homologues, which have both BH1 and BH2 domains conserved with respect to bcl-2, the M11 protein has a BH1 domain, but apparently lacks a BH2 domain. Transfection of HeLa cells with an epitope-tagged MHV-68 M11 construct showed that the protein is predominantly located in the cytoplasm of cells. In HeLa cells, M11 inhibited apoptosis induced by anti-Fas antibody and by TNF-α. Thus, despite its limited conservation with respect to other bcl-2 family members, the MHV-68 M11 protein is a potent inhibitor of apoptosis.Apoptosis is a host defence mechanism that the cell uses to limit production of infectious virus. Many viruses encode proteins that inhibit apoptosis (reviewed in O'Brien, 1998 ;Tschopp et al., 1998). Some viral proteins inhibit the proenzyme or activated form of caspases, while other viral proteins inhibit p53-mediated apoptosis or encode soluble TNF-α receptors. Several viruses including adenovirus, African swine fever virus and gammaherpesviruses encode homologues of the cellular bcl-2 protein.The bcl-2 family of proteins includes both anti-apoptotic (e.g. bcl-2, bcl-X L ) and pro-apoptotic (e.g. bak, bax) proteins. Members of this family contain up to four bcl-2 homology domains (BH1 to BH4). The BH1 and BH2 domains are important for bcl-2 and bcl-X L to form heterodimers with bax (reviewed in Kroemer, 1997). The BH3 domain of bak and bax is critical for these proteins to form heterodimers with bcl-2 and bcl-X L . The BH4 domain of bcl-2 is important for interactions with other cellular proteins.Most herpesvirus bcl-2 homologues show conservation of

Pneumonia virus of mice (PVM; family Paramyxoviridae) is a natural pathogen of rodents that reproduces important clinical features of severe respiratory syncytial virus infection in humans. As anticipated, PVM infection induces transcription of IFN antiviral response genes preferentially in wild-type over IFN-αβR gene-deleted (IFN-αβR−/−) mice. However, we demonstrate that PVM infection results in enhanced expression of eotaxin-2 (CCL24), thymus and activation-regulated chemokine (CCL17), and the proinflammatory RNase mouse eosinophil-associated RNase (mEar) 11, and decreased expression of monocyte chemotactic protein-5, IFN-γ-inducible protein-10, and TLR-3 in lung tissue of IFN-αβR−/− mice when compared with wild type. No differential expression of chemokines MIP-1α or MIP-2 or Th2 cytokines IL-4 or IL-5 was observed. Differential expression of proinflammatory mediators was associated with distinct patterns of lung pathology. The widespread granulocytic infiltration and intra-alveolar edema observed in PVM-infected, wild-type mice are replaced with patchy, dense inflammatory foci localized to the periphery of the larger blood vessels. Bronchoalveolar lavage fluid from IFN-αβR−/− mice yielded 7- to 8-fold fewer leukocytes overall, with increased percentages of eosinophils, monocytes, and CD4+ T cells, and decreased percentage of CD8+ T cells. Differential pathology is associated with prolonged survival of the IFN-αβR−/− mice (50% survival at 10.8 ± 0.6 days vs the wild type at 9.0 ± 0.3 days; p < 0.02) despite increased virus titers. Overall, our findings serve to identify novel transcripts that are differentially expressed in the presence or absence of IFN-αβR-mediated signaling, further elucidating interactions between the IFN and antiviral inflammatory responses in vivo.

No abstract