• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Jul 1988; 8(7): 2896–2909.
PMCID: PMC363509

Identification of upstream and intragenic regulatory elements that confer cell-type-restricted and differentiation-specific expression on the muscle creatine kinase gene.


Terminal differentiation of skeletal myoblasts is accompanied by induction of a series of tissue-specific gene products, which includes the muscle isoenzyme of creatine kinase (MCK). To begin to define the sequences and signals involved in MCK regulation in developing muscle cells, the mouse MCK gene has been isolated. Sequence analysis of 4,147 bases of DNA surrounding the transcription initiation site revealed several interesting structural features, some of which are common to other muscle-specific genes and to cellular and viral enhancers. To test for sequences required for regulated expression, a region upstream of the MCK gene from -4800 to +1 base pairs, relative to the transcription initiation site, was linked to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene. Introduction of this MCK-CAT fusion gene into C2 muscle cells resulted in high-level expression of CAT activity in differentiated myotubes and no detectable expression in proliferating undifferentiated myoblasts or in nonmyogenic cell lines. Deletion mutagenesis of sequences between -4800 and the transcription start site showed that the region between -1351 and -1050 was sufficient to confer cell type-specific and developmentally regulated expression on the MCK promoter. This upstream regulatory element functioned independently of position, orientation, or distance from the promoter and therefore exhibited the properties of a classical enhancer. This upstream enhancer also was able to confer muscle-specific regulation on the simian virus 40 promoter, although it exhibited a 3- to 5-fold preference for its own promoter. In contrast to the cell type- and differentiation-specific expression of the upstream enhancer, the MCK promoter was able to function in myoblasts and myotubes and in nonmyogenic cell lines when combined with the simian virus 40 enhancer. An additional positive regulatory element was identified within the first intron of the MCK gene. Like the upstream enhancer, this intragenic element functioned independently of position, orientation, and distance with respect to the MCK promoter and was active in differentiated myotubes but not in myoblasts. These results demonstrate that expression of the MCK gene in developing muscle cells is controlled by complex interactions among multiple upstream and intragenic regulatory elements that are functional only in the appropriate cellular context.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (3.1M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Jaynes JB, Johnson JE, Buskin JN, Gartside CL, Hauschka SD. The muscle creatine kinase gene is regulated by multiple upstream elements, including a muscle-specific enhancer. Mol Cell Biol. 1988 Jan;8(1):62–70. [PMC free article] [PubMed]
  • Baldwin AS, Jr, Kittler EL, Emerson CP., Jr Structure, evolution, and regulation of a fast skeletal muscle troponin I gene. Proc Natl Acad Sci U S A. 1985 Dec;82(23):8080–8084. [PMC free article] [PubMed]
  • Banerji J, Olson L, Schaffner W. A lymphocyte-specific cellular enhancer is located downstream of the joining region in immunoglobulin heavy chain genes. Cell. 1983 Jul;33(3):729–740. [PubMed]
  • Bergsma DJ, Grichnik JM, Gossett LM, Schwartz RJ. Delimitation and characterization of cis-acting DNA sequences required for the regulated expression and transcriptional control of the chicken skeletal alpha-actin gene. Mol Cell Biol. 1986 Jul;6(7):2462–2475. [PMC free article] [PubMed]
