Cell autonomous and non-cell autonomous functions of Otx2 in patterning the rostral brain

Development ◽  
1999 ◽  
Vol 126 (19) ◽  
pp. 4295-4304 ◽  
Author(s):  
M. Rhinn ◽  
A. Dierich ◽  
M. Le Meur ◽  
S. Ang
Keyword(s):  
Cell Adhesion ◽  
Target Genes ◽  
Homeobox Gene ◽  
Visceral Endoderm ◽  
Wild Type ◽  
Candidate Target ◽  
Important Regulator ◽  
Mutant Cells ◽  
The Brain ◽  
Brain Patterning

Previous studies have shown that the homeobox gene Otx2 is required first in the visceral endoderm for induction of forebrain and midbrain, and subsequently in the neurectoderm for its regional specification. Here, we demonstrate that Otx2 functions both cell autonomously and non-cell autonomously in neurectoderm cells of the forebrain and midbrain to regulate expression of region-specific homeobox and cell adhesion genes. Using chimeras containing both Otx2 mutant and wild-type cells in the brain, we observe a reduction or loss of expression of Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2 in mutant cells, whereas expression of En2 and Six3 is rescued by surrounding wild-type cells. Forebrain Otx2 mutant cells subsequently undergo apoptosis. Altogether, this study demonstrates that Otx2 is an important regulator of brain patterning and morphogenesis, through its regulation of candidate target genes such as Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2.

scholarly journals Ins1-Cre and Ins1-CreER Gene Replacement Alleles Are Susceptible To Silencing By DNA Hypermethylation

Endocrinology ◽  
2020 ◽  
Vol 161 (8) ◽  
Author(s):  
Elham Mosleh ◽  
Kristy Ou ◽  
Matthew W Haemmerle ◽  
Teguru Tembo ◽  
Andrew Yuhas ◽  
...  
Keyword(s):  
Beta Cell ◽  
Endocrine Cells ◽  
Target Genes ◽  
Cell Function ◽  
Gene Replacement ◽  
Wild Type ◽  
Dna Hypermethylation ◽  
Gene Ablation ◽  
Low Levels ◽  
The Brain

Abstract Targeted gene ablation studies of the endocrine pancreas have long suffered from suboptimal Cre deleter strains. In many cases, Cre lines purportedly specific for beta cells also displayed expression in other islet endocrine cells or in a subset of neurons in the brain. Several pancreas and endocrine Cre lines have experienced silencing or mosaicism over time. In addition, many Cre transgenic constructs were designed to include the hGH mini-gene, which by itself increases beta-cell replication and decreases beta-cell function. More recently, driver lines with Cre or CreER inserted into the Ins1 locus were generated, with the intent of producing β cell-specific Cre lines with faithful recapitulation of insulin expression. These lines were bred in multiple labs to several different mouse lines harboring various lox alleles. In our hands, the ability of the Ins1-Cre and Ins1-CreER lines to delete target genes varied from that originally reported, with both alleles displaying low levels of expression, increased levels of methylation compared to the wild-type allele, and ultimately inefficient or absent target deletion. Thus, caution is warranted in the interpretation of results obtained with these genetic tools, and Cre expression and activity should be monitored regularly when using these lines.

scholarly journals Barrier Activity in Candida albicans Mediates Pheromone Degradation and Promotes Mating

2007 ◽  
Vol 6 (6) ◽  
pp. 907-918 ◽  
Author(s):  
Dana Schaefer ◽  
Pierre Côte ◽  
Malcolm Whiteway ◽  
Richard J. Bennett
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Cell Growth ◽  
Cell Fusion ◽  
Aspartyl Protease ◽  
Wild Type ◽  
Pheromone Activity ◽  
Important Regulator ◽  
Mutant Cells ◽  
Type Strains

ABSTRACT Mating in Candida albicans and Saccharomyces cerevisiae is regulated by the secretion of peptide pheromones that initiate the mating process. An important regulator of pheromone activity in S. cerevisiae is barrier activity, involving an extracellular aspartyl protease encoded by the BAR1 gene that degrades the alpha pheromone. We have characterized an equivalent barrier activity in C. albicans and demonstrate that the loss of C. albicans BAR1 activity results in opaque a cells exhibiting hypersensitivity to alpha pheromone. Hypersensitivity to pheromone is clearly seen in halo assays; in response to alpha pheromone, a lawn of C. albicans Δbar1 mutant cells produces a marked zone in which cell growth is inhibited, whereas wild-type strains fail to show halo formation. C. albicans mutants lacking BAR1 also exhibit a striking mating defect in a cells, but not in α cells, due to overstimulation of the response to alpha pheromone. The block to mating occurs prior to cell fusion, as very few mating zygotes were observed in mixes of Δbar1 a and α cells. Finally, in a barrier assay using a highly pheromone-sensitive strain, we were able to demonstrate that barrier activity in C. albicans is dependent on Bar1p. These studies reveal that a barrier activity to alpha pheromone exists in C. albicans and that the activity is analogous to that caused by Bar1p in S. cerevisiae.

