Article

Deletion of CASK in mice is lethal and impairs synaptic function

Department of Neuroscience, University of Texas Southwestern Medical Center, Dallas, TX 75390-9111, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 03/2007; 104(7):2525-30. DOI: 10.1073/pnas.0611003104
Source: PubMed

ABSTRACT CASK is an evolutionarily conserved multidomain protein composed of an N-terminal Ca2+/calmodulin-kinase domain, central PDZ and SH3 domains, and a C-terminal guanylate kinase domain. Many potential activities for CASK have been suggested, including functions in scaffolding the synapse, in organizing ion channels, and in regulating neuronal gene transcription. To better define the physiological importance of CASK, we have now analyzed CASK "knockdown" mice in which CASK expression was suppressed by approximately 70%, and CASK knockout (KO) mice, in which CASK expression was abolished. CASK knockdown mice are viable but smaller than WT mice, whereas CASK KO mice die at first day after birth. CASK KO mice exhibit no major developmental abnormalities apart from a partially penetrant cleft palate syndrome. In CASK-deficient neurons, the levels of the CASK-interacting proteins Mints, Veli/Mals, and neurexins are decreased, whereas the level of neuroligin 1 (which binds to neurexins that in turn bind to CASK) is increased. Neurons lacking CASK display overall normal electrical properties and form ultrastructurally normal synapses. However, glutamatergic spontaneous synaptic release events are increased, and GABAergic synaptic release events are decreased in CASK-deficient neurons. In contrast to spontaneous neurotransmitter release, evoked release exhibited no major changes. Our data suggest that CASK, the only member of the membrane-associated guanylate kinase protein family that contains a Ca2+/calmodulin-dependent kinase domain, is required for mouse survival and performs a selectively essential function without being in itself required for core activities of neurons, such as membrane excitability, Ca2+-triggered presynaptic release, or postsynaptic receptor functions.

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    • "In addition to the synaptic phenotype in heterozygous NRXN1 mutant neurons, we observed a dramatic increase in CASK protein levels (Figure 7), validating the interaction of neurexin-1 with CASK (Hata et al., 1996). It seems likely that neurexin-1 normally regulates CASK levels, which might be critical for controlling neurotransmitter release; indeed, constitutive CASK deletion in mice results in an increased mESPC frequency in cortical neurons (Atasoy et al., 2007). Interestingly, mutations in the human CASK gene are also associated with ASDs, mental retardation, and brain malformations, similar to NRXN1 mutations (Najm et al., 2008; Hackett et al., 2010; Sanders et al., 2012). "
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    ABSTRACT: Heterozygous mutations of the NRXN1 gene, which encodes the presynaptic cell-adhesion molecule neurexin-1, were repeatedly associated with autism and schizophrenia. However, diverse clinical presentations of NRXN1 mutations in patients raise the question of whether heterozygous NRXN1 mutations alone directly impair synaptic function. To address this question under conditions that precisely control for genetic background, we generated human ESCs with different heterozygous conditional NRXN1 mutations and analyzed two different types of isogenic control and NRXN1 mutant neurons derived from these ESCs. Both heterozygous NRXN1 mutations selectively impaired neurotransmitter release in human neurons without changing neuronal differentiation or synapse formation. Moreover, both NRXN1 mutations increased the levels of CASK, a critical synaptic scaffolding protein that binds to neurexin-1. Our results show that, unexpectedly, heterozygous inactivation of NRXN1 directly impairs synaptic function in human neurons, and they illustrate the value of this conditional deletion approach for studying the functional effects of disease-associated mutations. Copyright © 2015 Elsevier Inc. All rights reserved.
    Cell Stem Cell 08/2015; DOI:10.1016/j.stem.2015.07.017 · 22.15 Impact Factor
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    • "Whether the vertebrate-specific CASK-liprin interaction has additional synaptic roles is not clear, although a recent study in mammalian neurons suggests that depletion of liprin-α2 at mature synapses results in less synaptic localization of CASK [72]. Deletion of CASK in mice has little effect on synapse formation or structure [76]. Interestingly, liprin’s interaction via CASK’s CAMK domain suggests that it may compete with other proteins that interact at this site, such as Caskin [77] and the evolutionarily conserved CASK-Mint1 interaction [61]; the mutation at position 268 might thus disrupt CASK’s affinity for liprin-α enough to shift the predominant binding partner in vivo. "
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    ABSTRACT: The overwhelming amount of available genomic sequence variation information demands a streamlined approach to examine known pathogenic mutations of any given protein. Here we seek to outline a strategy to easily classify pathogenic missense mutations that cause protein misfolding and are thus good candidates for chaperone-based therapeutic strategies, using previously identified mutations in the gene CASK. We applied a battery of bioinformatics algorithms designed to predict potential impact on protein structure to five pathogenic missense mutations in the protein CASK that have been shown to underlie pathologies ranging from X-linked mental retardation to autism spectrum disorder. A successful classification of the mutations as damaging was not consistently achieved despite the known pathogenicity. In addition to the bioinformatics analyses, we performed molecular modeling and phylogenetic comparisons. Finally, we developed a simple high-throughput imaging assay to measure the misfolding propensity of the CASK mutants in situ. Our data suggests that a phylogenetic analysis may be a robust method for predicting structurally damaging mutations in CASK. Mutations in two evolutionarily invariant residues (Y728C and W919R) exhibited a strong propensity to misfold and form visible aggregates in the cytosolic milieu. The remaining mutations (R28L, Y268H, and P396S) showed no evidence of aggregation and maintained their interactions with known CASK binding partners liprin-α3 Mint-1, and Veli, indicating an intact structure. Intriguingly, the protein aggregation caused by the Y728C and W919R mutations was reversed by treating the cells with a chemical chaperone (glycerol), providing a possible therapeutic strategy for treating structural mutations in CASK in the future.
    PLoS ONE 02/2014; 9(2):e88276. DOI:10.1371/journal.pone.0088276 · 3.23 Impact Factor
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    • "In its role as scaffold protein, CASK links different adaptors and molecules (including other channels) to elicit further downstream signaling, like the stability and trafficking of receptors towards the membrane (Hsueh, 2011) and vesicle release (Spangler et al., 2013). In brain synapses, CASK has a negative role, as facilitated glutamatergic release was observed in KO CASK mice (Atasoy et al., 2007), in agreement with the inhibitory effect of CASK over the P2X3 over-reactivity (Gnanasekaran et al., 2013). "
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