Journal of Neurocytology

, Volume 32, Issue 5–8, pp 905–913 | Cite as

Activity dependent removal of agrin from synaptic basal lamina by matrix metalloproteinase 3



Agrin is a heparan sulfate proteoglycan, which plays an essential role in the development and maintenance of the neuromuscular junction. Agrin is a stable component of the synaptic basal lamina and strong evidence supports the hypothesis that agrin directs the formation of the postsynaptic apparatus, including aggregates of AChRs, and junctional folds. Changes in the distribution of agrin during synaptic remodeling, denervation and reinnervation reveal that agrin can be quickly and efficiently removed from the synaptic basal lamina in a regulated manner. In order to fully understand this mechanism we sought to identify those molecules that were responsible for the removal of agrin. Matrix Metalloproteinases (MMPs) were the most likely molecules since MMPs are involved in the regulation of the pericellular space, including the cleavage of matrix proteins. In particular, MMP3 has been shown to be effective in cleaving heparan sulfate proteoglycans. Antibodies to MMP3 recognize molecules concentrated in the extracellular matrix of perisynaptic Schwann cells. MMP3 specific phylogenic compounds reveal that active MMP3 is localized to the neuromuscular junction. Purified recombinant MMP3 can directly cleave agrin, and it can also remove agrin from synaptic basal lamina. MMP3 activity is itself regulated as activation of MMP3 is lost in denervated muscles. MMP3 null mutant mice have altered neuromuscular junction structure and function, with increased AChRs, junctional folds and agrin immunoreactivity. Altogether these results support the hypothesis that synaptic activity induces the activation of MMP3, and the activated MMP3 removes agrin from the synaptic basal lamina.