scholarly journals Tao controls epithelial morphogenesis by promoting Fasciclin 2 endocytosis

2012 ◽  
Vol 199 (7) ◽  
pp. 1131-1143 ◽  
Author(s):  
Juan Manuel Gomez ◽  
Ying Wang ◽  
Veit Riechmann
Keyword(s):  
Cell Adhesion ◽  
Adhesion Molecule ◽  
Cell Adhesion Molecule ◽  
Neural Cell Adhesion Molecule ◽  
Squamous Epithelium ◽  
Wild Type ◽  
Lateral Membrane ◽  
Membrane Extension ◽  
Neural Cell Adhesion ◽  
Mutant Cells

Regulation of epithelial cell shape, for example, changes in relative sizes of apical, basal, and lateral membranes, is a key mechanism driving morphogenesis. However, it is unclear how epithelial cells control the size of their membranes. In the epithelium of the Drosophila melanogaster ovary, cuboidal precursor cells transform into a squamous epithelium through a process that involves lateral membrane shortening coupled to apical membrane extension. In this paper, we report a mutation in the gene Tao, which resulted in the loss of this cuboidal to squamous transition. We show that the inability of Tao mutant cells to shorten their membranes was caused by the accumulation of the cell adhesion molecule Fasciclin 2, the Drosophila N-CAM (neural cell adhesion molecule) homologue. Fasciclin 2 accumulation at the lateral membrane of Tao mutant cells prevented membrane shrinking and thereby inhibited morphogenesis. In wild-type cells, Tao initiated morphogenesis by promoting Fasciclin 2 endocytosis at the lateral membrane. Thus, we identify here a mechanism controlling the morphogenesis of a squamous epithelium.

scholarly journals Hepatocyte nuclear factor-1β regulates Wnt signaling through genome-wide competition with β-catenin/lymphoid enhancer binding factor

2019 ◽  
Vol 116 (48) ◽  
pp. 24133-24142 ◽  
Author(s):  
Siu Chiu Chan ◽  
Ying Zhang ◽  
Marco Pontoglio ◽  
Peter Igarashi
Keyword(s):  
Transcription Factor ◽  
Wnt Signaling ◽  
Target Genes ◽  
Cystic Kidney ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
Wild Type ◽  
Genome Wide ◽  
Binding Factor ◽  
Mutant Cells

Hepatocyte nuclear factor-1β (HNF-1β) is a tissue-specific transcription factor that is essential for normal kidney development and renal tubular function. Mutations of HNF-1β produce cystic kidney disease, a phenotype associated with deregulation of canonical (β-catenin–dependent) Wnt signaling. Here, we show that ablation of HNF-1β in mIMCD3 renal epithelial cells produces hyperresponsiveness to Wnt ligands and increases expression of Wnt target genes, including Axin2, Ccdc80, and Rnf43. Levels of β-catenin and expression of Wnt target genes are also increased in HNF-1β mutant mouse kidneys. Genome-wide chromatin immunoprecipitation sequencing (ChIP-seq) in wild-type and mutant cells showed that ablation of HNF-1β increases by 6-fold the number of sites on chromatin that are occupied by β-catenin. Remarkably, 50% of the sites that are occupied by β-catenin in HNF-1β mutant cells colocalize with HNF-1β–occupied sites in wild-type cells, indicating widespread reciprocal binding. We found that the Wnt target genes Ccdc80 and Rnf43 contain a composite DNA element comprising a β-catenin/lymphoid enhancer binding factor (LEF) site overlapping with an HNF-1β half-site. HNF-1β and β-catenin/LEF compete for binding to this element, and thereby HNF-1β inhibits β-catenin–dependent transcription. Collectively, these studies reveal a mechanism whereby a transcription factor constrains canonical Wnt signaling through direct inhibition of β-catenin/LEF chromatin binding.

scholarly journals An Integrated Statistical Approach to Compare Transcriptomics Data across Experiments: A Case Study on the Identification of Candidate Target Genes of the Transcription Factor PPARα

