To investigate whether the precursor Gag-Pol polyprotein was indeed incorporated into pseudoviral particles of mutants Y271A and I274A, products of the Gag-Pol polyprotein, integrase (IN), protease (PR) and p24, in wild type and mutant pseudoviral particles were examined by Western blotting. The results showed that, except for RT (Fig. 2B) , all products of the Gag-Pol polyprotein, IN p32, PR p11 and CA p24, were detected in the mutants Y271A and I274A at a level similar to that of the wild type pseudoviral particles (Fig. 4A) . Thus, Pr160 gag-pol was indeed incorporated into the virions and processed properly. To study if the RTs in the viral particles of Y271A and I274A mutants were degraded by proteolysis that made them undetectable, pseudoviruses of wild type and mutants were generated in the presence or absence of indinavir, a highly specific inhibitor of HIV-1 protease. Indinavir treatment was effective, because the treatment resulted in a dose-dependent inhibition of Pr55 Gag processing into p24 for wild type and mutant viruses (Fig. 4B ). As shown in Fig. 4C , in the absence of indinavir, both RT p66 and p51 were readily detected in the wild type pseudovirus, but not in the mutant viruses. In the presence of indinavir, however, both RT p66 and p55 were markedly reduced in the wild type virus but became detectable in mutant Y271A viral particles, while RT p66 was also detectable in mutant I274A virus. We also detected the RT subunits in the viral supernatants collected at earlier time points (12, 24 and 36 hours post-transfection), but the mutant RTs were still undetectable (data not shown). These results suggested that the Y271A and I274A