Europe PMC
Nothing Special   »   [go: up one dir, main page]

Europe PMC requires Javascript to function effectively.

Either your web browser doesn't support Javascript or it is currently turned off. In the latter case, please turn on Javascript support in your web browser and reload this page.

This website requires cookies, and the limited processing of your personal data in order to function. By using the site you are agreeing to this as outlined in our privacy notice and cookie policy.

Smoothelin, a novel cytoskeletal protein specific for smooth muscle cells.

Author information

Affiliations

  • 1. Department of Molecular Cell Biology & Genetics, University of Limburg, Maastricht, The Netherlands.
      Authors
    • van der Loop FT 1
    (1 author)

The Journal of Cell Biology, 01 Jul 1996, 134(2):401-411
https://doi.org/10.1083/jcb.134.2.401 PMID: 8707825 PMCID: PMC2120883

This article is in the Europe PMC Open access subset. Refer to the copyright information in the article for licensing details.
Free full text in Europe PMC

Abstract 

The characterization of a novel 59-kD cytoskeletal protein is described. It is exclusively observed in smooth muscle cells by Northern blotting and immunohistochemical analysis and therefore designated "smoothelin." A human smooth muscle cDNA library was screened with the monoclonal antibody R4A, and a full-size cDNA of the protein was selected. The cDNA was sequenced and appeared to contain a 1,113-bp open reading frame. Based on the cDNA sequence, the calculated molecular weight of the polypeptide was 40 kD and it was demonstrated to contain two N-glycosylation sites. Computer assisted analysis at the protein level revealed a 56-amino acid domain with homologies of approximately 40% with a sequence bordering the actin-binding domains of dystrophin, utrophin, beta-spectrin and alpha-actinin. In situ hybridization demonstrated that human smoothelin is encoded by a single copy gene which is located on chromosome 22. Immunohistochemistry and Western blotting revealed synthesis of smoothelin in smooth muscle of species evolutionarily as far apart as human and teleost. Northern blotting indicated that sequence as well as size of the mRNA (approximately 1,500 bases) are conserved among vertebrates. Cell fractionation studies and differential centrifugation showed that the protein cannot be extracted with Triton X-100, which indicates that it is a part of the cytoskeleton. Transfection of the human cDNA into smooth muscle cells and COS7 cells produced a protein of 59 kD, which assembled into a filamentous network. However, in rat heart-derived myoblasts association with stress fibers was most prominent. Smoothelin was not detected in primary or long term smooth muscle cell cultures. Also, transcription of smoothelin mRNA was almost instantly halted in smooth muscle tissue explants. We conclude that smoothelin is a new cytoskeletal protein that is only found in contractile smooth muscle cells and does not belong to one of the classes of structural proteins presently known.

Free full text 

Logo of jcellbiolLink to Publisher's site
J Cell Biol. 1996 Jul 2; 134(2): 401–411.
PMCID: PMC2120883
PMID: 8707825

Smoothelin, a novel cytoskeletal protein specific for smooth muscle cells

Copyright and License information Disclaimer

Abstract

The characterization of a novel 59-kD cytoskeletal protein is described. It is exclusively observed in smooth muscle cells by Northern blotting and immunohistochemical analysis and therefore designated "smoothelin." A human smooth muscle cDNA library was screened with the monoclonal antibody R4A, and a full-size cDNA of the protein was selected. The cDNA was sequenced and appeared to contain a 1,113-bp open reading frame. Based on the cDNA sequence, the calculated molecular weight of the polypeptide was 40 kD and it was demonstrated to contain two N-glycosylation sites. Computer assisted analysis at the protein level revealed a 56-amino acid domain with homologies of approximately 40% with a sequence bordering the actin-binding domains of dystrophin, utrophin, beta-spectrin and alpha-actinin. In situ hybridization demonstrated that human smoothelin is encoded by a single copy gene which is located on chromosome 22. Immunohistochemistry and Western blotting revealed synthesis of smoothelin in smooth muscle of species evolutionarily as far apart as human and teleost. Northern blotting indicated that sequence as well as size of the mRNA (approximately 1,500 bases) are conserved among vertebrates. Cell fractionation studies and differential centrifugation showed that the protein cannot be extracted with Triton X-100, which indicates that it is a part of the cytoskeleton. Transfection of the human cDNA into smooth muscle cells and COS7 cells produced a protein of 59 kD, which assembled into a filamentous network. However, in rat heart-derived myoblasts association with stress fibers was most prominent. Smoothelin was not detected in primary or long term smooth muscle cell cultures. Also, transcription of smoothelin mRNA was almost instantly halted in smooth muscle tissue explants. We conclude that smoothelin is a new cytoskeletal protein that is only found in contractile smooth muscle cells and does not belong to one of the classes of structural proteins presently known.

