Growth-Promoting Interaction of IGF-II with the Insulin Receptor during Mouse Embryonic Development

AbstractGenetic analyses of dwarfing phenotypes resulting from targeted mutagenesis of the genes encoding the insulin-like growth factors (IGF-I and IGF-II) and their cognate type 1 IGF receptor (IGF1R) have demonstrated that this signaling system is a major determinant of mouse embryonic growth. Of the two IGF ligands, IGF-I interacts exclusively with IGF1R, whereas IGF-II recognizes an additional receptor (XR), because the growth retardation of embryos lacking both IGR1R and IGF-II (30% of normal birthweight) is more severe than that manifested in either class of singleIgf1rorIgf2null mutants (45 and 60% of normal, respectively). To determine whether XR is the insulin receptor (IR), we examined embryos nullizygous for bothIgf1randInsr.While the growth of embryos lacking solely IR is affected very mildly and only at the end of gestation, concomitant absence of IGF1R results in a severe growth-deficiency phenotype (30% of normal size at birth) that is first detected at Embryonic Day 13.5 and is also characterized by transient edema, curly tail, generalized organ hypoplasia, including the muscles, developmental delays in ossification, and thin epidermis. TheIgf1r/Insrdouble nullizygotes are phenotypically indistinguishable from double mutants lacking IGF1R and IGF-II and from other double and triple mutants in which all of the IGF ligand/receptor interactions have been eliminated. Therefore, these results provide genetic evidence that the growth-promoting function of IGF-II during mouse embryogenesis is mediated in part by signaling through the insulin receptor

Molecular Cytogenetic Analysis and Resequencing of Contactin Associated Protein-Like 2 in Autism Spectrum Disorders

Autism spectrum disorders (ASD) are a group of related neurodevelopmental syndromes with complex genetic etiology.1 We identified a de novo chromosome 7q inversion disrupting Autism susceptibility candidate 2 (AUTS2) and Contactin Associated Protein-Like 2 (CNTNAP2) in a child with cognitive and social delay. We focused our initial analysis on CNTNAP2 based on our demonstration of disruption of Contactin 4 (CNTN4) in a patient with ASD;2 the recent finding of rare homozygous mutations in CNTNAP2 leading to intractable seizures and autism;3 and in situ and biochemical analyses reported herein that confirm expression in relevant brain regions and demonstrate the presence of CNTNAP2 in the synaptic plasma membrane fraction of rat forebrain lysates. We comprehensively resequenced CNTNAP2 in 635 patients and 942 controls. Among patients, we identified a total of 27 nonsynonymous changes; 13 were rare and unique to patients and 8 of these were predicted to be deleterious by bioinformatic approaches and/or altered residues conserved across all species. One variant at a highly conserved position, I869T, was inherited by four affected children in three unrelated families, but was not found in 4010 control chromosomes (p = 0.014). Overall, this resequencing data demonstrated a modest nonsignificant increase in the burden of rare variants in cases versus controls. Nonethless, when viewed in light of two independent studies published in this issue of AJHG showing a relationship between ASD and common CNTNAP2 alleles,4,5 the cytogenetic and mutation screening data suggest that rare variants may also contribute to the pathophysiology of ASD, but place limits on the magnitude of this contribution

Notch Lineages and Activity in Intestinal Stem Cells Determined by a New Set of Knock-In Mice

The conserved role of Notch signaling in controlling intestinal cell fate specification and homeostasis has been extensively studied. Nevertheless, the precise identity of the cells in which Notch signaling is active and the role of different Notch receptor paralogues in the intestine remain ambiguous, due to the lack of reliable tools to investigate Notch expression and function in vivo. We generated a new series of transgenic mice that allowed us, by lineage analysis, to formally prove that Notch1 and Notch2 are specifically expressed in crypt stem cells. In addition, a novel Notch reporter mouse, Hes1-EmGFPSAT, demonstrated exclusive Notch activity in crypt stem cells and absorptive progenitors. This roster of knock-in and reporter mice represents a valuable resource to functionally explore the Notch pathway in vivo in virtually all tissues