  • Bessman SP, Carpenter CL. The creatine-creatine phosphate energy shuttle. Annu Rev Biochem. 1985;54:831–862. [PubMed]
  • Blau HM, Pavlath GK, Hardeman EC, Chiu CP, Silberstein L, Webster SG, Miller SC, Webster C. Plasticity of the differentiated state. Science. 1985 Nov 15;230(4727):758–766. [PubMed]
  • Bouvagnet PF, Strehler EE, White GE, Strehler-Page MA, Nadal-Ginard B, Mahdavi V. Multiple positive and negative 5' regulatory elements control the cell-type-specific expression of the embryonic skeletal myosin heavy-chain gene. Mol Cell Biol. 1987 Dec;7(12):4377–4389. [PMC free article] [PubMed]
  • Caravatti M, Minty A, Robert B, Montarras D, Weydert A, Cohen A, Daubas P, Buckingham M. Regulation of muscle gene expression. The accumulation of messenger RNAs coding for muscle-specific proteins during myogenesis in a mouse cell line. J Mol Biol. 1982 Sep;160(1):59–76. [PubMed]
  • Chamberlain JS, Jaynes JB, Hauschka SD. Regulation of creatine kinase induction in differentiating mouse myoblasts. Mol Cell Biol. 1985 Mar;5(3):484–492. [PMC free article] [PubMed]
  • Chen EY, Seeburg PH. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. [PubMed]
  • Clemens MJ. A potential role for RNA transcribed from B2 repeats in the regulation of mRNA stability. Cell. 1987 Apr 24;49(2):157–158. [PubMed]
  • Davidson I, Fromental C, Augereau P, Wildeman A, Zenke M, Chambon P. Cell-type specific protein binding to the enhancer of simian virus 40 in nuclear extracts. Nature. 1986 Oct 9;323(6088):544–548. [PubMed]
  • Devlin RB, Emerson CP., Jr Coordinate regulation of contractile protein synthesis during myoblast differentiation. Cell. 1978 Apr;13(4):599–611. [PubMed]
  • Fambrough DM. Control of acetylcholine receptors in skeletal muscle. Physiol Rev. 1979 Jan;59(1):165–227. [PubMed]
  • Geyer PK, Fyrberg EA. 5'-flanking sequence required for regulated expression of a muscle-specific Drosophila melanogaster actin gene. Mol Cell Biol. 1986 Oct;6(10):3388–3396. [PMC free article] [PubMed]
  • Graham FL, van der Eb AJ. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. [PubMed]
  • Grichnik JM, Bergsma DJ, Schwartz RJ. Tissue restricted and stage specific transcription is maintained within 411 nucleotides flanking the 5' end of the chicken alpha-skeletal actin gene. Nucleic Acids Res. 1986 Feb 25;14(4):1683–1701. [PMC free article] [PubMed]
  • Grosschedl R, Baltimore D. Cell-type specificity of immunoglobulin gene expression is regulated by at least three DNA sequence elements. Cell. 1985 Jul;41(3):885–897. [PubMed]
  • Gustafson TA, Markham BE, Bahl JJ, Morkin E. Thyroid hormone regulates expression of a transfected alpha-myosin heavy-chain fusion gene in fetal heart cells. Proc Natl Acad Sci U S A. 1987 May;84(10):3122–3126. [PMC free article] [PubMed]
  • Hastings KE, Emerson CP., Jr cDNA clone analysis of six co-regulated mRNAs encoding skeletal muscle contractile proteins. Proc Natl Acad Sci U S A. 1982 Mar;79(5):1553–1557. [PMC free article] [PubMed]
  • Hayward LJ, Schwartz RJ. Sequential expression of chicken actin genes during myogenesis. J Cell Biol. 1986 Apr;102(4):1485–1493. [PMC free article] [PubMed]
  • Hickey R, Skoultchi A, Gunning P, Kedes L. Regulation of a human cardiac actin gene introduced into rat L6 myoblasts suggests a defect in their myogenic program. Mol Cell Biol. 1986 Sep;6(9):3287–3290. [PMC free article] [PubMed]
  • Hu MC, Sharp SB, Davidson N. The complete sequence of the mouse skeletal alpha-actin gene reveals several conserved and inverted repeat sequences outside of the protein-coding region. Mol Cell Biol. 1986 Jan;6(1):15–25. [PMC free article] [PubMed]
  • Imagawa M, Chiu R, Karin M. Transcription factor AP-2 mediates induction by two different signal-transduction pathways: protein kinase C and cAMP. Cell. 1987 Oct 23;51(2):251–260. [PubMed]
