Binding of serum response factor to cystic fibrosis transmembrane conductance regulator CArG-like elements, as a new potential CFTR transcriptional regulation pathway

René, Céline

doi:10.1093/nar/gki837

Cited by 23 publications

(38 citation statements)

References 87 publications

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“…The last result agrees with a report showing reduced expression of Kcnmb1 in the embryonic heart of Srf knock-out mice (40). Although previous reports have documented functional CArG boxes in the promoters to calcium (18) and chloride (64) transporters, the present results are the first to demonstrate functional CArG elements in an SMC-restricted ion channel subunit. Whether the CArG elements controlling KCNMB1 gene expression functionally regulate KCNIP1, whose first intron harbors KCNMB1, is presently unknown.…”

Section: Discussionsupporting

confidence: 93%

The Smooth Muscle Cell-restricted KCNMB1 Ion Channel Subunit Is a Direct Transcriptional Target of Serum Response Factor and Myocardin

Long

Tharp

Georger

et al. 2009

Journal of Biological Chemistry

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Large conductance calcium-activated potassium (MaxiK) channels play a pivotal role in maintaining normal arterial tone by regulating the excitation-contraction coupling process. MaxiK channels comprise ␣ and ␤ subunits encoded by Kcnma and the cell-restricted Kcnmb genes, respectively. Although the functionality of MaxiK channel subunits has been well studied, the molecular regulation of their transcription and modulation in smooth muscle cells (SMCs) is incomplete. Using several model systems, we demonstrate down-regulation of Kcnmb1 mRNA upon SMC phenotypic modulation in vitro and in vivo. As part of a broad effort to define all functional CArG elements in the genome (i.e. the CArGome), we discovered two conserved CArG boxes located in the proximal promoter and first intron of the human KCNMB1 gene. Gel shift and chromatin immunoprecipitation assays confirmed serum response factor (SRF) binding to both CArG elements. A luciferase assay showed myocardin (MYOCD)-mediated transactivation of the KCNMB1 promoter in a CArG element-dependent manner. In vivo analysis of the human KCNMB1 promoter disclosed activity in embryonic heart and aortic SMCs; mutation of both conserved CArG elements completely abolished in vivo promoter activity. Forced expression of MYOCD increased Kcnmb1 expression in a variety of rodent and human non-SMC lines with no effect on expression of the Kcnma1 subunit. Conversely, knockdown of Srf resulted in decreases of endogenous Kcnmb1. Functional studies demonstrated MYOCD-induced, iberiotoxin-sensitive potassium currents in porcine coronary SMCs. These results reveal the first ion channel subunit as a direct target of SRF-MYOCD transactivation, providing further insight into the role of MYOCD as a master regulator of the SMC contractile phenotype. Smooth muscle cells (SMCs)2 are of crucial importance in maintaining normal structural and contractile integrity of various organs and tissues throughout the vertebrate body. Unlike skeletal and cardiac muscle cells, which largely undergo irreversible terminal differentiation, SMCs have the inherent ability to alter their differentiated state in response to diverse stimulatory cues. This process of SMC phenotypic modulation is a hallmark of several human pathological conditions, including vascular occlusive disease, asthma, intestinal and bladder obstruction, and Alzheimer disease. SMC phenotypic modulation is often defined in terms of unique molecular signatures of gene expression involved with contraction and cytoskeletal architecture as well as extracellular matrix remodeling (1, 2). For example, vascular SMCs display reduced levels of contractile filaments following injury to the vessel wall, adopting the so-called synthetic phenotype characterized by an overabundance of rough endoplasmic reticulum (3). On the other hand, recent studies have shown an exaggerated SMC contractile phenotype thought to contribute to disease progression (4, 5).The majority of SMC-restricted genes contain one or more conserved CArG elements that bind the widely express...

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Section: Discussionsupporting

confidence: 93%

The Smooth Muscle Cell-restricted KCNMB1 Ion Channel Subunit Is a Direct Transcriptional Target of Serum Response Factor and Myocardin

Long

Tharp

Georger

et al. 2009

Journal of Biological Chemistry

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“…Electromobility shift assay Electromobility shift assay (EMSA) was performed as previously described [17].…”

Section: Quantitative Chromatin Immunoprecipitation Assaysmentioning

confidence: 99%

“…As FOXA and C/EBP factors do not act alone, we re-examined the in silico analysis data by using a lower cut-off in order to select other transcription factors that may contribute to CFTR regulation. Possible candidates of this transcription factor network included RREB-1 (the transcription factor with the highest score), several transcription factors with a known effect on CFTR gene expression, such as USF2, SRF and YY1 [17,18,35], and transcription factors involved in lung morphogenesis, but with a lower score than C/ EBPs and FOXAs (SOX17, FOXF1 and NKX2.1) [40]. We then used reporter assays to investigate whether these regulatory elements could participate in the temporal regulation of CFTR expression.…”

