Myosin X Interacts With N-cadherin To Regulate Neuronal Radial Migration

During neocortical brain development, postmitotic neurons newly generated in theventricular and subventricular zones migrate radially toward the pial surface along theradial glia cells to form the six-layered cortical structure which is essential for brainfunction. The neuron-neuron or neuron-glia adhesion is essential for the neuroal polaritydevelopment and radial migration. One of the features of neural cells in the developingcentral nervous system (CNS) is their ability to migrate away from their origins ingerminal zones to their final positions in cell assemblies. The research about themechanisms in neuronal migration will give new insights into that regulate migration andunderstand migration abnormalities in several naturally occurring genetic defects inhumans.Myosin X (Myo10), an uncharacteristic member of the myosin family, localizes atthe tips of filopodia and other actin-rich peripheral protrusions, and is critical for filopodiaformation as well as cell motility. Recently, Myo10has been implicated as requirementfor migration of cranial neural crest cell. During the brain development, a headless Myo10has been detected, which is involved in axon outgrowth and neuronal radial migrationcoordinating with full-length Myo10during the developing cerebral cortex, but themechanism is still largely unknown.Here, we provided evidence for the involvement of Myo10in N-cadherin mediatedcell adhesion and its participation in the neuronal migration. We showed that Myo10wasco-localized with N-cadherin at the cell contacts. Both Myo10and N-cadherin weredetected in the immunoprecipitate complexes from NLT cells and embryonic brain lysate.To determine the putative regions necessary for interaction, we designed a series ofMyo10deletion mutants and GST-N-cadherin cytoplasmic domain fusion proteinconstructs. Immunoprecipitation and glutathion-S-transferase (GST)-pulldown assaysconfirm that Myo10interacts with the last62amino acids of the cytoplasmic tail ofN-cadherin through a band4.1-ezrin-radixin-moesin (FERM) domain. We nextdetermined whether Myo10regulates N-cadherin subcellular localization.Immunofluorescence assay showed that the distribution of N-cadherin markedlydecreased at the cell-cell contacts in Myo10knockdown cells. Western-blotting confirmedthat knockdown of Myo10decreased the surface-associated fraction of N-cadherin. As ahomophilic adhesion molecule, N-cadherin regulates cell-cell interaction by theextracellular domains in a calcium-dependent way. The cytosolic domain of N-cadherin isassociated with the actin cytoskeleton through β-and α-catenins to modulate the celladhesion and motility. N-cadherin binding assay showed that knockdown of Myo10impaired the N-cadherin-mediated neuron adhesion. Previous studies have shown thatN-cadherin is important for the interaction between migration neurons and radial glial fibers. To investigate if Myo10also involves in radial glial-guilded neuronal migration invivo, we performed in utero electroporation. The results suggested that suppression ofMyo10impaired the adhesion between neurons and radial glial fibers, and some neuronsexhibit miss-oriented guiding processes in the radial migration zone. Furthermore, therescue expreiment showed that N-cadherin can partially rescue the migration deficiencycaused by Myo10-knockdown. These results suggest that Myo10plays a role in theregulation of N-cadherin-mediated adhesion during neuronal migration in corticaldevelopment.