An L314Q mutation in Map3k1 gene results in failure of eyelid fusion in the N-ethyl-N-nitrosourea-induced mutant line.
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ABSTRACT:
SUBMITTER: Chen B
PROVIDER: S-EPMC8614015 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BACKGROUND: N-ethyl-N-nitrosourea mutagenesis was used to induce a point mutation in C57BL/6?J mice. Pain-related phenotype screening was performed in 915?G3 mice. We report the detection of a heritable recessive mutant in meiotic recombinant N1F1 mice that caused an abnormal pain sensitivity phenotype with spontaneous skin inflammation in the paws and ears. METHODS: We investigated abnormal sensory processing, neuronal peptides, and behavioral responses after the induction of autoinflammatory disease. Single-nucleotide polymorphism (SNP) markers and polymerase chain reaction product sequencing were used to identify the mutation site. RESULTS: All affected mice developed paw inflammation at 4-8 weeks. Histological examinations revealed hyperplasia of the epidermis in the inflamed paws and increased macrophage expression in the spleen and paw tissues. Mechanical and thermal nociceptive response thresholds were reduced in the affected mice. Locomotor activity was decreased in affected mice with inflamed hindpaws, and this reduction was attributable to the avoidance of contact of the affected paw with the floor. Motor strength and daily activity in the home cage in the affected mice did not show any significant changes. Although Fos immunoreactivity was normal in the dorsal horn of affected mice, calcitonin gene-related peptide immunoreactivity significantly increased in the deep layer of the dorsal horn. The number of microglia increased in the spinal cord, hippocampus, and cerebral cortex in affected mice, and the proliferation of microglia was maintained for a couple of months. Two hundred eighty-five SNP markers were used to reveal the affected gene locus, which was found on the distal part of chromosome 18. A point mutation was detected at A to G in exon 8 of the pstpip2 gene, resulting in a conserved tyrosine residue at amino acid 180 replaced by cysteine (Y180 C). CONCLUSIONS: The data provide definitive evidence that a mutation in pstpip2 causes autoinflammatory disease in an N-ethyl-N-nitrosourea mutagenesis mouse model. Thus, our pstpip2 mutant mice provide a new model for investigating the potential mechanisms of inflammatory pain.
| S-EPMC3414809 | biostudies-literature
Project description:Embryonic eyelid closure involves forward movement and ultimate fusion of the upper and lower eyelids, an essential step of mammalian ocular surface development. Although its underlying mechanism of action is not fully understood, a functional mitogen-activated protein kinase kinase kinase 1 (MAP3K1) is required for eyelid closure. Here we investigate the molecular signatures of MAP3K1 in eyelid morphogenesis. At mouse gestational day E15.5, the developmental stage immediately prior to eyelid closure, MAP3K1 expression is predominant in the eyelid leading edge (LE) and the inner eyelid (IE) epithelium. We used laser capture microdissection (LCM) to obtain highly enriched LE and IE cells from wild type and MAP3K1-deficient fetuses and analyzed genome-wide expression profiles. The gene expression data led to the identification of three distinct developmental features of MAP3K1. First, MAP3K1 modulated Wnt and Sonic hedgehog signals, actin reorganization, and proliferation only in LE but not in IE epithelium, illustrating the temporal-spatial specificity of MAP3K1 in embryogenesis. Second, MAP3K1 potentiated AP-2α expression and SRF and AP-1 activity, but its target genes were enriched for binding motifs of AP-2α and SRF, and not AP-1, suggesting the existence of novel MAP3K1-AP-2α/SRF modules in gene regulation. Third, MAP3K1 displayed variable effects on expression of lineage specific genes in the LE and IE epithelium, revealing potential roles of MAP3K1 in differentiation and lineage specification. Using LCM and expression array, our studies have uncovered novel molecular signatures of MAP3K1 in embryonic eyelid closure.
