title: A single mutation responsible for temperature-sensitive entry and assembly defects in the VP1-2 protein of herpes simplex virus.
abstract: Evidence for an essential role of the herpes simplex virus type 1 (HSV-1) tegument protein VP1-2 originated from the analysis of the temperature-sensitive (ts) mutant tsB7. At the nonpermissive temperature (NPT), tsB7 capsids accumulate at the nuclear pore, with defective genome release and substantially reduced virus gene expression. We compared the UL36 gene of tsB7 with that of the parental strain HFEM or strain 17 and identified four amino acid substitutions, 1061D → G, 1453Y → H, 2273Y → H, and 2558T → I. We transferred the UL36 gene from tsB7, HFEM, or strain 17 into a KOS background. While KOS recombinants containing the HFEM or strain 17 UL36 gene exhibited no ts defect, recombinants containing the tsB7 UL36 VP1-2 exhibited a 5-log deficiency at the NPT. Incubation at the NPT resulted in little or no virus gene expression, though limited expression could be detected in a highly delayed fashion. Using shift-down regimes, gene expression recovered and recapitulated the time course normally observed, indicating that the initial block was in a reversible pathway. Using temperature shift-up regimes, a second defect later in the replication cycle was also observed in the KOS.ts viruses. We constructed a further series of recombinants which contained subsets of the four substitutions. A virus containing the wild-type (wt) residue at position 1453 and with the other three residues being from tsB7 VP1-2 exhibited wt plaquing efficiency. Conversely, a virus containing the three wt residues but the single Y → H change at position 1453 from tsB7 exhibited a 4- to 5-log drop in plaquing efficiency and was defective at both early and late stages of infection.
authors.affiliation: Imperial College, London W2 1PG, UK.
authors.forename: F
authors.initials: F
authors.lastname: Abaitua
authors.forename: T
authors.initials: T
authors.lastname: Daikoku
authors.forename: C M
authors.initials: CM
authors.lastname: Crump
authors.forename: M
authors.initials: M
authors.lastname: Bolstad
authors.forename: P
authors.initials: P
authors.lastname: O'Hare
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D056945:Hep G2 Cells; D006561:Herpes Simplex / Q000821:virology*; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D017354:Point Mutation; D016415:Sequence Alignment; D013696:Temperature; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D019065:Virus Assembly; D053586:Virus Internalization; D014779:Virus Replication
chemical_list: C078035:UL36 protein, Human herpesvirus 1; D014764:Viral Proteins