Fig. 8. JAM-2 recruits PAR-3 and ZO-1 in CHO cells. (A) CHO cells stably transfected with JAM-2 (JAM-2, left panel), the S281A mutant of JAM-2 (J-2 S281A, middle panel) or the S281D mutant of JAM-2 (J-2 S281D, right panel) were stained with a mAb against JAM-2. Wild-type JAM-2 is barely detectable at cell-cell junctions and appears as discrete punctate staining (small inset in left panel). By contrast, the S281A mutant of JAM-2 is predominantly clustered at intercellular contacts. The S281D mutant of JAM-2 behaves like wt JAM-2 and is rarely localized at cell-cell contacts. All three cell lines showed a comparable surface expression of the transfected constructs as analysed by FACS analysis (not shown). Bar, 100 µm. (B) CHO cells stably transfected with the S281A mutant of JAM-2 were simultaneously stained with antibodies against JAM-2 and either PAR-3, ZO-1 or HSP-90, followed by Cy-3-conjugated secondary antibodies to detect JAM-2 or Cy-2-conjugated secondary antibodies to detect PAR-3, ZO-1 or HSP-90. Both PAR-3 and ZO-1 were recruited by JAM-2 to sites of cell-cell contacts. Bar, 5 µm.