Showing papers by "Christopher A. Walsh published in 2011"

Recapitulation of premature ageing with iPSCs from Hutchinson–Gilford progeria syndrome

Abstract: Hutchinson-Gilford progeria syndrome (HGPS) is a rare and fatal human premature ageing disease, characterized by premature arteriosclerosis and degeneration of vascular smooth muscle cells (SMCs). HGPS is caused by a single point mutation in the lamin A (LMNA) gene, resulting in the generation of progerin, a truncated splicing mutant of lamin A. Accumulation of progerin leads to various ageing-associated nuclear defects including disorganization of nuclear lamina and loss of heterochromatin. Here we report the generation of induced pluripotent stem cells (iPSCs) from fibroblasts obtained from patients with HGPS. HGPS-iPSCs show absence of progerin, and more importantly, lack the nuclear envelope and epigenetic alterations normally associated with premature ageing. Upon differentiation of HGPS-iPSCs, progerin and its ageing-associated phenotypic consequences are restored. Specifically, directed differentiation of HGPS-iPSCs to SMCs leads to the appearance of premature senescence phenotypes associated with vascular ageing. Additionally, our studies identify DNA-dependent protein kinase catalytic subunit (DNAPKcs, also known as PRKDC) as a downstream target of progerin. The absence of nuclear DNAPK holoenzyme correlates with premature as well as physiological ageing. Because progerin also accumulates during physiological ageing, our results provide an in vitro iPSC-based model to study the pathogenesis of human premature and physiological vascular ageing.

Human Mutations in NDE1 Cause Extreme Microcephaly with Lissencephaly

Abstract: Genes disrupted in human microcephaly (meaning "small brain") define key regulators of neural progenitor proliferation and cell-fate specification. In comparison, genes mutated in human lissencephaly (lissos means smooth and cephalos means brain) highlight critical regulators of neuronal migration. Here, we report two families with extreme microcephaly and grossly simplified cortical gyral structure, a condition referred to as microlissencephaly, and show that they carry homozygous frameshift mutations in NDE1, which encodes a multidomain protein that localizes to the centrosome and mitotic spindle poles. Both human mutations in NDE1 truncate the C-terminal NDE1domains, which are essential for interactions with cytoplasmic dynein and thus for regulation of cytoskeletal dynamics in mitosis and for cell-cycle-dependent phosphorylation of NDE1 by Cdk1. We show that the patient NDE1 proteins are unstable, cannot bind cytoplasmic dynein, and do not localize properly to the centrosome. Additionally, we show that CDK1 phosphorylation at T246, which is within the C-terminal region disrupted by the mutations, is required for cell-cycle progression from the G2 to the M phase. The role of NDE1 in cell-cycle progression probably contributes to the profound neuronal proliferation defects evident in Nde1-null mice and patients with NDE1 mutations, demonstrating the essential role of NDE1 in human cerebral cortical neurogenesis.

Neurogenesis at the brain-cerebrospinal fluid interface.

Abstract: Cerebral cortical progenitor cells can be classified into several different types, and each progenitor type integrates cell-intrinsic and cell-extrinsic cues to regulate neurogenesis On one hand, cell-intrinsic mechanisms that depend upon appropriate apical-basal polarity are established by adherens junctions and apical complex proteins and are particularly important in progenitors with apical processes contacting the lateral ventricle The apical protein complexes themselves are concentrated at the ventricular surface, and apical complex proteins regulate mitotic spindle orientation and cell fate On the other hand, remarkably little is known about how cell-extrinsic cues signal to progenitors and couple with cell-intrinsic mechanisms to instruct neurogenesis Recent research shows that the cerebrospinal fluid, which contacts apical progenitors at the ventricular surface and bathes the apical complex of these cells, provides growth- and survival-promoting cues for neural progenitor cells in developing and adult brain This review addresses how the apical-basal polarity of progenitor cells regulates cell fate and allows progenitors to sample diffusible signals distributed by the cerebrospinal fluid We also review several classes of signaling factors that the cerebrospinal fluid distributes to the developing brain to instruct neurogenesis

COL4A1 mutations cause ocular dysgenesis, neuronal localization defects, and myopathy in mice and Walker-Warburg syndrome in humans.

