Involvement of myocyte enhancer factor 2c in the pathogenesis of autism spectrum disorder.
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ABSTRACT:
SUBMITTER: Chaudhary R
PROVIDER: S-EPMC8082549 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Mef2c haploinsufficiency is implicated in behavioral deficits related to autism, schizophrenia, and intellectual disability. Although perturbations in the cerebellum, notably Purkinje cells, have been linked to these neurological disorders, the underlying mechanisms remain poorly understood. In this study, we investigated the roles of Mef2c in cerebellar Purkinje cells during the first three weeks of postnatal development. Our analysis revealed that in comparison to other members of the Mef2 family, Mef2c expression is limited to postnatal Purkinje cells. Because the role of Mef2c has not been assessed in GABAergic neurons, we set out to determine the functional significance of Mef2c by knocking down the expression of Mef2c selectively in Purkinje cells. We found that the loss of Mef2c expression during the first and second postnatal week results in an increase in dendritic arborization without impact on the general growth and migration of Purkinje cells. The influence of Mef2c on dendritic arborization persists throughout the first three weeks, but is most prominent during the first postnatal week suggesting a critical period of Mef2c activity. Additionally, the loss of Mef2c expression results in an increase in the number of spines accompanied by an increase in Gad67 and vGluT1 puncta and decrease in vGluT2 puncta. Thus, our results reveal the specific expression and functional relevance of Mef2c in developing Purkinje cells and offer insight to how disruption of the expression of Mef2c in a GABAergic neuronal subtype may lead to pathogenesis of cerebellar-associated disorders.
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Project description:Rapid mineralization of cultured osteoblasts could be a useful characteristic in stem cell-mediated therapies for fracture and other orthopedic problems. Dimethyl sulfoxide (DMSO) is a small amphipathic solvent molecule capable of stimulating cell differentiation. We report that, in primary human osteoblasts, DMSO dose-dependently enhanced the expression of osteoblast differentiation markers alkaline phosphatase activity and extracellular matrix mineralization. Furthermore, similar DMSO-mediated mineralization enhancement was observed in primary osteoblast-like cells differentiated from mouse mesenchymal cells derived from fat, a promising source of starter cells for cell-based therapy. Using a convenient mouse pre-osteoblast model cell line MC3T3-E1, we further investigated this phenomenon showing that numerous osteoblast-expressed genes were elevated in response to DMSO treatment and correlated with enhanced mineralization. Myocyte enhancer factor 2c (Mef2c) was identified as the transcription factor most induced by DMSO, among the numerous DMSO-induced genes, suggesting a role for Mef2c in osteoblast gene regulation. Immunohistochemistry confirmed expression of Mef2c in osteoblast-like cells in mouse mandible, cortical, and trabecular bone. shRNAi-mediated Mef2c gene silencing resulted in defective osteoblast differentiation, decreased alkaline phosphatase activity, and matrix mineralization and knockdown of osteoblast specific gene expression, including osteocalcin and bone sialoprotein. A flow on knockdown of bone-specific transcription factors, Runx2 and osterix by shRNAi knockdown of Mef2c, suggests that Mef2c lies upstream of these two important factors in the cascade of gene expression in osteoblasts.
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Project description:Myocyte enhancer factor 2C (MEF2C) is increasingly expressed in mice along with breast cancer brain metastases (BCBM) development. We aim to ascertain MEF2C expression in human BCBM, establish the relationship with disease severity, disclose the involvement of vascular endothelial growth factor receptor-2 (VEGFR-2) and β-catenin, also known as KDR and CTNNB1, respectively, and investigate if matched primary tumors express the protein. We studied resected BCBM for the expression of MEF2C, VEGFR-2, and ß-catenin, as well as proliferation (Ki-67) and epithelial (pan Cytokeratin) markers, and related experimental and clinical data. MEF2C expression was further assessed in matched primary tumors and non-BCBM samples used as controls. MEF2C expression was observed in BCBM, but not in controls, and was categorized into three phenotypes (P): P1, with extranuclear location; P2, with extranuclear and nuclear staining, and P3, with nuclear location. Nuclear translocation increased with metastases extension and Ki-67-positive cells number. P1 was associated with higher VEFGR-2 plasma membrane immunoreactivity, whereas P2 and P3 were accompanied by protein dislocation. P1 was accompanied by β-catenin membrane expression, while P2 and P3 exhibited β-catenin nuclear translocation. Primary BC samples expressed MEF2C in mammary ducts and scattered cells in the parenchyma. MEF2C emerges as a player in BCBM associated with disease severity and VEGFR-2 and β-catenin signaling.
