title: Conserved Tryptophan Motifs in the Large Tegument Protein pUL36 Are Required for Efficient Secondary Envelopment of Herpes Simplex Virus Capsids.
abstract: Herpes simplex virus (HSV) replicates in the skin and mucous membranes, and initiates lytic or latent infections in sensory neurons. Assembly of progeny virions depends on the essential large tegument protein pUL36 of 3,164 amino acid residues that links the capsids to the tegument proteins pUL37 and VP16. Of the 32 tryptophans of HSV-1-pUL36, the tryptophan-acidic motifs (1766)WD(1767) and (1862)WE(1863) are conserved in all HSV-1 and HSV-2 isolates. Here, we characterized the role of these motifs in the HSV life cycle since the rare tryptophans often have unique roles in protein function due to their large hydrophobic surface. The infectivity of the mutants HSV-1(17(+))Lox-pUL36-WD/AA-WE/AA and HSV-1(17(+))Lox-CheVP26-pUL36-WD/AA-WE/AA, in which the capsid has been tagged with the fluorescent protein Cherry, was significantly reduced. Quantitative electron microscopy shows that there were a larger number of cytosolic capsids and fewer enveloped virions compared to their respective parental strains, indicating a severe impairment in secondary capsid envelopment. The capsids of the mutant viruses accumulated in the perinuclear region around the microtubule-organizing center and were not dispersed to the cell periphery but still acquired the inner tegument proteins pUL36 and pUL37. Furthermore, cytoplasmic capsids colocalized with tegument protein VP16 and, to some extent, with tegument protein VP22 but not with the envelope glycoprotein gD. These results indicate that the unique conserved tryptophan-acidic motifs in the central region of pUL36 are required for efficient targeting of progeny capsids to the membranes of secondary capsid envelopment and for efficient virion assembly.
  
  
  
  Herpesvirus infections give rise to severe animal and human diseases, especially in young, immunocompromised, and elderly individuals. The structural hallmark of herpesvirus virions is the tegument, which contains evolutionarily conserved proteins that are essential for several stages of the herpesvirus life cycle. Here we characterized two conserved tryptophan-acidic motifs in the central region of the large tegument protein pUL36 of herpes simplex virus. When we mutated these motifs, secondary envelopment of cytosolic capsids and the production of infectious particles were severely impaired. Our data suggest that pUL36 and its homologs in other herpesviruses, and in particular such tryptophan-acidic motifs, could provide attractive targets for the development of novel drugs to prevent herpesvirus assembly and spread.
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.|German Center for Infection Research (DZIF), Hannover, Germany.
authors.forename: Lyudmila
authors.initials: L
authors.lastname: Ivanova
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Buch
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Katinka
authors.initials: K
authors.lastname: Döhner
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Pohlmann
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Binz
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Ute
authors.initials: U
authors.lastname: Prank
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Malte
authors.initials: M
authors.lastname: Sandbaumhüter
authors.affiliation: Institute of Cell Biology, Hannover Medical School, Hannover, Germany Bauerfeind.Rudi@mh-hannover.de Sodeik.Beate@mh-hannover.de.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Bauerfeind
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany Bauerfeind.Rudi@mh-hannover.de Sodeik.Beate@mh-hannover.de.|German Center for Infection Research (DZIF), Hannover, Germany.
authors.forename: Beate
authors.initials: B
authors.lastname: Sodeik
mesh_terms: D020816:Amino Acid Motifs; D002213:Capsid / Q000378:metabolism* / Q000648:ultrastructure; D036022:Capsid Proteins / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D003593:Cytoplasm / Q000821:virology; D017405:Herpes Simplex Virus Protein Vmw65 / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008018:Life Cycle Stages; D008854:Microscopy, Electron; D009154:Mutation; D011485:Protein Binding; D000072417:Protein Domains; D014364:Tryptophan / Q000737:chemistry* / Q000378:metabolism; D015678:Viral Structural Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly
chemical_list: D036022:Capsid Proteins; D017405:Herpes Simplex Virus Protein Vmw65; D015678:Viral Structural Proteins; D014364:Tryptophan