Enhancing the actin flow–L1 linkage by shootin1 overexpression promotes L1-dependent neurite outgrowth. (A and B) DIC micrographs showing retrograde movement of L1-Fc–coated beads on minor process growth cones (one day in vitro) overexpressing myc-GST (A, left) or myc-shootin1 (B, left), and a time series of the indicated areas at 30-s intervals (A and B, right). (C) The percentage of beads coated with L1-Fc that showed retrograde flow on minor process growth cones expressing myc-GST (n = 28) or myc-shootin1 (n = 35), mean velocity of moving beads as means ± SEM (*, P < 0.05), and the percentage of moving beads with indicated velocities. (D–F) Neurons overexpressing myc-shootin1 or myc-GST were cultured on L1-Fc–coated coverslips for 24 h (D). They were also cotransfected with control shRNA or shRNA against L1. (E) The lengths of total minor processes (lengths of all neurites except the longest one) relative to that of the control neurons expressing myc-GST and control shRNA. (F) The percentage increase in total minor processes upon expression of myc-shootin1 (mean ± SEM; *, P < 0.05; n = 4 independent cultures, 554 neurons were examined). Bars: (B), 5 μm; (D) 50 μm.