Suppression of amber nonsense mutations of herpes simplex virus type 1 in a tissue culture system

Patel, A.H. , Subak-Sharpe, J.H., Stow, N.D., Marsden, H.S., Maclean, J.B. and Dargan, D.J. (1996) Suppression of amber nonsense mutations of herpes simplex virus type 1 in a tissue culture system. Journal of General Virology, 77(2), pp. 199-209. (doi: 10.1099/0022-1317-77-2-199)

Abstract

We have investigated the ability of monkey kidney cell lines (SupD3 and SupD12) inducibly expressing an amber suppressor tRNAser to suppress amber nonsense mutations in three genes of herpes simplex virus type 1 (HSV-1). HSV-1 mutant TK4, which contains a nonsense mutation in the non-essential viral thymidine kinase (TK) gene, synthesized a full-length TK polypeptide at about 30% of the wild-type (wt) level in induced SupD3 cells but not in the parental non-suppressor (Sup0) cells. Using complementing cells, we constructed HSV-1 mutants carrying nonsense mutations in an essential gene, UL8, encoding a protein essential for viral DNA replication (ambUL8) or in a partially dispensable gene, UL12, encoding alkaline nuclease (ambUL12). The growth of the mutants in Vero or Sup0 cells was either totally (ambUL8) or severely (ambUL12) impaired, whereas in cells expressing suppressor tRNA the mutants produced infectious virus. However, the yields were much lower than obtained with wt HSV-1. In Vero or Sup0 cells the mutants ambUL8 and ambUL12 failed to synthesize full-length UL8 and UL12 protein products, respectively. Western immuno-blotting showed that the virus ambUL12 produced full-length UL12 protein in SupD12 cells which yielded a level of 25.9% of the alkaline nuclease activity of the wt HSV-1 control. Our results show that the levels of suppression of the nonsense mutations in ambUL8 and ambUL12 are insufficient to allow their continuing propagation in the available Sup+ cells. Possible reasons are discussed.

• Item Type: Articles
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Stow, Dr Nigel and Dargan, Dr Derrick and Patel, Professor Arvind
• Authors: Patel, A.H., Subak-Sharpe, J.H., Stow, N.D., Marsden, H.S., Maclean, J.B., and Dargan, D.J.
• College/School: College of Medical Veterinary and Life Sciences > School of Infection & Immunity; College of Medical Veterinary and Life Sciences > School of Infection & Immunity > Centre for Virus Research
• Journal Name: Journal of General Virology
• Publisher: Society for General Microbiology
• ISSN: 0022-1317