Stable expression of human immunodeficiency virus type 1 Nef confers resistance against Fas-mediated apoptosis

To investigate the effect of Nef on Fas-mediated apoptosis, we compared T c ells, both population and subclones stably expressing Nef from HIV-1(NL432) , With Nef(-) control cells. Fas-mediated apoptosis was significantly delay ed in Nef(+) cells as determined by annexin V staining and the percentage o f apoptotic cells was lower in all Nef-expressing cells than in the control cells by a maximum of 10-fold, Next we measured cell surface levels of Fas to test whether the delayed apoptosis in Nef(+) cells was due to reduced c ell surface expression of Fas, We found that there was no significant diffe rence in the surface level of Fas between the Nef(+) and Nef(-) cells. To f urther define the steps affected by Nef in the Fas signaling pathway, the a ctivation of caspase-3 and caspase-8 was investigated. A reasonable correla tion was found between the magnitude of apoptosis measured by annexin V sta ining and the enzymatic activity of caspase-3, The overall level of caspase -8 activity in Nef(+) cells was also lower than in Nef(-) cells, although t he extent of inhibition was not as significant as seen for caspase-3. Overa ll, our results indicate that long-term stable expression of Nef, which mim icks persistent or latent infection in vivo, confers resistance against ant i-Fas Ab-induced apoptosis through inhibition of caspase-3 and caspase-8 ac tivation.