Background:  Stimulation of naïve CD4 T cells with anti-CD3/CD28-coated beads leads to rapid polarization of lipid rafts. Since neither stimulus alone can polarize lipid rafts, it has been postulated that a major role of co-stimulation is to facilitate lipid raft aggregation. CD86 is up-regulated or expressed aberrantly on immune cells in a wide array of autoimmune and infectious diseases, including HIV-1 infection.

Methods:  To ligate CD28 on human CD4 T cells, we used an immunoglobulin (Ig) fusion with the extracellular domain of CD86 (CD86Ig) bound to a magnetic bead or K562 cells expressing CD86. Cell-bead conjugates were plated onto coverslips, stained with anti-GM1 antiserum or choleratoxin B, and lipid raft polarization was visualized by digital immunofluorescence microscopy. Signaling pathways involved in T-cell activation, including increase of intracellular calcium, translocation of NFkB to the nucleus, and gene transcription were assessed.

Results:  Ligation of CD28 by natural ligand, but not by antibody, induced polarization of lipid rafts at the cell-bead interface of fresh human CD4 T cells, in the absence of TCR ligation. This correlated with activation of Vav-1, increase of the intracellular calcium concentration and nuclear translocation of NFkB p65, but did not result in T-cell proliferation or cytokine production. Using DNA microarrays, we detected induction of a small subset of genes, including members of the EGR1 family of transcription factors, upon CD28 ligation. Engagement of CTLA-4 blocked CD86Ig induction of both lipid raft polarization and new transcription.

Conclusions:  We show that lipid raft polarization in human CD4 T cells can occur in the absence of TCR triggering, driven solely by the CD28/CD86 interaction. This result has implications for mechanisms of T-cell activation. Abnormalities in this process could alter T- and B-cell tolerance and susceptibility to infection. HIV virions preferentially incorporate CD86 into their membranes and lipid rafts facilitate HIV entry. These virions have been shown to trigger NFkB activation in a CD86-dependent manner similar to that found in our studies. The heightened immune activation observed in HIV-infected individuals enhances CD86 expression, which in turn could induce lipid rafts polarization between infected cells and resting T cells, permitting formation of the virological synapse and HIV entry into non-cycling T cells. The ability of CD86 to induce lipid rafts may in part explain the susceptibility of resting T cells to HIV infection.

Keywords: T-cell Responses; Cellular Co-Factors; Virological Synapse