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Project description:Reversible proline-directed phosphorylation at Ser/Thr-Pro motifs has an essential role in myogenesis, a multistep process strictly regulated by several signaling pathways that impinge on two families of myogenic effectors, the basic helix-loop-helix myogenic transcription factors and the MEF2 (myocyte enhancer factor 2) proteins. The question of how these signals are deciphered by the myogenic effectors remains largely unaddressed. In this study, we show that the peptidyl-prolyl isomerase Pin1, which catalyzes the isomerization of phosphorylated Ser/Thr-Pro peptide bonds to induce conformational changes of its target proteins, acts as an inhibitor of muscle differentiation because its knockdown in myoblasts promotes myotube formation. With the aim of clarifying the mechanism of Pin1 function in skeletal myogenesis, we investigated whether MEF2C, a critical regulator of the myogenic program that is the end point of several signaling pathways, might serve as a/the target for the inhibitory effects of Pin1 on muscle differentiation. We show that Pin1 interacts selectively with phosphorylated MEF2C in skeletal muscle cells, both in vitro and in vivo. The interaction with Pin1 requires two novel critical phospho-Ser/Thr-Pro motifs in MEF2C, Ser(98) and Ser(110), which are phosphorylated in vivo. Overexpression of Pin1 decreases MEF2C stability and activity and its ability to cooperate with MyoD to activate myogenic conversion. Collectively, these findings reveal a novel role for Pin1 as a regulator of muscle terminal differentiation and suggest that Pin1-mediated repression of MEF2C function could contribute to this function.
| S-EPMC2966067 | biostudies-literature
Project description:A variety of cardiac transcription factors/cofactors, signaling pathways, and downstream structural genes integrate to form the regulatory hierarchies to ensure proper cardiogenesis in vertebrate. Major interaction proteins of the transcription cofactor vestigial like family member 4 (VGLL4) include myocyte enhancer factor 2 (MEF2) and TEA domain transcription factors (TEAD), both of which play important roles in embryonic cardiac development and in adulthood. In this study, we identified that the deficiency of zebrafish vgll4b paralog, a unique family member expressed in developing heart, led to an impaired valve development. Mechanistically, in vgll4b mutant embryos the disruption of Vgll4b-Mef2c complex, rather than that of Vgll4b-Tead complex, resulted in an aberrant expression of krüppel-like factor 2a (klf2a) in endocardium. Such misexpression of klf2a eventually evoked the valvulogenesis defects. Our findings suggest that zebrafish Vgll4b plays an important role in modulating the transcription activity of Mef2c on klf2a during valve development in a blood-flow-independent manner.
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Project description:N-Methyl-d-aspartate type glutamate receptors (NMDARs) are key mediators of synaptic activity-regulated gene transcription in neurons, both during development and in the adult brain. Developmental differences in the glutamate receptor ionotropic NMDA 2 (GluN2) subunit composition of NMDARs determines whether they activate the transcription factor cAMP-responsive element-binding protein 1 (CREB). However, whether the developmentally regulated GluN3A subunit also modulates NMDAR-induced transcription is unknown. Here, using an array of techniques, including quantitative real-time PCR, immunostaining, reporter gene assays, RNA-Seq, and two-photon glutamate uncaging with calcium imaging, we show that knocking down GluN3A in rat hippocampal neurons promotes the inducible transcription of a subset of NMDAR-sensitive genes. We found that this enhancement is mediated by the accumulation of phosphorylated p38 mitogen-activated protein kinase in the nucleus, which drives the activation of the transcription factor myocyte enhancer factor 2C (MEF2C) and promotes the transcription of a subset of synaptic activity-induced genes, including brain-derived neurotrophic factor (Bdnf) and activity-regulated cytoskeleton-associated protein (Arc). Our evidence that GluN3A regulates MEF2C-dependent transcription reveals a novel mechanism by which NMDAR subunit composition confers specificity to the program of synaptic activity-regulated gene transcription in developing neurons.
| S-EPMC7307204 | biostudies-literature
Project description:OBJECTIVE:Laminar flow activates myocyte enhancer factor 2 (MEF2) transcription factors in vitro to induce expression of atheroprotective genes in the endothelium. Here we sought to establish the role of Mef2c in the vascular endothelium in vivo. APPROACH AND RESULTS:To study endothelial Mef2c, we generated endothelial-specific deletion of Mef2c using Tie2-Cre or Cdh5-Cre-ERT2 and examined aortas and carotid arteries by en face immunofluorescence. We observed enhanced actin stress fiber formation in the Mef2c-deleted thoracic aortic endothelium (laminar flow region), similar to those observed in normal aortic inner curvature (disturbed flow region). Furthermore, Mef2c deletion resulted in the de novo formation of subendothelial intimal cells expressing markers of differentiated smooth muscle in the thoracic aortas and carotids. Lineage tracing showed that these cells were not of endothelial origin. To define early events in intimal development, we induced endothelial deletion of Mef2c and examined aortas at 4 and 12 weeks postinduction. The number of intimal cell clusters increased from 4 to 12 weeks, but the number of cells within a cluster peaked at 2 cells in both cases, suggesting ongoing migration but minimal proliferation. Moreover, we identified cells extending from the media through fenestrations in the internal elastic lamina into the intima, indicating transfenestral smooth muscle migration. Similar transfenestral migration was observed in wild-type carotid arteries ligated to induce neointimal formation. CONCLUSIONS:These results indicate that endothelial Mef2c regulates the endothelial actin cytoskeleton and inhibits smooth muscle cell migration into the intima.
