Analysis of a charge cluster mutation of herpes simplex virus type 1 UL34 and its extragenic suppressor suggests a novel interaction between pUL34 and pUL31 that is necessary for membrane curvature around capsids.
Interaction between pUL34 and pUL31 is essential for targeting both proteins to the inner nuclear membrane (INM). Sequences mediating the targeting interaction have been mapped by others with both proteins. We have previously reported identification of charge cluster mutants of herpes simplex virus type 1 UL34 that localize properly to the inner nuclear membrane, indicating interaction with UL31, but fail to complement a UL34 deletion. We have characterized one mutation (CL04) that alters a charge cluster near the N terminus of pUL34 and observed the following. (i) The CL04 mutant has a dominant-negative effect on pUL34 function, indicating disruption of some critical interaction. (ii) In infections with CL04 pUL34, capsids accumulate in close association with the INM, but no perinuclear enveloped viruses, cytoplasmic capsids, or virions or cell surface virions were observed, suggesting that CL04 UL34 does not support INM curvature around the capsid. (iii) Passage of UL34-null virus on a stable cell line that expresses CL04 resulted in selection of extragenic suppressor mutants that grew efficiently using the mutant pUL34. (iv) All extragenic suppressors contained an R229-->L mutation in pUL31 that was sufficient to suppress the CL04 phenotype. (v) Immunolocalization and coimmunoprecipitation experiments with truncated forms of pUL34 and pUL31 confirm that N-terminal sequences of pUL34 and a C-terminal domain of pUL31 mediate interaction but not nuclear membrane targeting. pUL34 and pUL31 may make two essential interactions-one for the targeting of the complex to the nuclear envelope and another for nuclear membrane curvature around capsids.
Roller
Richard J
RJ
Department of Microbiology, The University of Iowa, 3-432 Bowen Science Building, Iowa City, IA 52242, USA. richard-roller@uiowa.edu
Bjerke
Susan L
SL
Haugo
Alison C
AC
Hanson
Sara
S
D000818:Animals; D002213:Capsid / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D020125:Mutation, Missense; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D013489:Suppression, Genetic; D014763:Viral Matrix Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly
D009687:Nuclear Proteins; C108721:UL31 protein, Human herpesvirus 1; C403420:UL34 protein, Human herpesvirus 1; D014763:Viral Matrix Proteins; D014764:Viral Proteins