Inactivation and disassembly of the anaphase-promoting complex during human cytomegalovirus infection is associated with degradation of the APC5 and APC4 subunits and does not require UL97-mediated phosphorylation of Cdh1.
Infection of quiescent cells by human cytomegalovirus (HCMV) elicits severe cell cycle deregulation, resulting in a G(1)/S arrest, which can be partly attributed to the inactivation of the anaphase-promoting complex (APC). As we previously reported, the premature phosphorylation of its coactivator Cdh1 and/or the dissociation of the core complex can account for the inactivation. We have expanded on these results and further delineated the key components required for disabling the APC during HCMV infection. The viral protein kinase UL97 was hypothesized to phosphorylate Cdh1, and consistent with this, phosphatase assays utilizing a virus with a UL97 deletion mutation (ΔUL97 virus) indicated that Cdh1 is hypophosphorylated at early times in the infection. Mass spectrometry analysis demonstrated that UL97 can phosphorylate Cdh1 in vitro, and the majority of the sites identified correlated with previously characterized cyclin-dependent kinase (Cdk) consensus sites. Analysis of the APC core complex during ΔUL97 virus infection showed APC dissociation occurring at the same time as during infection with wild-type virus, suggesting that the UL97-mediated phosphorylation of Cdh1 is not required for this to occur. Further investigation of the APC subunits showed a proteasome-dependent loss of the APC5 and APC4 subunits that was temporally associated with the disassembly of the APC. Immediate early viral gene expression was not sufficient for the degradation of APC4 and APC5, indicating that a viral early gene product(s), possibly in association with a de novo-synthesized cellular protein(s), is involved.
Tran
Karen
K
Department of Cellular and Molecular Medicine and Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093-0712, USA.
Kamil
Jeremy P
JP
Coen
Donald M
DM
Spector
Deborah H
DH
D000595:Amino Acid Sequence; D064173:Anaphase-Promoting Complex-Cyclosome; D015703:Antigens, CD; D064192:Apc4 Subunit, Anaphase-Promoting Complex-Cyclosome; D064193:Apc5 Subunit, Anaphase-Promoting Complex-Cyclosome; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology; D003586:Cytomegalovirus Infections / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D017931:DNA Primers / Q000235:genetics; D064248:Geminin; D017353:Gene Deletion; D017781:Genes, Immediate-Early; D005814:Genes, Viral; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D061988:Proteasome Inhibitors; D055550:Protein Stability; D021122:Protein Subunits; D043743:Ubiquitin-Protein Ligase Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C577369:ANAPC4 protein, human; C577375:ANAPC5 protein, human; D015703:Antigens, CD; D064192:Apc4 Subunit, Anaphase-Promoting Complex-Cyclosome; D064193:Apc5 Subunit, Anaphase-Promoting Complex-Cyclosome; C513384:CDH1 protein, human; D015820:Cadherins; D018797:Cell Cycle Proteins; D017931:DNA Primers; C577995:GMNN protein, human; D064248:Geminin; D061988:Proteasome Inhibitors; D021122:Protein Subunits; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D017853:Phosphotransferases (Alcohol Group Acceptor); C075365:ganciclovir kinase