Human CD8⁺ T cells do not require the polarization of lipid rafts for activation and proliferation

Lipid rafts are important signaling platforms in T cells. Little is known about their properties in human CD8⁺ T cells. We studied polarization of lipid rafts by digital immunofluorescence microscopy in primary human T cells, using beads coated with anti-CD3 and anti-CD28 mAbs (CD3/28 beads). Unlike CD4⁺ T cells, CD8⁺ T cells did not polarize lipid rafts when stimulated with CD3/28 beads, when the anti-CD28 antibody was substituted with B7.21g, or if an anti-CD8 antibody was added to the CD3/28 beads. This phenomenon was also observed in human antigen-specific CD8⁺ T cells. On stimulation with CD3/28 beads, the T cell antigen receptor clustered at the cell/bead contact area in both CD4⁺ and CD8⁺ T cells. Examination of lipid rafts isolated by sucrose density gradient centrifugation revealed the constitutive expression of p⁵⁶Lck in the raft fractions of unstimulated CD8⁺ T cells, whereas p⁵⁶Lck was recruited to the raft fraction of CD4⁺ T cells only after stimulation with CD3/28 beads. Stimulation with CD3/28 beads induced marked calcium flux, recruitment of PKC-θ and F-actin to the cell/bead contact site, and similar proliferation patterns in CD4⁺ and CD8⁺ T cells. Thus, polarization of lipid rafts is not essential for early signal transduction events or proliferation of human CD8⁺ lymphocytes. It is possible that the lower stringency of CD8⁺ T cell activation obviates a requirement for raft polarization.