Section: Transcription Factors Involved In the Regulation Of Cftr Temmentioning

confidence: 99%

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Transcription factors and miRNAs that regulate fetal to adultCFTRexpression change are new targets for cystic fibrosis

Viart¹,

Bergougnoux²,

Bonini³

et al. 2014

Eur Respir J

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The CFTR gene displays a tightly regulated tissue-specific and temporal expression. Mutations in this gene cause cystic fibrosis (CF). In this study we wanted to identify trans-regulatory elements responsible for CFTR differential expression in fetal and adult lung, and to determine the importance of inhibitory motifs in the CFTR-3′UTR with the aim of developing new tools for the correction of disease-causing mutations within CFTR.We show that lung development-specific transcription factors (FOXA, C/EBP) and microRNAs (miR-101, miR-145, miR-384) regulate the switch from strong fetal to very low CFTR expression after birth. By using miRNome profiling and gene reporter assays, we found that miR-101 and miR-145 are specifically upregulated in adult lung and that miR-101 directly acts on its cognate site in the CFTR-3′UTR in combination with an overlapping AU-rich element. We then designed miRNA-binding blocker oligonucleotides (MBBOs) to prevent binding of several miRNAs to the CFTR-3′UTR and tested them in primary human nasal epithelial cells from healthy individuals and CF patients carrying the p.Phe508del CFTR mutation. These MBBOs rescued CFTR channel activity by increasing CFTR mRNA and protein levels.Our data offer new understanding of the control of the CFTR gene regulation and new putative correctors for cystic fibrosis. @ERSpublications Transcription factors/miRNAs that regulate fetal to adult CFTR expression change are new targets for CF treatment

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“…Total cellular proteins were harvested from the cultured fibroblasts as described elsewhere. 9 For mutation screening in the proband, all the coding sequences of the COL3A1 gene (cDNA) were sequenced bidirectionally in 10 overlapping fragments. For mutation confirmation in the proband and her parents, sequencing and denaturing high-performance liquid chromatography were carried out in genomic DNA.…”

Section: Mutation Analysismentioning

confidence: 99%

Homozygosity for a null allele of COL3A1 results in recessive Ehlers–Danlos syndrome

et al. 2009

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So far, mutations in the human COL3A1 gene have been associated with the predominantly inherited Ehlers-Danlos syndrome (EDS), vascular type. Genotype -phenotype correlation perspectives collapsed, as haploinsufficiency, which was long suggested to confer a milder or unrecognized phenotype, was reported in four patients with a phenotype similar to that of vascular EDS. Here, we study a case of recessive EDS in a young consanguineous girl of healthy parents. She fulfilled the vascular EDS criteria for laboratory testing. Total sequencing of COL3A1 cDNA identified a homozygous nucleotide duplication (c.479dupT) resulting in a premature termination codon (p.Lys161GlnfsX45). Studies in genomic DNA showed that this mutation was inherited from each parent. The expression analysis (RT-PCR, quantitative-PCR, immunohistochemistry, WB) showed strong mRNA decay and an absence of type III collagen in the proband. The expected COL3A1 haploinsufficiency in her healthy ascendants did not lead to the manifestations of vascular EDS. This case provides evidence of a stochastic effect of COL3A1 haploinsufficiency in humans, which could be explained by the relation between nonsense-mediated mRNA decay efficiency and the resulting dominant-negative effect depending on the position of the mutation and/or modifying factors. It opens up new perspectives for the understanding of COL3A1 genotype -phenotype correlations, which is required while considering targeted therapy.

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Binding of serum response factor to cystic fibrosis transmembrane conductance regulator CArG-like elements, as a new potential CFTR transcriptional regulation pathway

Cited by 23 publications

References 87 publications

The Smooth Muscle Cell-restricted KCNMB1 Ion Channel Subunit Is a Direct Transcriptional Target of Serum Response Factor and Myocardin

The Smooth Muscle Cell-restricted KCNMB1 Ion Channel Subunit Is a Direct Transcriptional Target of Serum Response Factor and Myocardin

Transcription factors and miRNAs that regulate fetal to adultCFTRexpression change are new targets for cystic fibrosis

Homozygosity for a null allele of COL3A1 results in recessive Ehlers–Danlos syndrome

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