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Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer

Tai, Phillip W. L. and Fisher-Aylor, Katherine I. and Himeda, Charis L. and Smith, Catherine L. and MacKenzie, Alexandra P. and Helterline, Deri L. and Angello, John C. and Welikson, Robert E. and Wold, Barbara J. and Hauschka, Stephen D. (2011) Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer. Skeletal Muscle, 1 . Art. No. 25. ISSN 2044-5040. PMCID PMC3157005.

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Background: Hundreds of genes, including muscle creatine kinase (MCK), are differentially expressed in fast- and slow-twitch muscle fibers, but the fiber type-specific regulatory mechanisms are not well understood. Results: Modulatory region 1 (MR1) is a 1-kb regulatory region within MCK intron 1 that is highly active in terminally differentiating skeletal myocytes in vitro. A MCK small intronic enhancer (MCK-SIE) containing a paired E-box/myocyte enhancer factor 2 (MEF2) regulatory motif resides within MR1. The SIE's transcriptional activity equals that of the extensively characterized 206-bp MCK 5'-enhancer, but the MCK-SIE is flanked by regions that can repress its activity via the individual and combined effects of about 15 different but highly conserved 9- to 24-bp sequences. ChIP and ChIP-Seq analyses indicate that the SIE and the MCK 5'-enhancer are occupied by MyoD, myogenin and MEF2. Many other E-boxes located within or immediately adjacent to intron 1 are not occupied by MyoD or myogenin. Transgenic analysis of a 6.5-kb MCK genomic fragment containing the 5'-enhancer and proximal promoter plus the 3.2-kb intron 1, with and without MR1, indicates that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers (types I and IIa, respectively), but is not required for expression in fast-twitch muscle fibers (types IIb and IId). Conclusions: In this study, we discovered that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers and that MR1's positive transcriptional activity depends on a paired E-box MEF2 site motif within a SIE. This is the first study to delineate the DNA controls for MCK expression in different skeletal muscle fiber types.

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Wold, Barbara J.0000-0003-3235-8130
Additional Information:© 2011 Tai et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Received: 5 January 2011; Accepted: 7 July 2011; Published: 7 July 2011. We thank members of the Hauschka and Jeff Chamberlain laboratories at the University of Washington, Q. Nguyen, D. Helterline, E. Nishiuchi, M. Haraguchi, P. Gregorevic, B. Sharma, A. Zebari and J. Buskin, for their technical assistance and useful comments. We thank members of the Wold Lab at the California Institute of Technology, D. Trout, B. King, H. Amrhein, L. Schaffer and I. Antoschken, for technical assistance, bioinformatics and/or critical discussions. This work was supported by National Institutes of Health (NIH) grant R01-0AR18860 (to SDH), NIH grant 1P01-NS046788 (to SDH), NIH grant 5732-HD07183 Developmental Biology Training Grant (to PWLT) and NIH grant T32-HL007312, Experimental Pathology of Cardiovascular Disease (to CLH). KF is supported by a Graduate Research Fellowship from the National Science Foundation and from the Beckman Foundation at the California Institute of Technology; BJW received funding from NIH grant U54 HG004576. Authors’ contributions: PWLT carried out the sequence alignments; made the test gene constructs; carried out the transfection assays, the ChIP analyses, the immunohistochemistry and immunofluorescence assays; and drafted parts of the manuscript describing Hauschka Lab data. KIFA conceived of the redesign of the ChIP-Seq fixation, performed and analyzed MEF2 ChIP-Seq and drafted portions of the manuscript describing Wold Lab data. CLH carried out the EMSA study and helped to draft the manuscript. CLS participated in the immunohistochemistry and immunofluorescence assays. APM participated in the transfection analyses. DLH carried out the whole muscle extract transgene expression assays. JCA and REW prepared and labeled the MYHC monoclonal antibodies and participated in the immunohistochemistry assays. BJW conceived of the global ChIP-Seq analysis of multiple myogenic transcription factors, participated in the design and coordination of the MEF2 ChIP-Seq studies and drafted parts of the manuscript describing the Wold Lab data. Together with PWLT, SDH conceived of the overall study, participated in its design and coordination between the two laboratories and played a major role in writing the manuscript. All authors read and approved the final manuscript.
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NSF Graduate Research FellowshipUNSPECIFIED
Caltech Beckman FoundationUNSPECIFIED
NIHU54 HG004576
PubMed Central ID:PMC3157005
Record Number:CaltechAUTHORS:20131107-133226265
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Official Citation:Tai et al.: Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer. Skeletal Muscle 2011 1:25
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:42308
Deposited By: Tony Diaz
Deposited On:07 Nov 2013 22:02
Last Modified:09 Mar 2020 13:19

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