Figure 3

The antiviral acitivity of equine tetherin orthologues. EIAV GagPol expressing construct (A) or pNL-r-HSAS (B) was transfected into 293 T cells along with increasing amounts of huTHN,horTHN, or donTHN (0, 10, 100, 200 ng). Cells were dissolved at 48 h after transfection and then subjected to SDS-PAGE. Viral proteins were analyzed by Western blotting using anti-EIAV serum, or a HIV p24^Gag monoclonal antibody (no. 3537). Intracellular expression of tetherin was detected by Western blotting using anti-HA antibody. (C) The EIAV reporter provirus pONY8.1luc, pGagPol_UK3, and VSV-G were co-transfected with increasing amount of human, horse or donkey tetherins into HEK293T cells. Expression of viral Gag protein and tetherins in cells were assessed by Western blotting. Amounts of EIAV particles in supernatants were determined by measuring viral reverse transcriptase (RT) activity. Levels of infectious HIV-1 were assessed by infecting the HEK293T cells. Values that were obtained from experiments with EIAV reporter constructs alone are set as 100.