Mutant EWI-2 with altered spreading and association properties. (A) MOLT-4 cells stably transduced with either EWI-2–FLAG or EWI-2/cCD2 were plated on VCAM-Ig–coated coverslips (5 μg/mL or 2.5 μg/mL), and photographed after 65 minutes using phase contrast (top row) or Hoffman objectives (bottom row). Percent of the spread cells (quantified as described in “Materials and methods”) is indicated in the lower part of the micrographs (original magnification, × 20). (B) MOLT-4 cells stably transduced with vector control (lane 1), CD2 (lane 2), EWI-2 (lane 3), or EWI-2/cCD2 were lysed in 1% Brij 58. Proteins were immunoprecipitated with M2 (anti-FLAG), A4PUJ (α4), or M38 (CD81) mAbs. In the first panel, cells were surface biotinylated prior to lysis. Sizes (kDa) of molecular weight markers are shown on the right.