2012 ◽  
Vol 6 ◽  
pp. BBI.S9529
Author(s):  
Mohammad Ohid Ullah ◽  
Michael Müller ◽  
Guido J.E.J. Hooiveld
Keyword(s):  
Transcription Factor ◽  
Multiple Testing ◽  
Target Genes ◽  
Gene Knockout ◽  
Statistical Tests ◽  
Multiple Hypothesis Testing ◽  
Wild Type ◽  
Anova Model ◽  
Candidate Target ◽  
Transcriptomics Data

An effective strategy to elucidate the signal transduction cascades activated by a transcription factor is to compare the transcriptional profiles of wild type and transcription factor knockout models. Many statistical tests have been proposed for analyzing gene expression data, but most tests are based on pair-wise comparisons. Since the analysis of microarrays involves the testing of multiple hypotheses within one study, it is generally accepted that one should control for false positives by the false discovery rate (FDR). However, it has been reported that this may be an inappropriate metric for comparing data across different experiments. Here we propose an approach that addresses the above mentioned problem by the simultaneous testing and integration of the three hypotheses (contrasts) using the cell means ANOVA model. These three contrasts test for the effect of a treatment in wild type, gene knockout, and globally over all experimental groups. We illustrate our approach on microarray experiments that focused on the identification of candidate target genes and biological processes governed by the fatty acid sensing transcription factor PPARα in liver. Compared to the often applied FDR based across experiment comparison, our approach identified a conservative but less noisy set of candidate genes with same sensitivity and specificity. However, our method had the advantage of properly adjusting for multiple testing while integrating data from two experiments, and was driven by biological inference. Taken together, in this study we present a simple, yet efficient strategy to compare differential expression of genes across experiments while controlling for multiple hypothesis testing.

scholarly journals Astrocytic modulation of excitatory synaptic signaling in a mouse model of Rett syndrome

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Benjamin Rakela ◽  
Paul Brehm ◽  
Gail Mandel
Keyword(s):  
Mouse Model ◽  
Rett Syndrome ◽  
Synaptic Activity ◽  
Wild Type ◽  
Synaptic Signaling ◽  
Network Function ◽  
Mosaic Distribution ◽  
Mutant Cells ◽  
The Brain ◽  
Expression Status

Studies linking mutations in Methyl CpG Binding Protein 2 (MeCP2) to physiological defects in the neurological disease, Rett syndrome, have focused largely upon neuronal dysfunction despite MeCP2 ubiquitous expression. Here we explore roles for astrocytes in neuronal network function using cortical slice recordings. We find that astrocyte stimulation in wild-type mice increases excitatory synaptic activity that is absent in male mice lacking MeCP2 globally. To determine the cellular basis of the defect, we exploit a female mouse model for Rett syndrome that expresses wild-type MeCP2-GFP in a mosaic distribution throughout the brain, allowing us to test all combinations of wild-type and mutant cells. We find that the defect is dependent upon MeCP2 expression status in the astrocytes and not in the neurons. Our findings highlight a new role for astrocytes in regulation of excitatory synaptic signaling and in the neurological defects associated with Rett syndrome.

scholarly journals The homeobox gene Hex is required in definitive endodermal tissues for normal forebrain, liver and thyroid formation

Development ◽  
2000 ◽  
Vol 127 (11) ◽  
pp. 2433-2445 ◽  
Author(s):  
J.P. Martinez Barbera ◽  
M. Clements ◽  
P. Thomas ◽  
T. Rodriguez ◽  
D. Meloy ◽  
...  
Keyword(s):  
Normal Development ◽  
Embryonic Stem ◽  
Homeobox Gene ◽  
Definitive Endoderm ◽  
Visceral Endoderm ◽  
Wild Type ◽  
Zona Limitans Intrathalamica ◽  
Ventral Thalamus ◽  
Septum Transversum ◽  
Early Mouse

The homeobox gene Hex is expressed in the anterior visceral endoderm (AVE) and rostral definitive endoderm of early mouse embryos. Later, Hex transcripts are detected in liver, thyroid and endothelial precursor cells. A null mutation was introduced into the Hex locus by homologous recombination in embryonic stem cells. Hex mutant embryos exhibit varying degrees of anterior truncation as well as liver and thyroid dysplasia. The liver diverticulum is formed but migration of hepatocytes into the septum transversum fails to occur. Development of the thyroid is arrested at the thyroid bud stage at 9.5 dpc. Brain defects are restricted to the rostral forebrain and have a caudal limit at the zona limitans intrathalamica, the boundary between dorsal and ventral thalamus. Analysis of Hex(−/−) mutants at early stages shows that the prospective forebrain ectoderm is correctly induced and patterned at 7.5 days post coitum (dpc), but subsequently fails to develop. AVE markers are expressed and correctly positioned but development of rostral definitive endoderm is greatly disturbed in Hex(−/−) embryos. Chimeric embryos composed of Hex(−/−) cells developing within a wild-type visceral endoderm show forebrain defects indicating that Hex is required in the definitive endoderm. All together, these results demonstrate that Hex function is essential in definitive endoderm for normal development of the forebrain, liver and thyroid gland.

