Fig. 1. Kalirin-7 produces a Cdk5-dependent response in PC12 cells. (A) Domains of Kalirin-7 and its N-terminal -splice variant; S, Sec14P-like; DH, Dbl homology; P, pleckstrin homology; T1590PAK, sequence of putative Cdk5 phosphorylation site. (B) PC12 cells were fixed 24 hours after transfection with vectors encoding GFP or Kalirin-7. Transfected GFP protein was visualized using fluorescence or Myc antibody (green); filamentous actin was visualized using TRITC-phalloidin (red). White arrowheads indicate protrusions, stars indicate nontransfected cells, arrow indicates nucleus. (C) PC12 cells transfected with dual promoter vector encoding GFP (green) and DN Cdk5 (blue); TRITC-phalloidin (red). GFP-expressing cells always expressed DN Cdk5. (D) PC12 cells cotransfected with vectors encoding Kalirin-7 (Myc; blue) and GFP/DN Cdk5 (GFP, green); TRITC-phalloidin (red). White stars indicate nontransfected cells. (E) Using the GFP or Myc signal to identify cell margins, perimeter and cell roundness were evaluated using SimplePCI software: control (GFP), n=18; Kalirin-7 (Kal7), n=15; Kalirin-7 with DN Cdk5, n=24; P values calculated using the Student's t-test (two-tailed). Data are means ± s.e.m.; #, P<0.001. Scale bars: 10 µm (B-D).