The Journal of Physiological Sciences

, Volume 60, Issue 5, pp 343–352 | Cite as

Skeletal muscle expression of bone morphogenetic protein-1 and tolloid-like-1 extracellular proteases in different fiber types and in response to unloading, food deprivation and differentiation

  • David L. AllenEmail author
  • Bradley J. Greyback
  • Andrea M. Hanson
  • Allison S. Cleary
  • Sarah F. Lindsay
Original Paper


Members of the bone morphogenetic protein-1/mammalian tolloid (BMP-1/mTLD) family of proteases cleave diverse extracellular proteins, including the growth inhibitor myostatin. The purpose of this work was to examine the expression of BMP-1/mTLD, tolloid-like-1 and -2 (TLL1 and TLL2) in hindlimb muscles of the mouse in vivo and in C2C12 muscle cells in vitro. Quantitative real-time polymerase chain reaction revealed that neither BMP-1/mTLD nor TLL1 mRNA levels differed between the predominantly fast-twitch tibialis anterior (TA) and gastrocnemius (GAST) muscles and the more slow-twitch soleus (SOL) muscle; TLL2 mRNA levels were not detectable in any of the muscles examined. Interestingly, however, immunohistochemical analysis revealed that BMP-1 protein was expressed in type I and IIa but not in IIb fibers. TLL1 mRNA levels significantly increased in the TA but not the SOL with 3 days of hindlimb suspension and significantly decreased in both TA and SOL in response to 2 days of food deprivation. In contrast, BMP-1/mTLD mRNA levels were unaffected in either muscle by either condition. In addition, BMP-1/mTLD and TLL1 mRNA levels significantly decreased during C2C12 myoblast differentiation in vitro, and activity of a 1,200-bp mouse TLL1 promoter construct was significantly decreased in C2C12 myotubes by differentiation, by mutation of an nuclear factor kappa-beta (NF-kappaB) site, or deletion of a sma/mothers against decapentaplegic (SMAD) site. Together, these data demonstrate that TLL1 mRNA levels are altered by loading, energy status, and differentiation, and thus its expression may be regulated so as to modulate activity of myostatin or other extracellular substrates during these adaptive states.


Myostatin Fast-twitch Slow-twitch Food deprivation Hindlimb suspension C2C12 



This work was partially supported by K01 grant AR0505-01 from the National Institutes of Health, and by two University of Colorado Innovative Seed Grants.


