title: Preparation of the Human Cytomegalovirus Nuclear Egress Complex and Associated Proteins.
abstract: Herpesviruses, like most DNA viruses, replicate their genomes in the host cell nucleus. Their DNA is then packaged and assembled into viral nucleocapsids, which, in most cases, are too large to pass through the nuclear pore complex. Instead, herpesviruses use a complex multistep pathway, termed nuclear egress, to exit the nucleus. Key players in this process include two conserved viral proteins that form the nuclear egress complex (NEC). In human cytomegalovirus, these NEC proteins are UL50, embedded in the inner nuclear membrane, and its nucleoplasmic partner UL53. Both are essential for viral nuclear egress. However, other viral components as well as host nuclear envelope proteins may also participate in nuclear egress. Identifying these viral and cellular factors may provide important insight into the herpesvirus lifecycle and its relationship to the underlying, yet still-mysterious, host nuclear egress pathway. We developed an immunoprecipitation-based protocol, described herein, to identify protein-protein interactions involving the NEC from the nuclear fraction of infected cells that express an epitope-tagged version of NEC subunit UL53.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA. Electronic address: mayuri_sharma@hms.harvard.edu.
authors.forename: Mayuri
authors.initials: M
authors.lastname: Sharma
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA; Department of Microbiology and Immunology, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Jeremy P
authors.initials: JP
authors.lastname: Kamil
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA. Electronic address: don_coen@hms.harvard.edu.
authors.forename: Donald M
authors.initials: DM
authors.lastname: Coen
mesh_terms: D002458:Cell Fractionation; D003587:Cytomegalovirus; D006801:Humans; D047468:Immunoprecipitation; D046912:Multiprotein Complexes / Q000302:isolation & purification*; D061251:Primary Cell Culture; D014764:Viral Proteins / Q000302:isolation & purification*
chemical_list: D046912:Multiprotein Complexes; D014764:Viral Proteins
keywords: Herpesvirus; Human cytomegalovirus; Nuclear egress complex; Nuclear envelope; Nuclear lamina; Protein kinase; Proteomics