Lim1 is required in both primitive streak-derived tissues and visceral endoderm for head formation in the mouse

Development ◽  
1999 ◽  
Vol 126 (22) ◽  
pp. 4925-4932 ◽  
Author(s):  
W. Shawlot ◽  
M. Wakamiya ◽  
K.M. Kwan ◽  
A. Kania ◽  
T.M. Jessell ◽  
...  
Keyword(s):  
Marker Gene ◽  
Embryonic Stem ◽  
Homeobox Gene ◽  
Primitive Streak ◽  
Mouse Embryos ◽  
Chimeric Mouse ◽  
Visceral Endoderm ◽  
Wild Type ◽  
Targeted Mutation ◽  
Head Formation

Lim1 is a homeobox gene expressed in the extraembryonic anterior visceral endoderm and in primitive streak-derived tissues of early mouse embryos. Mice homozygous for a targeted mutation of Lim1 lack head structures anterior to rhombomere 3 in the hindbrain. To determine in which tissues Lim1 is required for head formation and its mode of action, we have generated chimeric mouse embryos and performed tissue layer recombination explant assays. In chimeric embryos in which the visceral endoderm was composed of predominantly wild-type cells, we found that Lim1(−)(/)(−) cells were able to contribute to the anterior mesendoderm of embryonic day 7.5 chimeric embryos but that embryonic day 9.5 chimeric embryos displayed a range of head defects. In addition, early somite stage chimeras generated by injecting Lim1(−)(/)(−) embryonic stem cells into wild-type tetraploid blastocysts lacked forebrain and midbrain neural tissue. Furthermore, in explant recombination assays, anterior mesendoderm from Lim1(−)(/)(−) embryos was unable to maintain the expression of the anterior neural marker gene Otx2 in wild-type ectoderm. In complementary experiments, embryonic day 9.5 chimeric embryos in which the visceral endoderm was composed of predominantly Lim1(−)(/)(−) cells and the embryo proper of largely wild-type cells, also phenocopied the Lim1(−)(/)(−) headless phenotype. These results indicate that Lim1 is required in both primitive streak-derived tissues and visceral endoderm for head formation and that its inactivation in these tissues produces cell non-autonomous defects. We discuss a double assurance model in which Lim1 regulates sequential signaling events required for head formation in the mouse.

beta-thymosin is required for axonal tract formation in developing zebrafish brain

Development ◽  
1999 ◽  
Vol 126 (7) ◽  
pp. 1365-1374
Author(s):  
L.W. Roth ◽  
P. Bormann ◽  
A. Bonnet ◽  
E. Reinhard
Keyword(s):  
Antisense Rna ◽  
Zebrafish Embryos ◽  
Primary Neurons ◽  
Wild Type ◽  
Strong Reduction ◽  
Zebrafish Brain ◽  
Trigeminal Neurons ◽  
Somite Formation ◽  
Important Regulator ◽  
The Brain

beta-Thymosins are polypeptides that bind monomeric actin and thereby function as actin buffers in many cells. We show that during zebrafish development, β-thymosin expression is tightly correlated with neuronal growth and differentiation. It is transiently expressed in a subset of axon-extending neurons, essentially primary neurons that extend long axons, glia and muscle. Non-neuronal expression in the brain is restricted to a subset of glia surrounding newly forming axonal tracts. Skeletal muscle cells in somites, jaw and fin express beta-thymosin during differentiation, coinciding with the time of innervation. Injection of beta-thymosin antisense RNA into zebrafish embryos results in brain defects and impairment of the development of beta-thymosin-associated axon tracts. Furthermore, irregularities in somite formation can be seen in a subset of embryos. Compared to wild-type, antisense-injected embryos show slightly weaker and more diffuse engrailed staining at the midbrain-hindbrain boundary and a strong reduction of Isl-1 labeling in Rohon Beard and trigeminal neurons. The decreased expression is not based on a loss of neurons indicating that beta-thymosin may be involved in the maintenance of the expression of molecules necessary for neuronal differentiation. Taken together, our results strongly indicate that beta-thymosin is an important regulator of development.

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