Abstract: Muscle-eye-brain disease (MEB) and Walker Warburg Syndrome (WWS) belong to a spectrum of autosomal recessive diseases characterized by ocular dysgenesis, neuronal migration defects, and congenital muscular dystrophy. Until now, the pathophysiology of MEB/WWS has been attributed to alteration in dystroglycan post-translational modification. Here, we provide evidence that mutations in a gene coding for a major basement membrane protein, collagen IV alpha 1 (COL4A1), are a novel cause of MEB/WWS. Using a combination of histological, molecular, and biochemical approaches, we show that heterozygous Col4a1 mutant mice have ocular dysgenesis, neuronal localization defects, and myopathy characteristic of MEB/WWS. Importantly, we identified putative heterozygous mutations in COL4A1 in two MEB/WWS patients. Both mutations occur within conserved amino acids of the triple-helix-forming domain of the protein, and at least one mutation interferes with secretion of the mutant proteins, resulting instead in intracellular accumulation. Expression and posttranslational modification of dystroglycan is unaltered in Col4a1 mutant mice indicating that COL4A1 mutations represent a distinct pathogenic mechanism underlying MEB/WWS. These findings implicate a novel gene and a novel mechanism in the etiology of MEB/WWS and expand the clinical spectrum of COL4A1-associated disorders.

Congenital Microcephaly with a Simplified Gyral Pattern: Associated Findings and Their Significance

Abstract: BACKGROUND AND PURPOSE: Primary microcephaly is an incompletely understood malformation that is often associated with developmental brain anomalies, yet whether the associated anomalies result from the microcephaly itself or from associated developmental/genetic mishaps is not yet understood This study reviewed and analyzed a large number of MR imaging scans of children with microcephaly to determine the frequency of associated morphologic findings and to assess whether these findings were associated with the severity of the microcephaly MATERIALS AND METHODS: MR images of 119 patients with clinically diagnosed microcephaly were retrospectively reviewed, focusing on the degree of microcephaly, simplification of gyri, white matter volume, abnormalities of the corpus callosum, size and structure of posterior fossa contents, and myelination Associations among the findings were evaluated by using the Spearman correlation coefficient and the Fisher exact test RESULTS: Among 7 patients with mild, 42 with moderate, and 70 with extreme microcephaly, a significant correlation was identified between a greater degree of microcephaly and both a greater degree of simplified gyration and decreased white matter volume The severity of the callosal anomaly showed a lower but still significant correlation with the severity of microcephaly Degree of hypoplasia of posterior fossa structures, delay in myelination, and abnormality of the basal ganglia did not correlate with the degree of microcephaly CONCLUSIONS: A strong correlation was found between the degree of microcephaly, the volume of white matter, and the presence of a simplified gyral pattern These associations should be considered when attempting to use neuroimaging for segregation and classification of patients with microcephaly

A forward genetic screen with a thalamocortical axon reporter mouse yields novel neurodevelopment mutants and a distinct emx2 mutant phenotype

Abstract: The dorsal thalamus acts as a gateway and modulator for information going to and from the cerebral cortex. This activity requires the formation of reciprocal topographic axon connections between thalamus and cortex. The axons grow along a complex multistep pathway, making sharp turns, crossing expression boundaries, and encountering intermediate targets. However, the cellular and molecular components mediating these steps remain poorly understood. To further elucidate the development of the thalamocortical system, we first created a thalamocortical axon reporter line to use as a genetic tool for sensitive analysis of mutant mouse phenotypes. The TCA-tau-lacZ reporter mouse shows specific, robust, and reproducible labeling of thalamocortical axons (TCAs), but not the overlapping corticothalamic axons, during development. Moreover, it readily reveals TCA pathfinding abnormalities in known cortical mutants such as reeler. Next, we performed an unbiased screen for genes involved in thalamocortical development using random mutagenesis with the TCA reporter. Six independent mutant lines show aberrant TCA phenotypes at different steps of the pathway. These include ventral misrouting, overfasciculation, stalling at the corticostriatal boundary, and invasion of ectopic cortical cell clusters. An outcross breeding strategy coupled with a genomic panel of single nucleotide polymorphisms facilitated genetic mapping with small numbers of mutant mice. We mapped a ventral misrouting mutant to the Emx2 gene, and discovered that some TCAs extend to the olfactory bulbs in this mutant. Mapping data suggest that other lines carry mutations in genes not previously known for roles in thalamocortical development. These data demonstrate the feasibility of a forward genetic approach to understanding mammalian brain morphogenesis and wiring. A robust axonal reporter enabled sensitive analysis of a specific axon tract inside the mouse brain, identifying mutant phenotypes at multiple steps of the pathway, and revealing a new aspect of the Emx2 mutant. The phenotypes highlight vulnerable choice points and latent tendencies of TCAs, and will lead to a refined understanding of the elements and interactions required to form the thalamocortical system. See Commentary: http://www.biomedcentral.com/1741-7007/9/1

