title: Human Cytomegalovirus pUL47 Modulates Tegumentation and Capsid Accumulation at the Viral Assembly Complex.
abstract: Human cytomegalovirus (HCMV) tegument protein pUL47 is an interaction partner of pUL48 and highly conserved among herpesviruses. It is closely associated with the capsid and has an important function early in infection. Here, we report a specific role of pUL47 in the tegumentation of capsids in the cytoplasm. A newly generated mutant virus (TB-47stop), in which expression of pUL47 is blocked, exhibited a severe impairment in cell-to-cell spread and release of infectivity from infected cells. Ultrastructural analysis of TB-47stop-infected cells clearly showed cytoplasmic accumulations of nonenveloped capsids that were only partially tegumented, indicating that these capsids failed to complete tegumentation. Nevertheless, these accumulations were positive for HCMV inner tegument proteins pp150 and pUL48, suggesting that their attachment to capsids occurs independently of pUL47. Despite these morphological alterations, fully enveloped virus particles were found in the extracellular space and at the viral assembly complex (vAC) of TB-47stop-infected cells, indicating that pUL47 is not essential for the generation of virions. We confirmed findings that incorporation of pUL48 into virions is impaired in the absence of pUL47. Interestingly, pUL47 exhibited a strong nuclear localization in transfected cells, whereas it was found exclusively at the vAC in the context of virus infection. Colocalization of pUL47 and pUL48 at the vAC is consistent with their interaction. We also found a shift to a more nuclear localization of pUL47 when the expression of pUL48 was reduced. Summarizing our results, we hypothesize that pUL48 directs pUL47 to the vAC to promote tegumentation and secondary envelopment of capsids.
  
  
  OBJECTIVE
  Generation of infectious HCMV particles requires an organized and multistep process involving the action of several viral and cellular proteins as well as protein-protein interactions. A better understanding of these processes is important for understanding the biology of HCMV and may help to identify targets for antiviral intervention. Here, we identified tegument protein pUL47 to function in tegumentation and proper trafficking of capsids during late phases of infection. Although pUL47 is not essential for the generation and release of infectious virions, its absence led to massive accumulations of partially tegumented capsids at the cell periphery. Detection of pUL48 at these accumulations indicated a pUL47-independent attachment of pUL48 to the capsid. On the other hand, localization of pUL47 to the vAC during infection appeared to be dependent on tegument protein pUL48, which suggests an intricate interplay of these proteins for normal generation of infectious virus progeny.
authors.affiliation: Institute of Virology, University Medical Center Ulm, Ulm, Germany.
authors.forename: Ilaria
authors.initials: I
authors.lastname: Cappadona
authors.affiliation: Institute of Virology, University Medical Center Ulm, Ulm, Germany Electron Microscopy Facility, Ulm University, Ulm, Germany.
authors.forename: Clarissa
authors.initials: C
authors.lastname: Villinger
authors.affiliation: Institute of Virology, University Medical Center Ulm, Ulm, Germany.
authors.forename: Gabi
authors.initials: G
authors.lastname: Schutzius
authors.affiliation: Institute of Virology, University Medical Center Ulm, Ulm, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Mertens
authors.affiliation: Institute of Virology, University Medical Center Ulm, Ulm, Germany jens.von-einem@uniklinik-ulm.de.
authors.forename: Jens
authors.initials: J
authors.lastname: von Einem
mesh_terms: D002213:Capsid / Q000378:metabolism*; D003587:Cytomegalovirus / Q000235:genetics / Q000502:physiology*; D055786:Gene Knockout Techniques; D006801:Humans; D011485:Protein Binding; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly; D057074:Virus Release
chemical_list: C446319:UL47 protein, cytomegalovirus; C000591971:UL48 protein, human cytomegalovirus; D014764:Viral Proteins