Mechanism for down-regulation of CD28 by Nef

SIV and HIV Nef proteins disrupt T-cell receptor machinery by down-modulati ng cell surface expression of CD4 and expression or signaling of CD3-TCR. N ef also down-modulates class I major histocompatibility complex (MHC) surfa ce expression. We show that SIV and HIV-1 Nefs down-modulate CD28, a major co-stimulatory receptor that mediates effective T-cell activation, by accel erating CD28 endocytosis, The effects of Nef on CD28, CD4, CD3 and class I MHC expression are all genetically separable, indicating that all are selec ted independently. In cells expressing a Nef-green fluorescent protein (GFP ) fusion, CD28 co-localizes with the AP-2 clathrin adaptor and Nef-GFP. Mut ations that disrupt Nef interaction with AP-2 disrupt CD28 down-regulation. Furthermore, HIV and SIV Nefs use overlapping but distinct target sites in the membrane-proximal region of the CD28 cytoplasmic domain. Thus, Nef pro bably induces CD28 endocytosis via the AP-2 pathway, and this involves a te rnary complex containing Nef, AP-2 and CD28. The likely consequence of the concerted down-regulation of CD28, CD4 and/or CD3 by Nef is disruption of a ntigen-specific signaling machineries in infected T cells following a produ ctive antigen recognition event.