Cdk5 phosphorylation of ErbB4 is required for tangential migration of cortical interneurons.
Bottom Line: Interneuron dysfunction in humans is often associated with neurological and psychiatric disorders, such as epilepsy, schizophrenia, and autism.Some of these disorders are believed to emerge during brain formation, at the time of interneuron specification, migration, and synapse formation.This finding identifies Cdk5 as a crucial signaling factor in cortical interneuron development in mammals.
Affiliation: Department of Cell and Developmental Biology, University College London, London WC1 6BT, UK.Show MeSH
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Mentions: A number of studies have suggested a strong correlation between Cdk5 activity and the ErbB receptor pathway (Fu et al. 2001, 2005; Li et al. 2003; Xie et al. 2004, 2007). To determine whether p35/Cdk5 phosphorylates ErbB4 in vitro, we carried out experiments using COS7 cells that naturally express Cdk5, but not p35 or ErbB4. Cdk5 phosphorylates proteins containing a [S/T]Px[K/H/R] site (Fig. 3A). We focused on threonine 1152 (T1152PmH/R) that lies in close proximity to Cyt1-specific PI3-kinase-binding site Y1056. To initially test whether ErbB4 is a substrate of Cdk5, we used a phosphospecific ab that recognizes a minimal Cdk consensus site, phosphorylated threonine adjacent to a proline (T(PO4)P), and discovered a stronger ErbB4 phosphorylation in COS7 cells that co-expressed p35/Cdk5 compared with other experimental conditions (Fig. 4A). To further confirm that ErbB4 is indeed phosphorylated by Cdk5, we performed an in vitro p35/Cdk5 kinase assay using recombinant GST-fused ErbB4 fragments containing either the native T1152 residue (GST-T) or an alanine point mutation (T1152A), which was obtained by site-directed mutagenesis and cannot be phosphorylated [GST-T(A); Fig. 4B]. GST and histone 1, an established substrate of Cdk5, were used as negative and positive controls, respectively. This assay revealed a robust phosphorylation by p35/Cdk5 of intact (GST-T), but not mutated [GST-T(A)] ErbB4 fragments (Fig. 4C), identifying ErbB4 as a Cdk5 substrate in vitro.Figure 4.
Affiliation: Department of Cell and Developmental Biology, University College London, London WC1 6BT, UK.