, 1999; Rice and Curran, 2001). It was also reported that Reelin could BIBF 1120 price bind to an integrin receptor (Dulabon et al., 2000), although the effects of this interaction for neuronal migration is controversial (Magdaleno and Curran, 2001). In this study, we show that after Reelin binds to ApoER2/VLDLR,

it activates integrin α5β1 on the migrating neurons through the intracellular Dab1-Crk/CrkL-C3G-Rap1 pathway (“inside-out” activation of integrin) (Kinashi, 2005; Shattil et al., 2010), which promotes neuronal adhesion to fibronectin. Since fibronectin is present in the MZ, activated integrin α5β1 (a fibronectin receptor) then mediates terminal translocation through the PCZ. Furthermore, sequential in utero electroporation studies show that this integrin activation is indeed required for proper establishment of the eventual neuronal positioning in the mature cortex in vivo. Interestingly, whereas the Rap1-N-cadherin pathway is involved in the migration below the CP (Jossin and Cooper, 2011), we found that it could not promote neuronal entry into the PCZ by terminal translocation, suggesting that Rap1 has dual functions during different phases of neuronal migration and that Reelin changes the downstream adhesion molecules of Rap1 during terminal translocation. Our data

suggest that Reelin-dependent modulation of neuronal adhesion is critical for the eventual birthdate-dependent Selleckchem PARP inhibitor neuronal layering in the neocortex. Several studies have reported that Dab1 is required for terminal translocation, which is necessary for the establishment of the birthdate-dependent “inside-out” neuronal layering (Olson et al., 2006; Cooper, 2008; Franco et al., 2011; Sekine et al., 2011). Since Dab1 is a multifunctional adaptor protein that can selectively recruit several (-)-p-Bromotetramisole Oxalate downstream molecules to its specific phosphorylation sites (Honda et al., 2011), we first analyzed the effects of Dab1 phosphorylation on terminal translocation using various tyrosine mutants of Dab1. When a Dab1-knockdown (KD) vector was introduced into the mouse

embryonic neocortex by in utero electroporation at embryonic day 14.5 (E14.5), the transfected cells were mislocated just beneath the NeuN-negative region of the CP or the PCZ (Sekine et al., 2011) on postnatal day 0.5 (P0.5), 5 days after the electroporation (Figures 1A–1B′), suggesting that terminal translocation was disrupted. This Dab1-KD phenotype was rescued by cotransfection of the cells with wild-type Dab1 (Figures S1A and S1B available online). Dab1 has five potential tyrosine residues phosphorylated by Reelin (tyrosines 185, 198, 200, 220, and 232) and Dab1-5F, lacking all of these tyrosine residues, Dab1-3F, lacking the three main phosphorylation sites (198F, 220F, and 232F) (Keshvara et al.