Full Text

The Full Text of this article is available as a PDF (3.9M).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. [Abstract] [Google Scholar]
  • Auffray C, Rougeon F. Purification of mouse immunoglobulin heavy-chain messenger RNAs from total myeloma tumor RNA. Eur J Biochem. 1980 Jun;107(2):303–314. [Abstract] [Google Scholar]
  • Aviv H, Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. [Europe PMC free article] [Abstract] [Google Scholar]
  • Babai F, Musevi-Aghdam J, Schurch W, Royal A, Gabbiani G. Coexpression of alpha-sarcomeric actin, alpha-smooth muscle actin and desmin during myogenesis in rat and mouse embryos I. Skeletal muscle. Differentiation. 1990 Aug;44(2):132–142. [Abstract] [Google Scholar]
  • Baron MD, Davison MD, Jones P, Critchley DR. The sequence of chick alpha-actinin reveals homologies to spectrin and calmodulin. J Biol Chem. 1987 Dec 25;262(36):17623–17629. [Abstract] [Google Scholar]
  • Beggs AH, Byers TJ, Knoll JH, Boyce FM, Bruns GA, Kunkel LM. Cloning and characterization of two human skeletal muscle alpha-actinin genes located on chromosomes 1 and 11. J Biol Chem. 1992 May 5;267(13):9281–9288. [Abstract] [Google Scholar]
  • Berbers GA, Hoekman WA, Bloemendal H, de Jong WW, Kleinschmidt T, Braunitzer G. Proline- and alanine-rich N-terminal extension of the basic bovine beta-crystallin B1 chains. FEBS Lett. 1983 Sep 19;161(2):225–229. [Abstract] [Google Scholar]
  • Borrione AC, Zanellato AM, Giuriato L, Scannapieco G, Pauletto P, Sartore S. Nonmuscle and smooth muscle myosin isoforms in bovine endothelial cells. Exp Cell Res. 1990 Sep;190(1):1–10. [Abstract] [Google Scholar]
  • Campbell GR, Chamley-Campbell JH. Smooth muscle phenotypic modulation: role in atherogenesis. Med Hypotheses. 1981 Jun;7(6):729–735. [Abstract] [Google Scholar]
  • Campbell JH, Kocher O, Skalli O, Gabbiani G, Campbell GR. Cytodifferentiation and expression of alpha-smooth muscle actin mRNA and protein during primary culture of aortic smooth muscle cells. Correlation with cell density and proliferative state. Arteriosclerosis. 1989 Sep-Oct;9(5):633–643. [Abstract] [Google Scholar]
  • Church GM, Gilbert W. Genomic sequencing. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1991–1995. [Europe PMC free article] [Abstract] [Google Scholar]
  • Dhermy D. The spectrin super-family. Biol Cell. 1991;71(3):249–254. [Abstract] [Google Scholar]
  • Duband JL, Gimona M, Scatena M, Sartore S, Small JV. Calponin and SM 22 as differentiation markers of smooth muscle: spatiotemporal distribution during avian embryonic development. Differentiation. 1993 Dec;55(1):1–11. [Abstract] [Google Scholar]
  • Frid MG, Shekhonin BV, Koteliansky VE, Glukhova MA. Phenotypic changes of human smooth muscle cells during development: late expression of heavy caldesmon and calponin. Dev Biol. 1992 Oct;153(2):185–193. [Abstract] [Google Scholar]
  • Gabbiani G, Schmid E, Winter S, Chaponnier C, de Ckhastonay C, Vandekerckhove J, Weber K, Franke WW. Vascular smooth muscle cells differ from other smooth muscle cells: predominance of vimentin filaments and a specific alpha-type actin. Proc Natl Acad Sci U S A. 1981 Jan;78(1):298–302. [Europe PMC free article] [Abstract] [Google Scholar]
  • Glukhova MA, Kabakov AE, Belkin AM, Frid MG, Ornatsky OI, Zhidkova NI, Koteliansky VE. Meta-vinculin distribution in adult human tissues and cultured cells. FEBS Lett. 1986 Oct 20;207(1):139–141. [Abstract] [Google Scholar]
  • Gluzman Y. SV40-transformed simian cells support the replication of early SV40 mutants. Cell. 1981 Jan;23(1):175–182. [Abstract] [Google Scholar]
  • Gunning P, Gordon M, Wade R, Gahlmann R, Lin CS, Hardeman E. Differential control of tropomyosin mRNA levels during myogenesis suggests the existence of an isoform competition-autoregulatory compensation control mechanism. Dev Biol. 1990 Apr;138(2):443–453. [Abstract] [Google Scholar]
  • Haeberle JR, Hathaway DR, Smith CL. Caldesmon content of mammalian smooth muscles. J Muscle Res Cell Motil. 1992 Feb;13(1):81–89. [Abstract] [Google Scholar]
  • Karinch AM, Zimmer WE, Goodman SR. The identification and sequence of the actin-binding domain of human red blood cell beta-spectrin. J Biol Chem. 1990 Jul 15;265(20):11833–11840. [Abstract] [Google Scholar]
  • Kimes BW, Brandt BL. Properties of a clonal muscle cell line from rat heart. Exp Cell Res. 1976 Mar 15;98(2):367–381. [Abstract] [Google Scholar]
  • Koenig M, Monaco AP, Kunkel LM. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. [Abstract] [Google Scholar]
  • Köhler G, Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975 Aug 7;256(5517):495–497. [Abstract] [Google Scholar]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [Abstract] [Google Scholar]
  • Lees-Miller JP, Heeley DH, Smillie LB. An abundant and novel protein of 22 kDa (SM22) is widely distributed in smooth muscles. Purification from bovine aorta. Biochem J. 1987 Jun 15;244(3):705–709. [Europe PMC free article] [Abstract] [Google Scholar]