| S-EPMC3549005 | biostudies-literature
Project description:Epilepsy is one of the common neurological diseases that affects not only adults but also infants and children. Because epilepsy has been studied for a long time, there are several pharmacologically effective anticonvulsants, which, however, are not suitable as therapy for all patients. The genesis of epilepsy has been extensively investigated in terms of its occurrence after injury and as a concomitant disease with various brain diseases, such as tumors, ischemic events, etc. However, in the last decades, there are multiple reports that both genetic and epigenetic factors play an important role in epileptogenesis. Therefore, there is a need for further identification of genes and loci that can be associated with higher susceptibility to epileptic seizures. Use of mouse knockout models of epileptogenesis is very informative, but it has its limitations. One of them is due to the fact that complete deletion of a gene is not, in many cases, similar to human epilepsy-associated syndromes. Another approach to generating mouse models of epilepsy is N-Ethyl-N-nitrosourea (ENU)-directed mutagenesis. Recently, using this approach, we generated a novel mouse strain, soc (socrates, formerly s8-3), with epileptiform activity. Using molecular biology methods, calcium neuroimaging, and immunocytochemistry, we were able to characterize the strain. Neurons isolated from soc mutant brains retain the ability to differentiate in vitro and form a network. However, soc mutant neurons are characterized by increased spontaneous excitation activity. They also demonstrate a high degree of Ca2+ activity compared to WT neurons. Additionally, they show increased expression of NMDA receptors, decreased expression of the Ca2+-conducting GluA2 subunit of AMPA receptors, suppressed expression of phosphoinositol 3-kinase, and BK channels of the cytoplasmic membrane involved in protection against epileptogenesis. During embryonic and postnatal development, the expression of several genes encoding ion channels is downregulated in vivo, as well. Our data indicate that soc mutation causes a disruption of the excitation-inhibition balance in the brain, and it can serve as a mouse model of epilepsy.
| S-EPMC10707124 | biostudies-literature
Project description:Embryonic eyelid closure is a critical step of mammalian ocular surface development involving the forward movement and ultimate fusion of the upper and lower eyelids. Although its underlying mechanism of action is not fully understood, a functional mitogen-activated protein kinase kinase kinase 1 (MAP3K1) is essentially required for eyelid closure. We used microarray to characterize the central role that MAP3K1 plays in embryonic eyelid closure.
2012-09-01 | GSE39240 | GEO
Project description:Hirschsprung disease (HSCR), or colonic aganglionosis, is a congenital disorder characterized by the absence of intramural ganglia along variable lengths of the colon, resulting in intestinal obstruction. It is the most common cause of congenital intestinal obstruction, with an incidence of 1 in 5,000 live births. N-ethyl-N-nitrosourea (ENU)-induced mutagenesis is a powerful tool for the study of gene function and the generation of human disease models. In the current study, a novel mutant mouse with aganglionic megacolon and coat color spotting was generated by ENU-induced mutagenesis. Histological and acetylcholinesterase (AChE) whole-mount staining analysis showed a lack of ganglion cells in the colon in mutant mice. The mutation was mapped to chromosome 14 between markers rs30928624 and D14Mit205 (Chr 14 positions 103723921 bp and 105054651 bp). The Ednrb (Chr 14 position 103814625-103844173 bp) was identified as a potential candidate gene in this location. Mutation analysis revealed a T>C missense mutation at nucleotide 857 of the cDNA encoding endothelin receptor B (EDNRB) in which a proline was substituted for the highly conserved Lys-286 residue (L286P) in the fifth transmembrane (TM V) domain of this G protein-coupled receptor. The mutant mouse was named Ednrb(m1yzcm) (Ednrb; mutation 1, Yangzhou University Comparative Medicine Center). The results of the present study implicate the structural importance of the TM V domain in Ednrb function, and the Ednrb(m1yzcm) mouse represents a valuable model for the study of HSCR in humans.
| S-EPMC4976238 | biostudies-other
Project description:We describe the clinical, genetic, biochemical, and molecular characterization of a mouse that arose in the first generation (G(1)) of a random mutagenesis screen with the chemical mutagen ethyl-nitrosourea. The mouse was observed to have skeletal abnormalities inherited with an X-linked dominant pattern of inheritance. The causative mutation, named Skeletal abnormality 1 (Ska1), was shown to be a single base pair mutation in a splice donor site immediately following exon 8 of the Phex (phosphate-regulating gene with homologies to endopeptidases located on the X-chromosome) gene. This point mutation caused skipping of exon 8 from Phex mRNA, hypophosphatemia, and features of rickets. This experimentally induced phenotype mirrors the human condition X-linked hypophosphatemia; directly confirms the role of Phex in phosphate homeostasis, normal skeletal development, and rickets; and illustrates the power of mutagenesis in exploring animal models of human disease.
| S-EPMC1850771 | biostudies-literature
Project description:Embryonic eyelid closure is a critical step of mammalian ocular surface development involving the forward movement and ultimate fusion of the upper and lower eyelids. Although its underlying mechanism of action is not fully understood, a functional mitogen-activated protein kinase kinase kinase 1 (MAP3K1) is essentially required for eyelid closure. We used microarray to characterize the central role that MAP3K1 plays in embryonic eyelid closure. We used Laser Capture Microdissection (LCM) to obtain highly enriched eyelid leading edge and inner eyelid cells from E15.5 wild type and MAP3K1-deficient fetuses and analyze genome-wide expression profiles.
