title: The miR-143/145 cluster is a novel transcriptional target of Jagged-1/Notch signaling in vascular smooth muscle cells.
abstract: Activation of Notch signaling by Jagged-1 (Jag-1) in vascular smooth muscle cells (VSMC) promotes a differentiated phenotype characterized by increased expression of contractile proteins. Recent studies show that microRNAs (miR)-143/145 regulates VSMC phenotype. The serum response factor (SRF)/myocardin complex binds to CArG sequences to activate miR-143/145 transcription, but no other regulators are known in VSMC. Using miR arrays, we found miR-143/145 induced following expression of a constitutively active Notch1 intracellular domain (N1ICD). We hypothesized that miR-143/145 is required for Jag-1/Notch-induced VSMC differentiation. Activation of Notch receptors by Jag-1 caused CBF1-dependent up-regulation of miR-143/145, increased differentiation, and decreased proliferation. Conversely, inhibiting basal Notch signaling decreased steady state levels of miR-143/145. Using SRF knockdown, we found that Jag-1/Notch induction of miR-143/145 is SRF independent, although full acquisition of contractile markers requires SRF. Using miR-143/145 promoter reporter constructs we show Jag-1/Notch increases promoter activity, and this is dependent on intact CBF1 consensus sites within the promoter. Chromatin immunoprecipitation (ChIP) assays revealed that N1ICD-containing complexes bind to CBF1 sites in the miR-143/145 promoter. We also identified N1ICD complex binding to CBF1 sites within the endogenous human miR-143/145 promoter. Using miR-143/145-interfering oligonucleotides, we demonstrate that Jag-1/Notch signaling requires induction of both miR-143 and miR-145 to promote the VSMC contractile phenotype. Thus, miR-143/145 is a novel transcriptional target of Jag-1/Notch signaling in VSMC. We propose miR-143/145 as activated independently by Jag-1/Notch and SRF in parallel pathways. Multiple pathways converging on miR-143/145 provides potential for fine-tuning or amplification of VSMC differentiation signals.
authors.affiliation: Center for Molecular Medicine, Maine Medical Center Research Institute, Scarborough, Maine 04074, USA.
authors.forename: Joshua M
authors.initials: JM
authors.lastname: Boucher
authors.forename: Sarah M
authors.initials: SM
authors.lastname: Peterson
authors.forename: Sumithra
authors.initials: S
authors.lastname: Urs
authors.forename: Chunxiang
authors.initials: C
authors.lastname: Zhang
authors.forename: Lucy
authors.initials: L
authors.lastname: Liaw
mesh_terms: D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D055785:Gene Knockdown Techniques; D006801:Humans; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein / Q000235:genetics / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D000072100:Jagged-1 Protein; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D005810:Multigene Family / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000502:physiology; D000072099:Serrate-Jagged Proteins; D026362:Serum Response Factor / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D002135:Calcium-Binding Proteins; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D036341:Intercellular Signaling Peptides and Proteins; C000606273:JAG1 protein, human; D000072100:Jagged-1 Protein; C542359:MIRN143 microRNA, human; C541010:MIRN145 microRNA, human; D008565:Membrane Proteins; D035683:MicroRNAs; C497570:RBPJ protein, human; D051880:Receptors, Notch; C476858:SRF protein, human; D000072099:Serrate-Jagged Proteins; D026362:Serum Response Factor