Induction of Apoptosis in Rabbit Kidney Cell under High-Level Expression of Bovine Herpesvirus 1 Us ORF8 Product

Abstract

Objective: The open reading frame 8 (ORF8) within the short unique (U_s) region of the bovine herpesvirus 1 (BHV-1) genome was predicted to encode a protein with homology to U_s9 protein of other alphaherpesviruses. The aim of this study is to identify the protein encoded by the U_s ORF8 and to examine the effect of its expression in mammalian cells. Methods: A polyclonal antiserum was raised against U_s ORF8 protein expressed in Escherichia coli. A recombinant baculoviurs designated as Ac/CA8 was created by integrating U_s ORF8 under the control of the mammalian CAG promoter into the baculovirus genome. U_s ORF8 protein was expressed in rabbit kidney (RK13) cells transduced by Ac/CA8. Results: The antiserum reacted specifically with 27-and 32-kD polypeptides from the BHV-1-infected and Ac/CA8-transduced RK13 cells. High-level expression of U_s ORF8 protein in RK13 cells transduced by Ac/CA8 led to a distinct cytopathic effect and a reduction in cell viability; the onset of apoptotic cell death was induced as judged by DNA laddering and chromatin condensation analyses. Conclusions: These data represent the identification of BHV-1 U_sORF8 protein, which directly induces apoptosis in RK13 cells.