  • Inestrosa NC, Miller JB, Silberstein L, Ziskind-Conhaim L, Hall ZW. Developmental regulation of 16S acetylcholinesterase and acetylcholine receptors in a mouse muscle cell line. Exp Cell Res. 1983 Sep;147(2):393–405. [PubMed]
  • Ish-Horowicz D, Burke JF. Rapid and efficient cosmid cloning. Nucleic Acids Res. 1981 Jul 10;9(13):2989–2998. [PMC free article] [PubMed]
  • Jaynes JB, Chamberlain JS, Buskin JN, Johnson JE, Hauschka SD. Transcriptional regulation of the muscle creatine kinase gene and regulated expression in transfected mouse myoblasts. Mol Cell Biol. 1986 Aug;6(8):2855–2864. [PMC free article] [PubMed]
  • Klarsfeld A, Daubas P, Bourachot B, Changeux JP. A 5'-flanking region of the chicken acetylcholine receptor alpha-subunit gene confers tissue specificity and developmental control of expression in transfected cells. Mol Cell Biol. 1987 Feb;7(2):951–955. [PMC free article] [PubMed]
  • Konieczny SF, Emerson CP., Jr Differentiation, not determination, regulates muscle gene activation: transfection of troponin I genes into multipotential and muscle lineages of 10T1/2 cells. Mol Cell Biol. 1985 Sep;5(9):2423–2432. [PMC free article] [PubMed]
  • Konieczny SF, Emerson CP., Jr Complex regulation of the muscle-specific contractile protein (troponin I) gene. Mol Cell Biol. 1987 Sep;7(9):3065–3075. [PMC free article] [PubMed]
  • Lathrop B, Olson E, Glaser L. Control by fibroblast growth factor of differentiation in the BC3H1 muscle cell line. J Cell Biol. 1985 May;100(5):1540–1547. [PMC free article] [PubMed]
  • Mahdavi V, Chambers AP, Nadal-Ginard B. Cardiac alpha- and beta-myosin heavy chain genes are organized in tandem. Proc Natl Acad Sci U S A. 1984 May;81(9):2626–2630. [PMC free article] [PubMed]
  • Melloul D, Aloni B, Calvo J, Yaffe D, Nudel U. Developmentally regulated expression of chimeric genes containing muscle actin DNA sequences in transfected myogenic cells. EMBO J. 1984 May;3(5):983–990. [PMC free article] [PubMed]
  • Merlie JP, Buckingham ME, Whalen RG. Molecular aspects of myogenesis. Curr Top Dev Biol. 1977;11:61–114. [PubMed]
  • Minty A, Blau H, Kedes L. Two-level regulation of cardiac actin gene transcription: muscle-specific modulating factors can accumulate before gene activation. Mol Cell Biol. 1986 Jun;6(6):2137–2148. [PMC free article] [PubMed]
  • Minty A, Kedes L. Upstream regions of the human cardiac actin gene that modulate its transcription in muscle cells: presence of an evolutionarily conserved repeated motif. Mol Cell Biol. 1986 Jun;6(6):2125–2136. [PMC free article] [PubMed]
  • Mitchell PJ, Wang C, Tjian R. Positive and negative regulation of transcription in vitro: enhancer-binding protein AP-2 is inhibited by SV40 T antigen. Cell. 1987 Sep 11;50(6):847–861. [PubMed]
  • Miwa T, Kedes L. Duplicated CArG box domains have positive and mutually dependent regulatory roles in expression of the human alpha-cardiac actin gene. Mol Cell Biol. 1987 Aug;7(8):2803–2813. [PMC free article] [PubMed]
  • Moore DD, Marks AR, Buckley DI, Kapler G, Payvar F, Goodman HM. The first intron of the human growth hormone gene contains a binding site for glucocorticoid receptor. Proc Natl Acad Sci U S A. 1985 Feb;82(3):699–702. [PMC free article] [PubMed]
  • Nadal-Ginard B. Commitment, fusion and biochemical differentiation of a myogenic cell line in the absence of DNA synthesis. Cell. 1978 Nov;15(3):855–864. [PubMed]
  • Nudel U, Calvo JM, Shani M, Levy Z. The nucleotide sequence of a rat myosin light chain 2 gene. Nucleic Acids Res. 1984 Sep 25;12(18):7175–7186. [PMC free article] [PubMed]
  • Nudel U, Greenberg D, Ordahl CP, Saxel O, Neuman S, Yaffe D. Developmentally regulated expression of a chicken muscle-specific gene in stably transfected rat myogenic cells. Proc Natl Acad Sci U S A. 1985 May;82(10):3106–3109. [PMC free article] [PubMed]
