Human cytomegalovirus proteins encoded by UL37 exon 1 protect infected fibroblasts against virus-induced apoptosis and are required for efficient virus replication

Abstract

Human cytomegalovirus (HCMV) strain AD169 mutants carrying transposon insertions or large deletions in UL37 exon 1 (UL37x1) were recovered from modified bacterial artificial chromosomes by reconstitution in human fibroblasts expressing the adenovirus anti-apoptotic protein E1B19K. UL37x1 mutant growth was severely compromised in normal fibroblasts, with minimal release of infectious progeny. Growth in E1B19K-expressing cells was restored, but did not reach wild-type levels. Normal fibroblasts infected by UL37x1 mutants underwent apoptosis spontaneously between 48 and 96 h after infection. Apoptosis was inhibited by treatment of cells with the broad-spectrum caspase inhibitor z-Val-Ala-Asp(OMe)-fluoromethylketone, resulting in substantially increased release of virus. Inhibition of viral DNA replication by phosphonoformate or ganciclovir also inhibited apoptosis, implying that death was triggered by late viral functions or by replication and packaging of the viral genome. Immunofluorescent staining showed that although viral proteins accumulated normally during delayed-early phase and viral DNA replication compartments formed, viral late proteins were detected only rarely, suggesting that spontaneous apoptosis occurs early in late phase. These results demonstrate that anti-apoptotic proteins encoded by HCMV UL37x1 [pUL37x1 (vMIA), gpUL37 and gpUL37 M ] prevent apoptosis that would otherwise be initiated by the replication programme of the virus and are required for efficient and sustainable virus replication.

Mitochondrial and Secretory Human Cytomegalovirus UL37 Proteins Traffic into Mitochondrion-Associated Membranes of Human Cells

“…Consistent with that suggestion is the finding that pUL37x1 mutants (⌬2-23 and ⌬23-34) that are defective in trafficking through the MAM (the present study) and to mitochondria are defective in antiapoptosis (19,25). HCMV UL37x1 viral mutants, defective in antiapoptotic activities, are also defective in their growth (37,42). Taken together, these studies are important because they provide insight into how UL37 proteins traffic from the ER to mitochondria, where they play important roles for the survival of the infected cell.…”

Mechanisms for human cytomegalovirus-induced cytoplasmic p53 sequestration in endothelial cells

“…Studies from this laboratory and others have also demonstrated that HCMV infection can cause severe endothelial dysfunction, e.g. dysregulated apoptosis (Kovacs et al, 1996;Reboredo et al, 2004;Shen et al, 2004) and hampered apoptosis in HeLa cells (Zhu et al, 1995), cancer cells (Michaelis et al, 2004) and fibroblasts (Reboredo et al, 2004). We previously reported that p53 is stabilized in the nucleus at the early stage of HCMV infection and becomes sequestrated in the cytoplasm at the later stage of the infection (Kovacs et al, 1996;Wang et al, 2001;Wang et al, 2000).…”

Structure–function analysis of the interaction between Bax and the cytomegalovirus-encoded protein vMIA

“…In addition, CMV causes the accumulation of DNp73a, a host-encoded inhibitor of p53 and p73 (Terrasson et al, 2005) and stimulates the antiapoptotic phosphatidylinositol 3,4,5 triphosphate kinase pathway (Smith et al, 2004). vMIA protects CMV-infected cells from apoptosis in the late phase of the viral life cycle (Reboredo et al, 2004), and vMIA-deficient/vICA-deficient CMV cannot replicate (because it kills the infected cells prematurely) unless it infects cells that overexpress a Bcl-2-like apoptosis inhibitor (Reboredo et al, 2004). A laboratory strain of vICA-expressing/vMIA-deficient CMV can replicate on human cells (McCormick et al, 2005), but a freshly isolated clinical strain of CMV encoding functionally active vICA loses its replication ability upon knocking out vMIA expression, suggesting that at least in some clinical CMV strains, vMIA is indispensable for viral replication (Jurak and Brune, 2006).…”