Poster abstracts

Poster number 84 submitted by Feifei Wang

The Effect of SAMHD1 on Murine Leukemia Virus Infection in Mouse NIH3T3 Cells

Feifei Wang (Center for Retrovirus Research,Veterinary Biosciences, College of Veterinary Medicine), Kate Scherer (Center for Retrovirus Research,Veterinary Biosciences, College of Veterinary Medicine), Corine St. Gelais (Center for Retrovirus Research,Veterinary Biosciences, College of Veterinary Medicine), Li Wu (Center for Retrovirus Research,Veterinary Biosciences, College of Veterinary Medicine)

Abstract:
SAMHD1 is a cellular protein that has been proposed to negatively regulate the innate immune response. Mutations in the SAMHD1 gene are associated with Aicardi-Goutières Syndrome (AGS). AGS is an early onset autoimmune disease that causes neurologic symptoms. Myeloid cells such as dendritic cells and macrophages highly express SAMHD1. Human SAMHD1 blocks the replication of human immunodeficiency virus type 1 (HIV-1) in myeloid cells and resting CD4+ T-cells. SAMHD1 mediates HIV-1 restriction by decreasing the intracellular concentration of deoxynucleoside triphosphates (dNTPs) in noncycling cells, which blocks HIV-1 reverse transcription during the infection. Murine leukemia virus (MLV) is a mouse retrovirus that can cause leukemia in certain mouse strains. However, it is unknown whether mouse SAMHD1 can block MLV infection in mouse cells. The goal of this study was to examine whether mouse SAMHD1 protein can block MLV infection in the mouse fibroblast cell line NIH3T3. This was accomplished by first transfecting a human cell line to generate retroviral vectors expressing human and mouse SAMHD1 and an antibiotic selection marker. Then NIH3T3 cells were transduced with retroviral vectors that express SAMHD1. A stable cell line expressing human SAMHD1 was generated by puromycin selection. Human SAMHD1 expression was confirmed by Western blot analysis. Then the NIH3T3 cells were infected with a single-cycle luciferase reporter MLV. The infection was quantified by measuring luciferase activity in the cell lysate post-infection. Our preliminary data suggest that human SAMHD1 can block MLV infection in NIH3T3 cells by decreasing the intracellular dNTP pool. We are currently generating stable NIH3T3 cells that express two isoforms of mouse SAMHD1 and optimizing HIV-1 and MLV infections in these cell lines. These studies will enrich our knowledge about the mechanisms underlying SAMHD1-mediated retroviral restriction in mouse cells.

Keywords: SAMHD1, MLV