  • McHugh KM, Crawford K, Lessard JL. A comprehensive analysis of the developmental and tissue-specific expression of the isoactin multigene family in the rat. Dev Biol. 1991 Dec;148(2):442–458. [Abstract] [Google Scholar]
  • Nagai R, Kuro-o M, Babij P, Periasamy M. Identification of two types of smooth muscle myosin heavy chain isoforms by cDNA cloning and immunoblot analysis. J Biol Chem. 1989 Jun 15;264(17):9734–9737. [Abstract] [Google Scholar]
  • Nanaev AK, Shirinsky VP, Birukov KG. Immunofluorescent study of heterogeneity in smooth muscle cells of human fetal vessels using antibodies to myosin, desmin, and vimentin. Cell Tissue Res. 1991 Dec;266(3):535–540. [Abstract] [Google Scholar]
  • O'Farrell PH. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [Europe PMC free article] [Abstract] [Google Scholar]
  • Owens GK. Regulation of differentiation of vascular smooth muscle cells. Physiol Rev. 1995 Jul;75(3):487–517. [Abstract] [Google Scholar]
  • Pless DD, Lennarz WJ. Enzymatic conversion of proteins to glycoproteins. Proc Natl Acad Sci U S A. 1977 Jan;74(1):134–138. [Europe PMC free article] [Abstract] [Google Scholar]
  • Quax-Jeuken YE, Quax WJ, Bloemendal H. Primary and secondary structure of hamster vimentin predicted from the nucleotide sequence. Proc Natl Acad Sci U S A. 1983 Jun;80(12):3548–3552. [Europe PMC free article] [Abstract] [Google Scholar]
  • Ramaekers FC, Puts JJ, Moesker O, Kant A, Huysmans A, Haag D, Jap PH, Herman CJ, Vooijs GP. Antibodies to intermediate filament proteins in the immunohistochemical identification of human tumours: an overview. Histochem J. 1983 Jul;15(7):691–713. [Abstract] [Google Scholar]
  • Ramaekers FC, Moesker O, Huysmans A, Schaart G, Westerhof G, Wagenaar SS, Herman CJ, Vooijs GP. Intermediate filament proteins in the study of tumor heterogeneity: an in-depth study of tumors of the urinary and respiratory tracts. Ann N Y Acad Sci. 1985;455:614–634. [Abstract] [Google Scholar]
  • Rost B, Sander C. Prediction of protein secondary structure at better than 70% accuracy. J Mol Biol. 1993 Jul 20;232(2):584–599. [Abstract] [Google Scholar]
  • Rost B, Sander C. Combining evolutionary information and neural networks to predict protein secondary structure. Proteins. 1994 May;19(1):55–72. [Abstract] [Google Scholar]
  • Sanger F, Coulson AR, Barrell BG, Smith AJ, Roe BA. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. [Abstract] [Google Scholar]
  • Schaart G, Pieper FR, Kuijpers HJ, Bloemendal H, Ramaekers FC. Baby hamster kidney (BHK-21/C13) cells can express striated muscle type proteins. Differentiation. 1991 Mar;46(2):105–115. [Abstract] [Google Scholar]
  • Skalli O, Ropraz P, Trzeciak A, Benzonana G, Gillessen D, Gabbiani G. A monoclonal antibody against alpha-smooth muscle actin: a new probe for smooth muscle differentiation. J Cell Biol. 1986 Dec;103(6 Pt 2):2787–2796. [Europe PMC free article] [Abstract] [Google Scholar]
  • Takahashi K, Hiwada K, Kokubu T. Vascular smooth muscle calponin. A novel troponin T-like protein. Hypertension. 1988 Jun;11(6 Pt 2):620–626. [Abstract] [Google Scholar]
  • Takeuchi K, Takahashi K, Abe M, Nishida W, Hiwada K, Nabeya T, Maruyama K. Co-localization of immunoreactive forms of calponin with actin cytoskeleton in platelets, fibroblasts, and vascular smooth muscle. J Biochem. 1991 Feb;109(2):311–316. [Abstract] [Google Scholar]
  • Tinsley JM, Blake DJ, Roche A, Fairbrother U, Riss J, Byth BC, Knight AE, Kendrick-Jones J, Suthers GK, Love DR, et al. Primary structure of dystrophin-related protein. Nature. 1992 Dec 10;360(6404):591–593. [Abstract] [Google Scholar]
  • Turner CE, Burridge K. Detection of metavinculin in human platelets using a modified talin overlay assay. Eur J Cell Biol. 1989 Jun;49(1):202–206. [Abstract] [Google Scholar]
  • Vaughan KT, Weber FE, Einheber S, Fischman DA. Molecular cloning of chicken myosin-binding protein (MyBP) H (86-kDa protein) reveals extensive homology with MyBP-C (C-protein) with conserved immunoglobulin C2 and fibronectin type III motifs. J Biol Chem. 1993 Feb 15;268(5):3670–3676. [Abstract] [Google Scholar]
  • Voorter C, Joos S, Bringuier PP, Vallinga M, Poddighe P, Schalken J, du Manoir S, Ramaekers F, Lichter P, Hopman A. Detection of chromosomal imbalances in transitional cell carcinoma of the bladder by comparative genomic hybridization. Am J Pathol. 1995 Jun;146(6):1341–1354. [Europe PMC free article] [Abstract] [Google Scholar]
  • Winder SJ, Hemmings L, Maciver SK, Bolton SJ, Tinsley JM, Davies KE, Critchley DR, Kendrick-Jones J. Utrophin actin binding domain: analysis of actin binding and cellular targeting. J Cell Sci. 1995 Jan;108(Pt 1):63–71. [Abstract] [Google Scholar]
  • Winkelmann JC, Chang JG, Tse WT, Scarpa AL, Marchesi VT, Forget BG. Full-length sequence of the cDNA for human erythroid beta-spectrin. J Biol Chem. 1990 Jul 15;265(20):11827–11832. [Abstract] [Google Scholar]
Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