  • Olson EN, Caldwell KL, Gordon JI, Glaser L. Regulation of creatine phosphokinase expression during differentiation of BC3H1 cells. J Biol Chem. 1983 Feb 25;258(4):2644–2652. [PubMed]
  • Olson EN, Spizz G, Tainsky MA. The oncogenic forms of N-ras or H-ras prevent skeletal myoblast differentiation. Mol Cell Biol. 1987 Jun;7(6):2104–2111. [PMC free article] [PubMed]
  • Olson EN, Sternberg E, Hu JS, Spizz G, Wilcox C. Regulation of myogenic differentiation by type beta transforming growth factor. J Cell Biol. 1986 Nov;103(5):1799–1805. [PMC free article] [PubMed]
  • Perriard JC, Caravatti M, Perriard ER, Eppenberger HM. Quantitation of creatine kinase isoenzyme transition in differentiating chicken embryonic breast muscle and myogenic cell cultures by immunoadsorption. Arch Biochem Biophys. 1978 Nov;191(1):90–100. [PubMed]
  • Picard D, Schaffner W. A lymphocyte-specific enhancer in the mouse immunoglobulin kappa gene. Nature. 1984 Jan 5;307(5946):80–82. [PubMed]
  • Poncz M, Solowiejczyk D, Ballantine M, Schwartz E, Surrey S. "Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method. Proc Natl Acad Sci U S A. 1982 Jul;79(14):4298–4302. [PMC free article] [PubMed]
  • Roman D, Billadello J, Gordon J, Grace A, Sobel B, Strauss A. Complete nucleotide sequence of dog heart creatine kinase mRNA: conservation of amino acid sequence within and among species. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8394–8398. [PMC free article] [PubMed]
  • Rossi P, de Crombrugghe B. Identification of a cell-specific transcriptional enhancer in the first intron of the mouse alpha 2 (type I) collagen gene. Proc Natl Acad Sci U S A. 1987 Aug;84(16):5590–5594. [PMC free article] [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Schmid CW, Jelinek WR. The Alu family of dispersed repetitive sequences. Science. 1982 Jun 4;216(4550):1065–1070. [PubMed]
  • Schwartz RJ, Rothblum KN. Gene switching in myogenesis: differential expression of the chicken actin multigene family. Biochemistry. 1981 Jul 7;20(14):4122–4129. [PubMed]
  • Seiler-Tuyns A, Eldridge JD, Paterson BM. Expression and regulation of chicken actin genes introduced into mouse myogenic and nonmyogenic cells. Proc Natl Acad Sci U S A. 1984 May;81(10):2980–2984. [PMC free article] [PubMed]
  • Sen R, Baltimore D. Multiple nuclear factors interact with the immunoglobulin enhancer sequences. Cell. 1986 Aug 29;46(5):705–716. [PubMed]
  • Sen R, Baltimore D. Inducibility of kappa immunoglobulin enhancer-binding protein Nf-kappa B by a posttranslational mechanism. Cell. 1986 Dec 26;47(6):921–928. [PubMed]
  • Shani M. Tissue-specific expression of rat myosin light-chain 2 gene in transgenic mice. Nature. 1985 Mar 21;314(6008):283–286. [PubMed]
  • Shani M. Tissue-specific and developmentally regulated expression of a chimeric actin-globin gene in transgenic mice. Mol Cell Biol. 1986 Jul;6(7):2624–2631. [PMC free article] [PubMed]
  • Spizz G, Roman D, Strauss A, Olson EN. Serum and fibroblast growth factor inhibit myogenic differentiation through a mechanism dependent on protein synthesis and independent of cell proliferation. J Biol Chem. 1986 Jul 15;261(20):9483–9488. [PubMed]
  • Wilson C, Cross GS, Woodland HR. Tissue-specific expression of actin genes injected into Xenopus embryos. Cell. 1986 Nov 21;47(4):589–599. [PubMed]
  • Yaffe D, Saxel O. Serial passaging and differentiation of myogenic cells isolated from dystrophic mouse muscle. Nature. 1977 Dec 22;270(5639):725–727. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • MedGen
    Related information in MedGen
  • Nucleotide
    Published Nucleotide sequences
  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...