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Project description:Myocyte Enhancer Factor 2 (MEF2) proteins are a small family of transcription factors that play pivotal role in morphogenesis and myogenesis of skeletal, cardiac, and smooth muscle cells. In vertebrates, there are four MEF2 genes, referred to as MEF2A, -B, -C, and -D, that are located on different chromosomes. After birth MEF2A, MEF2B, MEF2D transcriptions are expressed ubiquitously, whereas MEF2C transcripts are restricted to skeletal muscle, brain, and spleen. In this study, on the basis of the sequences of the bovine chromosome 7 genomic contig, available in the GenBank database, sets of PCR primers were designed and to amplify the bovine MEF2C gene promoter region, exon 1 (5'UTR) and part sequence of the intron 1. Seven overlapping fragments of the bovine MEF2C gene were amplified and then sequenced. Altogether, these fragments were composed in the 3,120-bp sequence which was deposited in the GenBank database under accession no. GU211007. The sequence fragment included the putative site of the promoter region and transcription start of the exon 1. The sequence analysis of these fragments in individual animals representing different Bos taurus breeds revealed four variations in promoter region: g.-1606C>T, g.-1336_-1335DelG, g.-818C>T, g.-613_-612DelA and four SNPs within intron 1: g.2711A>G, g. 2913A>G, g.2962G>T and g.3014A>G. No polymorphism was found within sequence of the exon 1 (5'UTR). These polymorphisms were identified for first time using these sequences and were confirmed by RFLP or MSSCP methods.
| S-EPMC3024517 | biostudies-literature
Project description:Cardiomyocyte regeneration is limited in adults. The adipose tissue-derived stromal vascular fraction (Ad-SVF) contains pluripotent stem cells that rarely transdifferentiate into spontaneously beating cardiomyocyte-like cells (beating CMs). However, the characteristics of beating CMs and the factors that regulate the differentiation of Ad-SVF toward the cardiac lineage are unknown. We developed a simple culture protocol under which the adult murine inguinal Ad-SVF reproducibly transdifferentiates into beating CMs without induction. The beating CMs showed the striated ventricular phenotype of cardiomyocytes and synchronised oscillation of the intracellular calcium concentration among cells on day 28 of Ad-SVF primary culture. We also identified beating CM-fated progenitors (CFPs) and performed single-cell transcriptome analysis of these CFPs. Among 491 transcription factors that were differentially expressed (≥ 1.75-fold) in CFPs and the beating CMs, myocyte-specific enhancer 2c (Mef2c) was key. Transduction of Ad-SVF cells with Mef2c using a lentiviral vector yielded CFPs and beating CMs with ~ tenfold higher cardiac troponin T expression, which was abolished by silencing of Mef2c. Thus, we identified the master gene required for transdifferentiation of Ad-SVF into beating CMs. These findings will facilitate the development of novel cardiac regeneration therapies based on gene-modified, cardiac lineage-directed Ad-SVF cells.
| S-EPMC7810870 | biostudies-literature
Project description:The role of microRNA-214-3p (miR-214-3p) in cardiac hypertrophy was not well illustrated. The present study aimed to investigate the expression and potential target of miR-214-3p in angiotensin II (Ang-II)-induced mouse cardiac hypertrophy. In mice with either Ang-II infusion or transverse aortic constriction (TAC) model, miR-214-3p expression was markedly decreased in the hypertrophic myocardium. Down-regulation of miR-214-3p was observed in the myocardium of patients with cardiac hypertrophy. Expression of miR-214-3p was upregulated in Ang-II-induced hypertrophic neonatal mouse ventricular cardiomyocytes. Cardiac hypertrophy was attenuated in Ang-II-infused mice by tail vein injection of miR-214-3p. Moreover, miR-214-3p inhibited the expression of atrial natriuretic peptide (ANP) and β-myosin heavy chain (MHC) in Ang-II-treated mouse cardiomyocytes in vitro. Myocyte-specific enhancer factor 2C (MEF2C), which was increased in Ang-II-induced hypertrophic mouse myocardium and cardiomyocytes, was identified as a target gene of miR-214-3p. Functionally, miR-214-3p mimic, consistent with MEF2C siRNA, inhibited cell size increase and protein expression of ANP and β-MHC in Ang-II-treated mouse cardiomyocytes. The NF-κB signal pathway was verified to mediate Ang-II-induced miR-214-3p expression in cardiomyocytes. Taken together, our results revealed that MEF2C is a novel target of miR-214-3p, and attenuation of miR-214-3p expression may contribute to MEF2Cexpressionin cardiac hypertrophy.
| S-EPMC5087095 | biostudies-literature