THE CYTOPLASMIC DOMAIN OF CD4 IS SUFFICIENT FOR ITS DOWN-REGULATION FROM THE CELL-SURFACE BY HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NEF

Human immunodeficiency virus type 1 Nef down-regulates surface express ion of murine and human CD4 but not human CD8. We recently reported th at the cytoplasmic domain of CD4 is required for its down-regulation b y Nef. Using a chimeric molecule composed of the extracellular and tra nsmembrane domains of human CD8 fused to the cytoplasmic domain of hum an CD4, we show here that the cytoplasmic domain of CD4 is sufficient for dean-regulation by Nef. Since the cytoplasmic domain of CD4 is als o the site of its association with p56(lck), we used a series of CD4 m utants to determine whether the regions of the cytoplasmic domain of C D4 required for down-regulation by Nef are the same as those required for p56(lck) binding. Our results indicate that the portion of the cyt oplasmic domain required for the down-regulation of CD4 by Nef overlap s with the binding site of p56(lck), but the cysteine residues which a re essential for the association of CD4 with p56(lck) are not required . This observation raised the possibility that Nef competes with p56(l ck) for binding to CD4. However, under conditions which are considerab ly milder than those permissive for coimmunoprecipitation of CD4 and p 56(lck), we found no evidence for an association between Nef and CD4. While a decrease in total CD4 was observed in lysates of cells express ing Nef, the levels of p56(lck) were not significantly affected. Pulse -chase experiments further revealed a decrease in the half-life of CD4 in Nef-expressing cells. These results show that the decrease in surf ace CD4 expression induced by Nef is mediated at least in part by a de crease in the half-life of CD4 protein. These results also indicate th at a large portion of p56(lck) is free of CD4 in T cells expressing Ne f, which could have a significant effect on T-cell function.