title: Proteomic analysis of the multimeric nuclear egress complex of human cytomegalovirus.
abstract: Herpesviral capsids are assembled in the host cell nucleus before being translocated into the cytoplasm for further maturation. The crossing of the nuclear envelope represents a major event that requires the formation of the nuclear egress complex (NEC). Previous studies demonstrated that human cytomegalovirus (HCMV) proteins pUL50 and pUL53, as well as their homologs in all members of Herpesviridae, interact with each other at the nuclear envelope and form the heterodimeric core of the NEC. In order to characterize further the viral and cellular protein content of the multimeric NEC, the native complex was isolated from HCMV-infected human primary fibroblasts at various time points and analyzed using quantitative proteomics. Previously postulated components of the HCMV-specific NEC, as well as novel potential NEC-associated proteins such as emerin, were identified. In this regard, interaction and colocalization between emerin and pUL50 were confirmed by coimmunoprecipitation and confocal microscopy analyses, respectively. A functional validation of viral and cellular NEC constituents was achieved through siRNA-mediated knockdown experiments. The important role of emerin in NEC functionality was demonstrated by a reduction of viral replication when emerin expression was down-regulated. Moreover, under such conditions, reduced production of viral proteins and deregulation of viral late cytoplasmic maturation were observed. Combined, these data prove the functional importance of emerin as an NEC component, associated with pUL50, pUL53, pUL97, p32/gC1qR, and further regulatory proteins. Summarized, our findings provide the first proteomics-based characterization and functional validation of the HCMV-specific multimeric NEC.
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany;
authors: Jens
authors: J
authors: Milbradt
authors: Université Grenoble Alpes, iRTSV-BGE, F-38000 Grenoble, France; ¶CEA, iRTSV-BGE, F-38000 Grenoble, France; INSERM, BGE, F-38000 Grenoble, France;
authors: Alexandra
authors: A
authors: Kraut
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany;
authors: Corina
authors: C
authors: Hutterer
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany;
authors: Eric
authors: E
authors: Sonntag
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany;
authors: Cathrin
authors: C
authors: Schmeiser
authors: Université Grenoble Alpes, iRTSV-BGE, F-38000 Grenoble, France; ¶CEA, iRTSV-BGE, F-38000 Grenoble, France; INSERM, BGE, F-38000 Grenoble, France;
authors: Myriam
authors: M
authors: Ferro
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany;
authors: Sabrina
authors: S
authors: Wagner
authors: **Department of Histology and Embryology, Faculty of Medicine, University of Rijeka, 51000 Rijeka, Croatia;
authors: Tihana
authors: T
authors: Lenac
authors: ‡‡Institute for Virology, University of Leipzig, 04103 Leipzig, Germany;
authors: Claudia
authors: C
authors: Claus
authors: §§Institute of Biotechnology, University of Technology Berlin, 13353 Berlin, Germany;
authors: Sandra
authors: S
authors: Pinkert
authors: ¶¶Virology Division, SEALS Microbiology, Prince of Wales Hospital, University of New South Wales, 2052 Sydney, Australia;
authors: Stuart T
authors: ST
authors: Hamilton
authors: ¶¶Virology Division, SEALS Microbiology, Prince of Wales Hospital, University of New South Wales, 2052 Sydney, Australia;
authors: William D
authors: WD
authors: Rawlinson
authors: ‖‖Division of Bioinformatics, Institute of Biochemistry, University of Erlangen-Nuremberg, 91054 Erlangen, Germany.
authors: Heinrich
authors: H
authors: Sticht
authors: Université Grenoble Alpes, iRTSV-BGE, F-38000 Grenoble, France; ¶CEA, iRTSV-BGE, F-38000 Grenoble, France; INSERM, BGE, F-38000 Grenoble, France; manfred.marschall@viro.med.uni-erlangen.de yohann.coute@cea.fr.
authors: Yohann
authors: Y
authors: Couté
authors: From the Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, 91054 Erlangen, Germany; manfred.marschall@viro.med.uni-erlangen.de yohann.coute@cea.fr.
authors: Manfred
authors: M
authors: Marschall
mesh_terms: D000818:Animals; D003587:Cytomegalovirus / Q000502:physiology*; D005347:Fibroblasts / Q000821:virology; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D009687:Nuclear Proteins; D014764:Viral Proteins; C092710:emerin; C523758:pUL50 protein, cytomegalovirus; C523759:pUL53 protein, cytomegalovirus