HERV-K Gag polyprotein cleavage requires a functional retroviral protease. Western blot analysis of HERV-K Gag, expressed by transient vaccinia virus-driven expression. HERV-K Gag was detected with the HERV-K CA-specific monoclonal antibody mix HERMA  and an HRP-coupled anti-mouse antibody followed by ECL detection (Amersham, Freiburg). A: Lys: cell lysates; sup: concentrated cell culture supernatants. Lanes 1-3: HERV-K Gag expression with a functional protease; Lanes 4-6: HERV-K Gag expression without protease; +: plasmid transfection with additional MVA infection. Positions of the Gag polyprotein and CA are indicated. B: Lys: cell lysates; sup: concentrated cell culture supernatants. Lane 1: lysate of untreated 293T cells; lanes 2-3: plasmid driven HERV-K Gag expression, samples prepared 48 hrs after transfection; Lanes 4-5: MVA-HERV-Kcon driven HERV-K Gag expression, samples prepared 24 hrs after infection. The loading volume was one-third of the volume used in lanes 1, 2 and 3.