Id4 Downstream of Notch2 Maintains Neural Stem Cell Quiescence in the Adult Hippocampus

Neural stem cells (NSCs) in the adult mouse hippocampal dentate gyrus (DG) are mostly quiescent, and only a few are in cell cycle at any point in time. DG NSCs become increasingly dormant with age and enter mitosis less frequently, which impinges on neurogenesis. How NSC inactivity is maintained is largely unknown. Here, we found that Id4 is a downstream target of Notch2 signaling and maintains DG NSC quiescence by blocking cell-cycle entry. Id4 expression is sufficient to promote DG NSC quiescence and Id4 knockdown rescues Notch2-induced inhibition of NSC proliferation. Id4 deletion activates NSC proliferation in the DG without evoking neuron generation, and overexpression increases NSC maintenance while promoting astrogliogenesis at the expense of neurogenesis. Together, our findings indicate that Id4 is a major effector of Notch2 signaling in NSCs and a Notch2-Id4 axis promotes NSC quiescence in the adult DG, uncoupling NSC activation from neuronal differentiation.ISSN:2666-3864ISSN:2211-124

Functional Synergy between Cholecystokinin Receptors CCKAR and CCKBR in Mammalian Brain Development

<div><p>Cholecystokinin (CCK), a peptide hormone and one of the most abundant neuropeptides in vertebrate brain, mediates its actions via two G-protein coupled receptors, CCKAR and CCKBR, respectively active in peripheral organs and the central nervous system. Here, we demonstrate that the CCK receptors have a dynamic and largely reciprocal expression in embryonic and postnatal brain. Using compound homozygous mutant mice lacking the activity of both CCK receptors, we uncover their additive, functionally synergistic effects in brain development and demonstrate that CCK receptor loss leads to abnormalities of cortical development, including defects in the formation of the midline and corpus callosum, and cortical interneuron migration. Using comparative transcriptome analysis of embryonic neocortex, we define the molecular mechanisms underlying these defects. Thus we demonstrate a developmental, hitherto unappreciated, role of the two CCK receptors in mammalian neocortical development.</p></div

Defects in tangentially migrating interneurons in <i>Cckar/br</i> mutants.

<p>(A,B,E,F) Interneuron progenitors are generated normally in <i>Cckar/br</i> mutant embryos compared to controls, as demonstrated by in situ hybridization with <i>Gad1</i> (A,B) and <i>Lhx6</i> (E,F). However their tangential migration appears delayed in the mutants. Arrows in (A,B) indicate the onset of tangential routes employed by migrating interneurons. Note a delay/reduction in tangential migration in <i>Cckar/br</i> mutants. Red marks in (E,F) indicate the extent of tangential migration (deep route) in the neocortex. Scale bar (B,D), 200 μm. (C,D,G,H) In situ hybridization with <i>Gad1</i> and <i>Lhx6</i> at birth (P0, C,D) or late embryogenesis (E17.5, G,H) demonstrates that despite the delay in migration, a majority of interneurons settle into the neocortex. Scale bar (F,H), 200 μm.</p

Defects in cortical development in <i>Cckar/br</i> mutant mice.

<p>(A and B) Nissl staining of coronal sections of control and <i>Cckar/br</i> mutant brains at postnatal day 14 (P14) reveals thickening of the cingulate cortex, agenesis of the corpus callosum, and lateral displacement of the hippocampus. (C and D) In situ hybridization with <i>claudin 11</i>, which is expressed in myelinated oligodentrocytes thus highlighting white matter tracts, shows that the commissural axons of the corpus callosum do not cross to the contralateral hemisphere in <i>Cckar/br</i> mutants (D) compared to control (C). Of a total of 16 brains from <i>Cckar/br</i> mutants that were fully sectioned and analyzed for callosal development, 13 had complete and 3 had partial agenesis of the corpus callosum. Scale bar, 100 μm. (E and F) Labeling of callosal projections with DiI shows that colossal axons fail to cross the midline in <i>Cckar/br</i> double mutants (F). Green lines (in E,F) and asterisk (in F) indicate the midline and Probst bundles, respectively. Scale bar, 200 μm.</p