  1. 1.
    Hopkins DR, Keles S, Greenspan DS (2007) The bone morphogenetic protein 1/Tolloid-like metalloproteinases. Matrix Biol 26:508–523CrossRefPubMedGoogle Scholar
  2. 2.
    Ge G, Greenspan DS (2006) Developmental roles of the BMP1 metalloproteinases. Birth Defects Res C Embryo Today 78:47–68CrossRefPubMedGoogle Scholar
  3. 3.
    Wolfman NM, McPherron AC, Pappano WN, Davies MV, Song K, Tomkinson KN, Wright JF, Zhao L, Sebald SM, Greenspan DS, Lee SJ (2003) Activation of latent myostatin by the BMP-1/tolloid family of metalloproteinases. Proc Natl Acad Sci USA 100:15842–15846CrossRefPubMedGoogle Scholar
  4. 4.
    Lee SJ (2008) Genetic analysis of the role of proteolysis in the activation of latent myostatin. PLoS One 3:e1628CrossRefPubMedGoogle Scholar
  5. 5.
    Fukagawa M, Suzuki N, Hogan BL, Jones CM (1994) Embryonic expression of mouse bone morphogenetic protein-1 (BMP-1), which is related to the Drosophila dorsoventral gene tolloid and encodes a putative astacin metalloendopeptidase. Dev Biol 163:175–183CrossRefPubMedGoogle Scholar
  6. 6.
    Takahara K, Lyons GE, Greenspan DS (1994) Bone morphogenetic protein-1 and a mammalian tolloid homologue (mTld) are encoded by alternatively spliced transcripts which are differentially expressed in some tissues. J Biol Chem 269:32572–32578PubMedGoogle Scholar
  7. 7.
    Takahara K, Brevard R, Hoffman GG, Suzuki N, Greenspan DS (1996) Characterization of a novel gene product (mammalian tolloid-like) with high sequence similarity to mammalian tolloid/bone morphogenetic protein-1. Genomics 34:157–165CrossRefPubMedGoogle Scholar
  8. 8.
    Scott IC, Blitz IL, Pappano WN, Imamura Y, Clark TG, Steiglitz BM, Thomas CL, Maas SA, Takahara K, Cho KW, Greenspan DS (1999) Mammalian BMP-1/tolloid-related metalloproteinases, including novel family member mammalian Tolloid-like 2, have differential enzymatic activities and distributions of expression relevant to patterning and skeletogenesis. Dev Biol 213:283–300CrossRefPubMedGoogle Scholar
  9. 9.
    Allen DL, Linderman JK, Roy RR, Grindeland RE, Mukku V, Edgerton VR (1997) Growth hormone/IGF-I and/or resistive exercise maintains myonuclear number in hindlimb unweighted muscles. J Appl Physiol 83:1857–1861PubMedGoogle Scholar
  10. 10.
    Allen DL, Linderman JK, Roy RR, Bigbee AJ, Grindeland RE, Mukku V, Edgerton VR (1997) Apoptosis: a mechanism contributing to remodeling of skeletal muscle in response to hindlimb unweighting. Am J Physiol 273:C579–C587PubMedGoogle Scholar
  11. 11.
    Allen DL, Bandstra ER, Harrison BC, Thorng S, Stodieck LS, Kostenuik PJ, Morony S, Lacey DL, Hammond TG, Leinwand LL, Argraves WS, Bateman TA, Barth JL (2009) Effects of spaceflight on murine skeletal muscle gene expression. J Appl Physiol 106:582–595CrossRefPubMedGoogle Scholar
  12. 12.
    Allen DL, Unterman TG (2007) Regulation of myostatin expression and myoblast differentiation by FoxO and SMAD transcription factors. Am J Physiol Cell Physiol 292:C188–C199CrossRefPubMedGoogle Scholar
  13. 13.
    Allen DL, Cleary AS, Speaker KJ, Lindsay SF, Uyenishi J, Reed JM, Madden MC, Mehan RS (2008) Myostatin, activin receptor IIb, and follistatin-like-3 gene expression are altered in adipose tissue and skeletal muscle of obese mice. Am J Physiol Endocrinol Metab 294:E918–E927CrossRefPubMedGoogle Scholar
  14. 14.
    Carlson CJ, Booth FW, Gordon SE (1999) Skeletal muscle myostatin mRNA expression is fiber-type specific and increases during hindlimb unloading. Am J Physiol 277:R601–R606PubMedGoogle Scholar
  15. 15.
    Sakuma K, Watanabe K, Sano M, Uramoto I, Totsuka T (2000) Differential adaptation of growth and differentiation factor 8/myostatin, fibroblast growth factor 6 and leukemia inhibitory factor in overloaded, regenerating and denervated rat muscles. Biochim Biophys Acta 1497:77–88CrossRefPubMedGoogle Scholar
  16. 16.
    Salerno MS, Thomas M, Forbes D, Watson T, Kambadur R, Sharma M (2004) Molecular analysis of fiber type-specific expression of murine myostatin promoter. Am J Physiol Cell Physiol 287:C1031–C1040CrossRefPubMedGoogle Scholar
  17. 17.
    Sakuma K, Watanabe K, Sano M, Kitajima S, Sakamoto K, Uramoto I, Totsuka T (2000) The adaptive response of transforming growth factor-beta 2 and -beta RII in the overloaded, regenerating and denervated muscles of rats. Acta Neuropathol 299:177–185CrossRefGoogle Scholar
  18. 18.
    Lecker SH, Jagoe RT, Gilbert A, Gomes M, Baracos V, Bailey J, Price SR, Mitch WE, Goldberg AL (2004) Multiple types of skeletal muscle atrophy involve a common program of changes in gene expression. FASEB J 18:39–51CrossRefPubMedGoogle Scholar
  19. 19.
    Sacheck JM, Hyatt JP, Raffaello A, Jagoe RT, Roy RR, Edgerton VR, Lecker SH, Goldberg AL (2007) Rapid disuse and denervation atrophy involve transcriptional changes similar to those of muscle wasting during systemic diseases. FASEB J 21:140–155CrossRefPubMedGoogle Scholar
  20. 20.
    Ríos R, Carneiro I, Arce VM, Devesa J (2001) Myostatin regulates cell survival during C2C12 myogenesis. Biochem Biophys Res Commun 280:561–566CrossRefPubMedGoogle Scholar
  21. 21.
    Mendler L, Zádor E, Ver Heyen M, Dux L, Wuytack F (2000) Myostatin levels in regenerating rat muscles and in myogenic cell cultures. J Muscle Res Cell Motil 21:551–563CrossRefPubMedGoogle Scholar
  22. 22.
    Ma K, Mallidis C, Artaza J, Taylor W, Gonzalez-Cadavid N, Bhasin S (2001) Characterization of 5′-regulatory region of human myostatin gene: regulation by dexamethasone in vitro. Am J Physiol Endocrinol Metab 281:E1128–E1136PubMedGoogle Scholar
  23. 23.
    Kaltschmidt B, Baeuerle PA, Kaltschmidt C (1993) Potential involvement of the transcription factor NF-kappa B in neurological disorders. Mol Aspects Med 14:171–190CrossRefPubMedGoogle Scholar
  24. 24.
    Tamura G, Olson D, Miron J, Clark TG (2005) Tolloid-like 1 is negatively regulated by stress and glucocorticoids. Brain Res Mol Brain Res 142:81–90CrossRefPubMedGoogle Scholar

Copyright information

© The Physiological Society of Japan and Springer 2010

Authors and Affiliations

  • David L. Allen
    • 1
    Email author
  • Bradley J. Greyback
    • 2
  • Andrea M. Hanson
    • 2
  • Allison S. Cleary
    • 1
  • Sarah F. Lindsay
    • 1
  1. 1.Department of Integrative PhysiologyUniversity of ColoradoBoulderUSA
  2. 2.Department of Aerospace EngineeringUniversity of ColoradoBoulderUSA

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