The serine/threonine kinase Cdk5 plays an essential role in neuronal positioning during corticogenesis, but the underlying mechanisms are unknown. In nonneuronal cells, the tyrosine kinase FAK is a major regulator of cell motility through focal adhesions. It is unclear whether FAK plays a role in brain development. Here, we show that FAK phosphorylation by Cdk5 at S732 is important for microtubule organization, nuclear movement, and neuronal migration. In cultured neurons, S732-phosphorylated FAK is enriched along a centrosome-associated microtubule fork that abuts the nucleus. Overexpression of the nonphosphorylatable mutant FAK S732A results in disorganization of the microtubule fork and impairment of nuclear movement in vitro, and neuronal positioning defects in vivo. These observations are reminiscent of what is seen in the Cdk5-deficient mice. Taken together, these results suggest that Cdk5 phosphorylation of FAK is critical for neuronal migration through regulation of a microtubule fork important for nuclear translocation.

Neurons in the developing mammalian brain are generated from progenitor cells in the proliferative ventricular zone, and control of progenitor division is essential to produce the correct number of neurons during neurogenesis. Here we establish that Gbetagamma subunits of heterotrimeric G proteins are required for proper mitotic-spindle orientation of neural progenitors in the developing neocortex. Interfering with Gbetagamma function in progenitors causes a shift in spindle orientation from apical-basal divisions to planar divisions. This results in hyperdifferentiation of progenitors into neurons as a consequence of both daughter cells adopting a neural fate instead of the normal asymmetric cell fates. Silencing AGS3, a nonreceptor activator of Gbetagamma, results in defects similar to the impairment of Gbetagamma, providing evidence that AGS3-Gbetagamma signaling in progenitors regulates apical-basal division and asymmetric cell-fate decisions. Furthermore, our observations indicate that the cell-fate decision of daughter cells is coupled to mitotic-spindle orientation in progenitors.

Centrosome- and microtubule-associated proteins have been shown to be important for maintaining the neural progenitor pool during neocortical development by regulating the mitotic spindle. It remains unclear whether these proteins may control neurogenesis by regulating other microtubule-dependent processes such as nuclear migration. Here, we identify Cep120, a centrosomal protein preferentially expressed in neural progenitors during neocortical development. We demonstrate that silencing Cep120 in the developing neocortex impairs both interkinetic nuclear migration (INM), a characteristic pattern of nuclear movement in neural progenitors, and neural progenitor self-renewal. Furthermore, we show that Cep120 interacts with transforming acidic coiled-coil proteins (TACCs) and that silencing TACCs also causes defects in INM and neural progenitor self-renewal. Our data suggest a critical role for Cep120 and TACCs in both INM and neurogenesis. We propose that sustaining INM may be a mechanism by which microtubule-regulating proteins maintain the neural progenitor pool during neocortical development.

The mechanisms controlling neurogenesis during brain development remain relatively unknown. Through a differential protein screen with developmental versus mature neural tissues, we identified a group of developmentally enriched microtubule-associated proteins (MAPs) including doublecortin-like kinase (DCLK), a protein that shares high homology with doublecortin (DCX). DCLK, but not DCX, is highly expressed in regions of active neurogenesis in the neocortex and cerebellum. Through a dynein-dependent mechanism, DCLK regulates the formation of bipolar mitotic spindles and the proper transition from prometaphase to metaphase during mitosis. In cultured cortical neural progenitors, DCLK RNAi Lentivirus disrupts the structure of mitotic spindles and the progression of M phase, causing an increase of cell-cycle exit index and an ectopic commitment to a neuronal fate. Furthermore, both DCLK gain and loss of function in vivo specifically promote a neuronal identity in neural progenitors. These data provide evidence that DCLK controls mitotic division by regulating spindle formation and also determines the fate of neural progenitors during cortical neurogenesis.

Disabled-1 regulates laminar organization in the developing mammalian brain. Although mutation of the disabled-1 gene in scrambler mice results in abnormalities in neuronal positioning, migratory behavior linked to Disabled-1 signaling is not completely understood. Here we show that newborn neurons in the scrambler cortex remain attached to the process of their parental radial glia during the entire course of radial migration, whereas wild-type neurons detach from the glial fiber in the later stage of migration. This abnormal neuronal-glial adhesion is highly linked to the positional abnormality of scrambler neurons and depends intrinsically on Disabled-1 Tyr220 and Tyr232, potential phosphorylation sites during corticogenesis. Importantly, phosphorylation at those sites regulates alpha3 integrin levels, which is critical for the timely detachment of migrating neurons from radial glia. Altogether, these results outline the molecular mechanism by which Disabled-1 signaling controls the adhesive property of neurons to radial glia, thereby maintaining proper neuronal positioning during corticogenesis.