Full text links 

Read article at publisher's site: https://doi.org/10.1083/jcb.134.2.401

Read article for free, from open access legal sources, via Unpaywall: https://rupress.org/jcb/article-pdf/134/2/401/1265922/401.pdfUnpaywall

Citations & impact 

Impact metrics

145
Citations
Jump to Citations
7
Data citations
Jump to Data

Citations of article over time

Alternative metrics

Altmetric item for https://www.altmetric.com/details/3558702
Altmetric
Discover the attention surrounding your research
https://www.altmetric.com/details/3558702

Smart citations by scite.ai
Smart citations by scite.ai include citation statements extracted from the full text of the citing article. The number of the statements may be higher than the number of citations provided by EuropePMC if one paper cites another multiple times or lower if scite has not yet processed some of the citing articles.
Explore citation contexts and check if this article has been supported or disputed.
https://scite.ai/reports/10.1083/jcb.134.2.401

Supporting
Mentioning
Contrasting
14
183
0

Article citations

Go to all (145) article citations

Other citations

Data 

Data behind the article

This data has been text mined from the article, or deposited into data resources.

BioStudies: supplemental material and supporting data

Similar Articles 

To arrive at the top five similar articles we use a word-weighted algorithm to compare words from the Title and Abstract of each citation.

Funding 

Funders who supported this work.

NHLBI NIH HHS (1)