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David A Keays

Publications and source records attributed to David A Keays.

6 recordsLinked to original sources
PubMed2026

A complex of MAST1 and 14-3-3η regulates Tau phosphorylation in the developing cortex.

The MAST family of serine/threonine kinases has been implicated in a spectrum of human neurodevelopmental disorders. However, little is known about their biological function or regulation. Seeking to fill these gaps in our knowledge, we have identified upstream and downstream partners of MAST1. 14-3-3η, a neuronal 14-3-3 paralog, specifically interacts with MAST1 at two regulatory serines, S90 and S161. p21-activated kinase (PAK), a neuronal regulator of the actin cytoskeleton, phosphorylates MAST1 to regulate its interaction with 14-3-3η. Exploiting mouse models of human Mega-Corpus-Callosum Syndrome (MCC) and whole brain phosphoproteomics, we identify the microtubule-associated protein Tau as a candidate substrate of MAST1. We show that pathogenic MAST1 mutations perturb protein function either through misfolding or attenuation of kinase activity. Our data are consistent with a model in which the MAST kinases couple PAK, a neuronal regulator of the actin cytoskeleton, to microtubule remodeling during the differentiation and specification of cortical neurons.

Animals↗
PubMed2007

Mutations in alpha-tubulin cause abnormal neuronal migration in mice and lissencephaly in humans.

The development of the mammalian brain is dependent on extensive neuronal migration. Mutations in mice and humans that affect neuronal migration result in abnormal lamination of brain structures with associated behavioral deficits. Here, we report the identification of a hyperactive N-ethyl-N-nitrosourea (ENU)-induced mouse mutant with abnormalities in the laminar architecture of the hippocampus and cortex, accompanied by impaired neuronal migration. We show that the causative mutation lies in the guanosine triphosphate (GTP) binding pocket of alpha-1 tubulin (Tuba1) and affects tubulin heterodimer formation. Phenotypic similarity with existing mouse models of lissencephaly led us to screen a cohort of patients with developmental brain anomalies. We identified two patients with de novo mutations in TUBA3, the human homolog of Tuba1. This study demonstrates the utility of ENU mutagenesis in the mouse as a means to discover the basis of human neurodevelopmental disorders.

Amino Acid Sequence↗
PubMed2006

Estimating the number of coding mutations in genotypic- and phenotypic-driven N-ethyl-N-nitrosourea (ENU) screens.

N-ethyl-N-nitrosourea (ENU) is a widely used mutagen in genotypic and phenotypic screens aimed at elucidating gene function. The high rate at which ENU induces point mutations raises the possibility that an observed phenotype may be to the result of another unidentified linked mutation. This article presents methods for estimating the probability of additional linked coding mutations (1) in a given region of DNA using both Poisson and Bayesian models and in (2) an F(1) animal exposed to ENU that has undergone b number of backcrosses. Applying these methods to the mouse data set of Quwailid et al., we estimate that the probability that a confounding mutation is linked to a cloned mutation when the candidate region is 5 Mb is very slim (p < 0.002). Where mutants are identified by genotypic methods, we show that backcrossing in the absence of marker-assisted selection is an inefficient means of eliminating linked confounding mutations.

Animals↗
PubMed2004

New semidominant mutations that affect mouse development.

Dominantly acting mutations that produce visible phenotypes are frequently recovered, either during routine maintenance of colonies or from mutagenesis experiments. We have studied 12 dominant mouse mutations that cause a tail dysmorphology, a coat spotting phenotype, or a combination of these. The majority of these mutations act in a semidominant manner with the homozygous state associated with embryonic lethality and a visible phenotype at or before midgestation. The homozygous phenotypes include axis truncation and neural crest cell defects, as may be expected from the heterozygous phenotypes. The majority of mutations, however, also produced other phenotypes that include neural tube closure defects and aberrant heart looping. In one coat spotting mutant the homozygous condition is lethal before neural crest cell production commences. The mutated genes often function in processes additional to those alluded to by the heterozygous phenotype.

Alkylating Agents↗
PubMed2004

Determining sequences and post-translational modifications of novel conotoxins in Conus victoriae using cDNA sequencing and mass spectrometry.

A combination of cDNA cloning and detailed mass spectrometric analyses was employed to identify novel conotoxins from Conus victoriae. Eleven conotoxin sequences were determined using molecular methods: one belonging to the A superfamily (Vc1.1), six belonging to the O superfamily (Vc6.1-Vc6.6) and four members of the T superfamily (Vc5.1-Vc5.4). In order to verify the sequences and identify the post-translational modifications (excluding the disulfide connectivity) of three Conus victoriae conotoxins, vc1a, vc5a and vc6a, deduced from sequences Vc1.1, Vc5.1, and Vc6.1, respectively, liquid chromatography/electrospray ionization ion trap mass spectrometry, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and nanospray ionization ion trap mass spectrometry with collisionally induced dissociation were performed on reduced and alkylated venom fractions. We report that vc1a, the native form of alpha-conotoxin Vc1.1 (an unmodified 16 amino acid residue peptide that has notable pain-relieving capabilities), includes a hydroxyproline and a gamma-carboxyglutamate residue. Conotoxin vc5a is a 10-residue peptide with two disulfide bonds and a hydroxyproline and vc6a is a 25 amino acid peptide with three disulfide bonds.

Amino Acid Sequence↗
PubMed2003

N-ethyl-N-nitrosourea mouse mutants in the dissection of behavioural and psychiatric disorders.

Twin and adoption studies have consistently implicated genetics in the aetiology of psychiatric and behavioural disorders. The identification of the genes and molecular pathways that are associated with these traits using linkage studies has been difficult because psychiatric disorders are almost always non-mendelian, heterogeneous, involve multiple genetic loci and are influenced significantly by environmental factors. Mouse models that are based on intermediate signatures of psychiatric disease and pharmacological responsiveness hold promise as a complementary approach to dissecting the molecular basis of neurobehavioural disorders. This has been made possible by the development and refinement of gene targeting technologies and the use of super-efficient chemical mutagens. N-ethyl-N-nitrosourea (ENU) mutagenesis in the mouse, when coupled to a battery of sensitive behavioural screens, is an effective way of creating and identifying novel mouse behavioural mutants. Here, the concept of screening for ENU mutants is introduced while progress with two behavioural screens, an "anxiety" screen and a circadian screen, are presented. It is hoped that the study of mouse mutants that have arisen from these screens will provide new insights into the genetic basis of abnormal behaviour and that they might lead to the development of novel therapeutic compounds for human psychiatric disease.

Animals↗