title: MiR-133 promotes cardiac reprogramming by directly repressing Snai1 and silencing fibroblast signatures.
abstract: Fibroblasts can be directly reprogrammed into cardiomyocyte-like cells (iCMs) by overexpression of cardiac transcription factors or microRNAs. However, induction of functional cardiomyocytes is inefficient, and molecular mechanisms of direct reprogramming remain undefined. Here, we demonstrate that addition of miR-133a (miR-133) to Gata4, Mef2c, and Tbx5 (GMT) or GMT plus Mesp1 and Myocd improved cardiac reprogramming from mouse or human fibroblasts by directly repressing Snai1, a master regulator of epithelial-to-mesenchymal transition. MiR-133 overexpression with GMT generated sevenfold more beating iCMs from mouse embryonic fibroblasts and shortened the duration to induce beating cells from 30 to 10 days, compared to GMT alone. Snai1 knockdown suppressed fibroblast genes, upregulated cardiac gene expression, and induced more contracting iCMs with GMT transduction, recapitulating the effects of miR-133 overexpression. In contrast, overexpression of Snai1 in GMT/miR-133-transduced cells maintained fibroblast signatures and inhibited generation of beating iCMs. MiR-133-mediated Snai1 repression was also critical for cardiac reprogramming in adult mouse and human cardiac fibroblasts. Thus, silencing fibroblast signatures, mediated by miR-133/Snai1, is a key molecular roadblock during cardiac reprogramming.
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Muraoka
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamakawa
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Miyamoto
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Taketaro
authors.initials: T
authors.lastname: Sadahiro
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Umei
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Isomi
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Hanae
authors.initials: H
authors.lastname: Nakashima
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Mizuha
authors.initials: M
authors.lastname: Akiyama
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Rie
authors.initials: R
authors.lastname: Wada
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Kohei
authors.initials: K
authors.lastname: Inagawa
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Takahiko
authors.initials: T
authors.lastname: Nishiyama
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Ruri
authors.initials: R
authors.lastname: Kaneda
authors.affiliation: Department of Physiology and Cell Biology, Tokyo Medical and Dental University, Bunkyo-ku Tokyo, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Fukuda
authors.affiliation: Department of Physiology and Cell Biology, Tokyo Medical and Dental University, Bunkyo-ku Tokyo, Japan.
authors.forename: Shu
authors.initials: S
authors.lastname: Takeda
authors.affiliation: Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Shugo
authors.initials: S
authors.lastname: Tohyama
authors.affiliation: Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Hisayuki
authors.initials: H
authors.lastname: Hashimoto
authors.affiliation: Japan Biological Informatics Consortium (JBiC), Koto-ku Tokyo, Japan.
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Kawamura
authors.affiliation: Molecular Profiling Research Center for Drug Discovery, National Institute of Advanced Industrial Science and Technology, Koto-ku Tokyo, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Goshima
authors.affiliation: Division of Cardiovascular Surgery, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Aeba
authors.affiliation: Department of Pediatrics, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamagishi
authors.affiliation: Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan.
authors.forename: Keiichi
authors.initials: K
authors.lastname: Fukuda
authors.affiliation: Department of Clinical and Molecular Cardiovascular Research, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan Department of Cardiology, Keio University School of Medicine, Shinjuku-ku Tokyo, Japan JST CREST, Shinjuku-ku Tokyo, Japan mieda@z8.keio.jp.
authors.forename: Masaki
authors.initials: M
authors.lastname: Ieda
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D054338:Cell Transdifferentiation / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000502:physiology*; D055785:Gene Knockdown Techniques; D049452:Green Fluorescent Proteins; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism*; D046228:Microarray Analysis; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: D035683:MicroRNAs; C517732:Mirn133 microRNA, mouse; C000604407:SNAI1 protein, human; C000604429:Snai1 protein, mouse; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; D049452:Green Fluorescent Proteins
keywords: Snai1; cardiomyocyte; microRNA; reprogramming; transcription factor