Expression of the Nef protein encoded by human and simian immunodeficiency viruses results in the specific down-regulation of CD4 from the cell surface in both lymphoid and non-lymphoid cells. In this report, we examine the biosynthesis and cell surface expression of CD4 in the human T cell line, CEM-SS, that has been stably transduced with the SIV nef gene. Quantification of CD4 in Nef-expressing cells reveals that the steady state level of CD4 is significantly reduced as compared to control transductants. The presence of Nef in these cells promotes the degradation of newly synthesized CD4 protein. The biosynthesis and oligosaccharide processing of CD4 in Nef-expressing T cells appears to be normal through the endoplasmic reticulum and Golgi compartments, suggesting that the degradation of CD4 is a late event in the biosynthetic pathway. Treatment with the lysosomotropic agents chloroquine and primaquine prevents the degradation of CD4 in Nef-expressing CEM-SS cells, indicating that the degradation of CD4 likely occurs in an acidic compartment. Thus the reduced cell surface expression observed in Nef-expressing CEM-SS cells is the likely consequence of a Nef-induced sorting of CD4 into a cellular compartment where CD4 is then degraded.