Mutation of the 3-Phosphoinositide-Dependent Protein Kinase 1 (PDK1) Substrate-Docking Site in the Developing Brain Causes Microcephaly with Abnormal Brain Morphogenesis Independently of Akt, Leading to Impaired Cognition and Disruptive Behaviors

Abstract

The phosphoinositide (PI) 3-kinase/Akt signaling pathway plays essential roles during neuronal development. 3-Phosphoinositide-dependent protein kinase 1 (PDK1) coordinates the PI 3-kinase signals by activating 23 kinases of the AGC family, including Akt. Phosphorylation of a conserved docking site in the substrate is a requisite for PDK1 to recognize, phosphorylate, and activate most of these kinases, with the exception of Akt. We exploited this differential mechanism of regulation by generating neuron-specific conditional knock-in mice expressing a mutant form of PDK1, L155E, in which the substrate-docking site binding motif, termed the PIF pocket, was disrupted. As a consequence, activation of all the PDK1 substrates tested except Akt was abolished. The mice exhibited microcephaly, altered cortical layering, and reduced circuitry, leading to cognitive deficits and exacerbated disruptive behavior combined with diminished motivation. The abnormal patterning of the adult brain arises from the reduced ability of the embryonic neurons to polarize and extend their axons, highlighting the essential roles that the PDK1 signaling beyond Akt plays in mediating the neuronal responses that regulate brain development. The phosphoinositide 3-kinase (PI3K) signaling pathway regulates cell survival, proliferation, growth, and motility, as well as metabolism, in response to extracellular signals. Class I PI3Ks phosphorylate the membrane phospholipid phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P 2 ] to generate the phosphatidylinositol-3,4,5-trisphosphate [PtdIns(3,4,5)P 3 ] second messenger (1, 2). In neurons, stimulation of PI3K by neurotrophic factors, neurotransmitters, or guidance cues results in the activation of protein kinase B (PKB) (also termed Akt), the most studied downstream effector of this signaling pathway. Akt phosphorylates and inactivates a number of cellular substrates controlling different aspects of neuronal development. They include PRAS40 and TSC2, leading to mammalian TORC1 (mTORC1) activation, which in turn promotes the synthesis of selected sets of proteins involved in the differentiation program (3); glycogen synthase kinase 3 (GSK3), which regulates cytoskeleton dynamics and participates in the establishment and maintenance of neuronal polarity (4, 5); and FOXO, which promotes the expression of genes inhibiting apoptosis (6). Genetic analysis in mice has uncovered the functional significance of PI3K for brain morphology and physiology (7-10), whereas deregulation of this signaling pathway has pathophysiological consequences in human neurodevelopmental disorders, such as schizophrenia (11-13) and autism (14, 15).3-Phosphoinositide-dependent protein kinase 1 (PDK1) transduces many agonist-induced cellular responses by activating an entire set of AGC kinase family members, in addition to Akt (16). They include S6K, SGK, RSK, and protein kinase C (PKC) isoforms. Upon cell stimulation, PDK1 is enabled to phosphorylate the T loops of all these AGC kinases, resulting in their activation (17, 18).Sin...

Conditional Deletion of PDK1 in the Forebrain Causes Neuron Loss and Increased Apoptosis during Cortical Development

“…However, the density of NeuN+ cells in PDK1 cKO mice was increased as compared to control animals ( Figure 2E ). This finding was in agreement with that reported on PDK1 conditional knock-in mice ( Cordon-Barris et al, 2016 ). Overall, conditional deletion of PDK1 in the forebrain resulted in remarkable loss of mature neurons.…”

“…Recent work showed that conditional deletion of PDK1 through GFAP-Cre mediated gene recombination causes microcephaly in mice, indicating a critical role of PDK1 in brain development ( Chalhoub et al, 2009 ). A conditional knock-in mouse model expressing the PDK1 L155E mutation displays microcephaly as well ( Cordon-Barris et al, 2016 ). In addition, it has been demonstrated that PDK1 in neural progenitor cells (NPCs) is important for the generation of oligodendrocyte precursor cells ( Watatani et al, 2012 ) and neuronal migration ( Itoh et al, 2016 ).…”

SGK1 Is a Critical Component of an AKT-Independent Pathway Essential for PI3K-Mediated Tumor Development and Maintenance

“…The Pdk1 L155E genetic model that uncouples AKT-dependent and AKT-independent pathways downstream of PI3K has been previously utilized to show that PI3K-dependent, AKT-independent pathways are required for T cell progenitors proliferation (27), leucine-induced mTORC1/S6K activation in the cardiac muscle (28), and proper brain patterning (29). Our data further extend these concepts by providing clear genetic evidence that activation of AGC kinases controlled by PDK1 via its PIF pocket is critical not only for the proliferative response to PI3K activation, but also for the ability of PI3K to drive neoplastic transformation and tumor progression in three well-characterized mouse models of follicular, poorly differentiated, and anaplastic thyroid cancer, respectively.…”

Clinical significance of miR-138 in patients with malignant melanoma through targeting of PDK1 in the PI3K/AKT autophagy signaling pathway

“…As an upstream gene of AKT, PDK1 regulates the activities of AKT and other downstream kinases of PI3K/ AKT, which enables PDK1 to regulate a series of biological responses in cells by the PI3K/AKT signaling pathway (27). PDK1 is a serine/threonine kinase of the AGC protein kinase family.…”