Spinophilin is recruited to the IS. (a and b) DCs (day six) were cocultured in the presence (+Antigen) or absence (−Antigen) of agonist peptide (MCC; a and b, respectively) and splenic CD4+ T cells. Confocal sections along the z axis of conjugated cells are shown in a and b. (a) In the presence of an antigen, topographical distribution of spinophilin (red), MHCII (green), and CD3, a component of the TCR (Cy5), reveals the polarization of spinophilin on the DC toward activated T cells. (b) In the absence of antigen, spinophilin remains localized close to the plasma membrane, as seen in mature DCs cultured in the absence of T cells (Fig. 1). Bar, 5 μm. (c) Correlation of spinophilin polarization in DCs with TCR clustering within T cells. Within DCs, the polarization of spinophilin toward contacting T cells correlated with TCR clustering to the contact site but not with MHCII. Error bars indicate the SEM.