Phenotypic heterogeneity in Woodhouse-Sakati syndrome: two new families with a mutation in the C2orf37 gene.

Abstract: Hypogonadism, alopecia, diabetes mellitus, mental retardation, and extrapyramidal syndrome [also known as Woodhouse-Sakati syndrome (WSS)] is a rare autosomal recessive neuroendocrine and ectodermal disorder. The syndrome was first described by Woodhouse and Sakati in 1983, and 47 patients from 23 families have been reported so far. We report on an additional seven patients (four males and three females) from two highly consanguineous Arab families from Qatar, presenting with a milder phenotype of WSS. These patients show the spectrum of clinical features previously found in WSS, but lack evidence of diabetes mellitus and extrapyramidal symptoms. These two new families further illustrate the natural course and the interfamilial phenotypic variability of WSS that may lead to challenges in making the diagnosis. In addition, our study suggests that WSS may not be as infrequent in the Arab world as previously thought.

Variable disease severity in Saudi Arabian and Sudanese families with c.3924 + 2 T > C mutation of LAMA2.

Abstract: Background: Congenital muscular dystrophy type 1A is caused by mutations in the LAMA2 gene that encodes the laminin a2 chain, a component of the skeletal muscle extracellular matrix protein laminin-211. The clinical spectrum of the disease is more heterogeneous than previously thought, particularly in terms of motor achievement and disease progression. We investigated clinical findings and performed molecular genetic analysis in 3 families from Saudi Arabia and 1 from Sudan in whom congenital muscular dystrophy 1A was suspected based on homozygosity mapping and laminin a2 chain deficiency. Methods: We investigated 9 affected individuals from 1 Sudanese and 3 Saudi families in whom MDC1A was suggested by clinical, neuroimaging and/or pathological findings and by homozygosity mapping at the LAMA2 locus. Morphological and immunohistochemical analysis were performed in 3 patients from the 3 Saudi families. SSCP analysis, DNA sequencing and microsatellite analysis were carried out in the 4 index cases. Results: A previously described mutation in the LAMA2 gene, a homozygous T > C substitution at position +2 of the consensus donor splice site of exon 26, was found in the 4 index patients. Clinical evaluation of 9 patients from the 4 families revealed variable disease severity particularly as regards motor achievement and disease progression. Microsatellite analysis showed an identical mutation-associated haplotype in the 4 index cases indicating a founder effect of the mutation in all 4 families. Conclusions: Our data provide further evidence that the clinical spectrum of MDC1A due to a single mutation is heterogeneous, particularly in terms of motor achievement and disease progression, making it difficult to give a reliable prognosis even in patients with identical LAMA2-associated haplotype. The c.3924 + 2 T > C mutation to date has been found only in patients originating from the Middle East or Sudan; therefore laminin 2 chain deficiency in patients from those regions should initially prompt a search for this mutation.

Response to “The Role of Cytomegalovirus in Schizencephaly” by Spalice et al.

Abstract: Response to ‘‘The Role of Cytomegalovirus in Schizencephaly’’ by Spalice et al. Annapurna Poduri,* Cecilia Mellado, and Christopher A. Walsh Department of Neurology, Children’s Hospital Boston, Boston, Massachusetts Division of Genetics, Children’s Hospital Boston, The Manton Center for Orphan Disease Research, Boston, Massachusetts Department of Neurology, Beth Israel Deaconess Medical Center, Boston, Massachusetts Howard Hughes Medical Institute, Chevy Chase, Maryland