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  1. ASTROW, S. H., QIANG, H. & KO, C. P. (1998) Perisynaptic Schwann cells at neuromuscular junctions revealed by a novel monoclonal antibody. J. Neurocytol. 27(9), 66–681.Google Scholar
  2. BALICE-GORDON, R. J. & LICHTMAN, J. W. (1994) Long-term synapse loss induced by focal blockade of postsynaptic receptors. Nature 372(6506), 51–524.PubMedGoogle Scholar
  3. BURGESS, R. W., SKARNES, W. C. & SANES, J. R. (2000) Agrin isoforms with distinct amino termini: Differential expression, localization, and function. Journal of Cell Biology 151(1), 4–52.PubMedGoogle Scholar
  4. FERNS, M. J., CAMPANELLI, J. T., HOCH, W., SCHELLER, R. H. & HALL, Z. (1993) The ability of agrin to cluster AChRs depends on alternative splicing and on cell surface proteoglycans. Neuron 11(3), 49–502.PubMedGoogle Scholar
  5. GAUTAM, M., NOAKES, P. G., MOSCOSO, L., RUPP, F., SCHELLER, R. H., MERLIE, J. P. & SANES, J. R. (1996) Defective neuromuscular synaptogenesis in agrindeficient mutant mice. Cell 85(4), 52–535.PubMedGoogle Scholar
  6. GUERIN, C. W. & HOLLAND, P. C. (1995) Synthesis and secretion of matrix-degrading metalloproteases by human skeletal muscle satellite cells. Dev. Dyn. 202(1), 9–99.PubMedGoogle Scholar
  7. HERRERA, A. A., BANNER, L. R. & NAGAYA, N. (1990) Repeated, in vivo observation of frog neuromuscular junctions: Remodelling involves concurrent growth and retraction. Journal of Neurocytology 19(1), 8–99.PubMedGoogle Scholar
  8. HUGHES, C., MURPHY, G. & HARDINGHAM, T. E. (1991) Metalloproteinase digestion of cartilage proteoglycan. Pattern of cleavage by stromelysin and susceptibility to collagenase. Bichemical Journal 279(Pt 3), 73– 739.Google Scholar
  9. JONES, G., MOORE, C., HASHEMOLHOSSEINI, S. & BRENNER, H. R. (1999) Constitutively active MuSK is clustered in the absence of agrin and induces ectopic jostsynaptic-like membranes in skeletal muscle fibers. Journal of Neuroscience 19(9), 337–3383.PubMedGoogle Scholar
  10. KAHL, J. & CAMPANELLI, J. T. (2003) A role for the juxtamembrane domain of beta-dystroglycan in agrininduced acetylcholine receptor clustering. Journal of Neuroscience 23(2), 39–402.PubMedGoogle Scholar
  11. KHERIF, S., LAFUMA, C., DEHAUPAS, M., LACHKAR, S., FOURNIER, J. G., VERDIERE-SAHUQUE, M., FARDEAU, M. & ALAMEDDINE, H. S. (1999) Expression of matrix metalloproteinases 2 and 9 in regenerating skeletal muscle: A study in experimentally injured and mdx muscles. Dev. Biol. 205(1), 15–170.PubMedGoogle Scholar
  12. LEONARD, J. P. & SALPETER M. M. (1979) Agonistinduced myopathy at the neuromuscular junction is mediated by calcium. Journal of Cell Biology 82(3), 81– 819.PubMedGoogle Scholar
  13. LETINSKY, M. S., FISCHBECK, K. H. & MCMAHAN, U. J. (1976) Precision of reinnervation of original postsynaptic sites in frog muscle after a nerve crush. Journal of Neurocytology 5(6), 69–718.PubMedGoogle Scholar
  14. LIN, W., BURGESS, R. W., DOMINGUEZ, B., PFAFF, S. L., SANES, J. R. & LEE, K. F. (2001) Distinct roles of nerve and muscle in postsynaptic differentiation of the neuromuscular synapse. Nature 410, 105–1064.PubMedGoogle Scholar
  15. LIU, Y., FIELDS, R. D., FITZGERALD, S., FESTOFF, B. W. & NELSON, P. G. (1994) Proteolytic activity, synapse elimination, and the Hebb synapse. Journal of Neurobiology 25(3), 32–335.PubMedGoogle Scholar
  16. MARSHALL, L. M., SANES, J. R. & MCMAHAN, U. J. (1977) Reinnervation of original synaptic sites on muscle fiber basement membrane after disruption of the muscle cells. Proceedings of the National Acadamy of Science USA 77(7), 307–3077.Google Scholar
  17. MCMAHAN, U. J., HORTON, S. E., WERLE, M. J., HONIG, L. J., KRÖGER, S., RUEGG, M. A. & ESCHER, G. (1992). Agrin isoforms and their role in synaptogenesis. Current Opinions in Cell Biology 4, 86–874.Google Scholar
  18. NAGASE, H. & WOESSNER, J. F. (1999) Matrix metalloproteinases. Journal of Biological Chemistry 274, 2149–21494.PubMedGoogle Scholar
  19. NITKIN, R. M., SMITH, M. A., MAGILL, C., FALLON, J. R., YAO, Y. M., WALLACE, B. G. & MCMAHAN, U. J. (1987) Identification of agrin, a synaptic organizing protein fromTorpedo electric organ. Journal of Cell Biology 105(6 Pt 1), 247–2478.PubMedGoogle Scholar
  20. O'BRIEN, R. A., OSTBERG, A. J. & VRBOVA, G. (1978) Observations on the elimination of polyneuronal innervation in developing mammalian skeletal muscle. Journal of Physiology 282, 57–582.PubMedGoogle Scholar
  21. REIST, N. E., MAGILL, C. & MCMAHAN, U. J. (1987) Agrin-like molecules at synaptic sites in normal, denervated, and damaged skeletal muscles. Journal of Cell Biology 105(6 Pt 1), 245–2469.PubMedGoogle Scholar
  22. REIST, N. E. & SMITH, S. J. (1992) Neurally evoked calcium transients in terminal Schwann cells at the neuromuscular junction. Proceedings of the National Academy of Sciences USA 89(16), 762–7659.Google Scholar
  23. REUNANEN, N., LI, S. P., AHONEN, M., FOSCHI, M., HAN, J. & KAHARI, V. M. (2002) Activation of p38 alpha MAPK enhances collagenase-1 (matrix metalloproteinase (MMP)-1) and stromelysin-1 (MMP3) expression by mRNA stabilization. Journal of Biological Chemistry 277(35), 3236–32368.PubMedGoogle Scholar
  24. SCHOSER, B. G. & BLOTTNER, D. (1999) Matrix metalloproteinases MMP-2, MMP-7 and MMP-9 in denervated human muscle. Neuroreport. 10(13), 279–2797.PubMedGoogle Scholar
  25. STANCO, A. M. & WERLE, M. J. (1997) Agrin and acetylcholine receptors are removed from abandoned synaptic sites at reinnervated frog neuromuscular junctions. Journal of Neurobiology 33, 99–1018.PubMedGoogle Scholar
  26. STANCO, A. M. & WERLE, M. J. (1998) Agrin and acetylcholine receptor distribution following electrical stimulation. Muscle and Nerve 21, 40–409.PubMedGoogle Scholar
  27. STERNLICHT, M. D. & WERB, Z. (2001) How matrix metalloproteinases regulate cell behavior. Annual Reviews in Cell Developmental Biology 17, 46–516.Google Scholar
  28. VANSAUN, M. & WERLE, M. J. (2000) Matrix metalloproteinase-3 removes agrin from synaptic basal lamina. Journal of Neurobiology 43, 14–149.PubMedGoogle Scholar
  29. VANSAUN, M., HERRERA, A. A. & WERLE, M. J. (2004) Structural alterations at the neuromuscular junctions of matrix metalloproteinase 3 null mutant mice. J. Neurosci. In Press.Google Scholar
  30. VANSAUN, M. & WERLE, M. J. (2002) Activity of matrix metalloproteinase 3 during periods of neuromuscular junction remodeling. Society for Neuroscience Abstracts 32, 629.7.Google Scholar
  31. WALLACE, B. G. (1988) Regulation of agrin-induced acetylcholine receptor aggregation by Ca++ and phorbol ester. Journal of Cell Biology 107(1), 26–278.PubMedGoogle Scholar
  32. WALSH, M. K. & LICHTMAN, J. W. (2003) In vivo timelapse imaging of synaptic takeover associated with naturally occurring synapse elimination. Neuron 37(1), 6–73.PubMedGoogle Scholar
  33. WERLE, M. J. & SOJKA, A. M. (1996) Anti-agrin staining is absent in empty gutters of frog neuromuscular junctions. Journal of Neurobiology 30, 29–302.PubMedGoogle Scholar
  34. WERLE, M. J., JONES, M. A. & STANCO, A. M. (1999) Agrin produced by Schwann cells does not induce the aggregation of AChRs at the frog neuromuscular junction. Journal of Neurobiology 40, 4–54.PubMedGoogle Scholar

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© Kluwer Academic Publishers 2003

Authors and Affiliations

  1. 1.Department of Anatomy and Cell BiologyUniversity of Kansas Medical CenterKansas CityUSA

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