2012-09-01 | E-GEOD-39240 | biostudies-arrayexpress
Project description:Kyphosis and scoliosis are common spinal disorders that occur as part of complex syndromes or as nonsyndromic, idiopathic diseases. Familial and twin studies implicate genetic involvement, although the causative genes for idiopathic kyphoscoliosis remain to be identified. To facilitate these studies, we investigated progeny of mice treated with the chemical mutagen N-ethyl-N-nitrosourea (ENU) and assessed them for morphological and radiographic abnormalities. This identified a mouse with kyphoscoliosis due to fused lumbar vertebrae, which was inherited as an autosomal dominant trait; the phenotype was designated as hereditary vertebral fusion (HVF) and the locus as Hvf. Micro-computed tomography (?CT) analysis confirmed the occurrence of nonsyndromic kyphoscoliosis due to fusion of lumbar vertebrae in HVF mice, consistent with a pattern of blocked vertebrae due to failure of segmentation. ?CT scans also showed the lumbar vertebral column of HVF mice to have generalized disc narrowing, displacement with compression of the neural spine, and distorted transverse processes. Histology of lumbar vertebrae revealed HVF mice to have irregularly shaped vertebral bodies and displacement of intervertebral discs and ossification centers. Genetic mapping using a panel of single nucleotide polymorphic (SNP) loci arranged in chromosome sets and DNA samples from 23 HVF (eight males and 15 females) mice, localized Hvf to chromosome 4A3 and within a 5-megabase (Mb) region containing nine protein coding genes, two processed transcripts, three microRNAs, five small nuclear RNAs, three large intergenic noncoding RNAs, and 24 pseudogenes. However, genome sequence analysis in this interval did not identify any abnormalities in the coding exons, or exon-intron boundaries of any of these genes. Thus, our studies have established a mouse model for a monogenic form of nonsyndromic kyphoscoliosis due to fusion of lumbar vertebrae, and further identification of the underlying genetic defect will help elucidate the molecular mechanisms involved in kyphoscoliosis. © 2018 The Authors. JBMR Plus is published by Wiley Periodicals, Inc. on behalf of the American Society for Bone and Mineral Research.
| S-EPMC6124210 | biostudies-literature
Project description:Single-cell sequencing has shown that thousands of mutations accumulate with age in most human tissues. While there is ample evidence that some mutations can clonally amplify and lead to disease, the total burden of mutations a cell tolerates without functional decline remains unknown. Here we addressed this question by exposing human primary fibroblasts to multiple, low doses of N-ethyl-N-nitrosourea (ENU) and analyzed somatic mutation burden using single-cell whole genome sequencing. The results indicate that individual cells can sustain ~60,000 single-nucleotide variants (SNVs) with only a slight adverse effect on growth rate. We provide evidence that such high levels of mutations are only tolerated through negative selection against variants in gene coding regions, and in sequences associated with genetic pathways for maintaining basic cellular function and growth. Since most tissues in adults are non-dividing, these results suggest that somatic mutations in the absence of negative selection may have functionally adverse effects.
2024-07-22 | GSE271867 | GEO
Project description:Developmental eyelid closure is an evolutionarily conserved morphogenetic event requiring proliferation, differentiation, cytoskeleton reorganization, and migration of epithelial cells at the tip of the developing eyelid. Many signaling events take place during eyelid closure, but how the signals converge to regulate the morphogenetic process remains an open and intriguing question. Here we show that mitogen-activated protein kinase kinase kinase 1 (MAP3K1) highly expressed in the developing eyelid epithelium, forms with c-Jun, a regulatory axis that orchestrates morphogenesis by integrating two different networks of eyelid closure signals. A TGF-?/EGFR-RhoA module initiates one of these networks by inducing c-Jun expression which, in a phosphorylation-independent manner, binds to the Map3k1 promoter and causes an increase in MAP3K1 expression. RhoA knockout in the ocular surface epithelium disturbs this network by decreasing MAP3K1 expression, and causes delayed eyelid closure in Map3k1 hemizygotes. The second network is initiated by the enzymatic activity of MAP3K1, which phosphorylates and activates a JNK-c-Jun module, leading to AP-1 transactivation and induction of its downstream genes, such as Pai-1. MAP3K1 inactivation reduces AP-1 activity and PAI-1 expression both in cells and developing eyelids. MAP3K1 is therefore the nexus of an intracrine regulatory loop connecting the TGF-?/EGFR/RhoA-c-Jun and JNK-c-Jun-AP-1 pathways in developmental eyelid closure.
| S-EPMC3198332 | biostudies-literature