SP100B as a Repressor of Viral Gene Expression

Abstract

Abstract: : Purpose: After infection viral DNAs localize adjacent to nuclear PML bodies. It has been proposed that a component of PML bodies is a repressor of viral gene expression, since PML bodies increase in size and number following interferon (IFN) treatment and viruses destroy these bodies. SP100B, a splice variant of the PML body protein SP100A, is detected in interferon treated cells and is a potent repressor of expression of transfected genes. The studies reported here were designed to test whether SP100B expression could repress viral gene expression. Methods: Cell lines were created that inducibly express SP100A or SP100B as GFP or flag–tagged proteins under the regulation of the tet–repressor. To determine whether expression of SP100B repressed viral gene expression, the cells were induced to expressed SP100 proteins and then infected with recombinant herpes simplex viruses (HSV) that express reporter genes under the control of immediate early, early, or late viral promoters. Viral gene expression levels in SP100B expressing cells were compared with those from infected cells induced to express SP100A or not induced. The combined effects of SP100B expression and IFN treatment were assessed on HSV gene expression. The effect of HSV infection on SP100 levels was examined by western blot. Colocalization of SP100A and SP100B with viral proteins was examined by immunofluorescence. Transcription of viral genes was examined by RT–PCR. Results: Expression of SP100B, but not SP100A inhibited viral gene expression. Immediate early gene expression was reduced to a limited extent (2–fold), but early and late gene expression showed greater inhibition (6–fold). IFN treatment resulted in a 3–fold reduction in expression, but combination of SP100B expression and IFN treatment led to a greater than 50–fold reduction in expression from the viral thymidine kinase promoter. HSV infection resulted in the loss of SP100A, but a relocalization of SP100B, colocalizing with ICP4. Conclusions:SP100B expression resulted in a repression of viral gene expression, especially affecting early and late viral genes. IFN treatment combined with SP100B expression showed dramatic repressive activity. SP100B may be an important component of the innate antiviral immune response of cells, especially following IFN treatment.

Keywords: herpes simplex virus • immunomodulation/immunoregulation • antiviral drugs