title: Identification of conserved amino acids in pUL34 which are critical for function of the pseudorabies virus nuclear egress complex.
abstract: Nuclear egress of herpesvirus capsids is mediated by a conserved heterodimeric complex of two viral proteins, designated pUL34 and pUL31 in herpes simplex virus and pseudorabies virus (PrV). pUL34, a tail-anchored membrane protein, is targeted to the nuclear envelope and recruits pUL31 to the inner nuclear membrane (INM) to provide the docking and envelopment machinery for the nascent capsid. While the less conserved C-terminal part of pUL34 is required for correct positioning of the nuclear egress complex (NEC) at the INM, the conserved N-terminal part functions as a docking site for pUL31. Since no crystal structure of NEC is available yet, structure-function studies depend on mutational analyses, with several approaches already being performed for different herpesvirus NECs. Here, we extended our studies on PrV pUL34 and identified two asparagine residues (N75, N103) and a dileucine motif (LL166/167), adjacent to an endoplasmic reticulum retention signal, which are absolutely required for NEC function. While the pUL34-N75A substitution mutant is unable to interact with pUL31, the pUL34-N103A mutant is nonfunctional, despite continuing complex formation. Surprisingly, mutant pUL34-G77A, which does not efficiently recruit pUL31 to the nuclear rim after cotransfection, nonetheless complements a UL34 deletion mutant, indicating that the NEC may be stabilized by additional viral factors during infection.
  
  
  OBJECTIVE
  In the absence of a crystal structure of the nuclear egress complex (NEC) required for herpesvirus maturation, site-directed mutagenesis studies provide important information on critical amino acid residues. Here, we identify conserved amino acid residues in the membrane-bound component of the NEC which are relevant for its function.
authors.affiliation: Institute of Molecular Biology, Friedrich-Loeffler-Institut, Greifswald-Insel Riems, Germany.
authors.forename: Lars
authors.initials: L
authors.lastname: Paßvogel
authors.forename: Una
authors.initials: U
authors.lastname: Janke
authors.forename: Barbara G
authors.initials: BG
authors.lastname: Klupp
authors.forename: Harald
authors.initials: H
authors.lastname: Granzow
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Mettenleiter
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D017124:Conserved Sequence / Q000235:genetics*; D005455:Fluorescent Antibody Technique; D005816:Genetic Complementation Test; D011558:Herpesvirus 1, Suid / Q000235:genetics*; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011817:Rabbits; D017422:Sequence Analysis, DNA; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D057074:Virus Release / Q000235:genetics*
chemical_list: C403420:UL34 protein, Human herpesvirus 1; D014764:Viral Proteins