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. 2006 Mar 16;116(5):1346–1359. doi: 10.1172/JCI27414

Figure 9. Clustering of CD16 chimeric proteins and induction of tyrosine phosphorylation on clustered CD16/7/nephrinCD (CD16/7/NCD).

Figure 9

(A) Schematic representation of CD16 fusion proteins prepared for these experiments. (B) Confocal microscopy imaging of NIH 3T3 cells expressing the CD16/7/GFP fusion protein before and after addition to media of live cells of anti-CD16 antibody and rhodamine-conjugated anti-IgG antibody. In the merged image at right, rhodamine IgG (red) and GFP (green) colocalized (yellow) on the plasma membrane, demonstrating clustering of the CD16/7/GFP chimeric protein. Note absence of clusters where anti-CD16 antibody was not present. Magnification, ×600. (C) NIH 3T3 cells expressing CD16/7/nephrinCD were treated with clustering antibodies (1° + 2° Ab) or as indicated for the time periods shown, then immunoblotted with P-nephrin antibody to detect phosphorylation of Y1191 and Y1208. Pretreatment of cells with PP2 blocked tyrosine phosphorylation on these sites following induction of clusters.