FAK-MAPK-dependent adhesion disassembly downstream of L1 contributes to semaphorin3A-induced collapse

Axonal receptors for class 3 semaphorins (Sema3s) are heterocomplexes of neuropilins (Nrps) and Plexin-As signalling coreceptors. In the developing cerebral cortex, the Ig superfamily cell adhesion molecule L1 associates with Nrp1. Intriguingly, the genetic removal of L1 blocks axon responses of cortical neurons to Sema3A in vitro despite the expression of Plexin-As in the cortex, suggesting either that L1 substitutes for Plexin-As or that L1 and Plexin-A are both required and mediate distinct roles. We report that association of Nrp1 with L1 but not Plexin-As mediates the recruitment and activation of a Sema3A-induced focal adhesion kinase-mitogen-activated protein kinase cascade. This signalling downstream of L1 is needed for the disassembly of adherent points formed in growth cones and subsequently their collapse response to Sema3A. Plexin-As and L1 are coexpressed and present in common complexes in cortical neurons and both dominant-negative forms of Plexin-A and L1 impair their response to Sema3A. Consistently, Nrp1-expressing cortical projections are defective in mice lacking Plexin-A3, Plexin-A4 or L1. This reveals that specific signalling activities downstream of L1 and Plexin-As cooperate for mediating the axon guidance effects of Sema3A.     

Semaphorin3A enhances endocytosis at sites of receptor-F-actin colocalization during growth cone collapse

Axonal growth cone collapse is accompanied by a reduction in filopodial F-actin. We demonstrate here that semaphorin 3A (Sema3A) induces a coordinated rearrangement of Sema3A receptors and F-actin during growth cone collapse. Differential interference contrast microscopy reveals that some sites of Sema3A-induced F-actin reorganization correlate with discrete vacuoles, structures involved in endocytosis. Endocytosis of FITC-dextran by the growth cone is enhanced during Sema3A treatment, and sites of dextran accumulation colocalize with actin-rich vacuoles and ridges of membrane. Furthermore, the Sema3A receptor proteins, neuropilin-1 and plexin, and the Sema3A signaling molecule, rac1, also reorganize to vacuoles and membrane ridges after Sema3A treatment. These data support a model whereby Sema3A stimulates endocytosis by focal and coordinated rearrangement of receptor and cytoskeletal elements. Dextran accumulation is also increased in retinal ganglion cell (RGC) growth cones, in response to ephrin A5, and in RGC and DRG growth cones, in response to myelin and phorbol-ester. Therefore, enhanced endocytosis may be a general principle of physiologic growth cone collapse. We suggest that growth cone collapse is mediated by both actin filament rearrangements and alterations in membrane dynamics.