Abstract
Leech neurons in culture sprout rapidly when attached to extracts from connective tissue surrounding the nervous system. Laminin-like molecules that promote sprouting have now been isolated from this extracellular matrix. Two mAbs have been prepared that react on immunoblots with a approximately equal to 220- and a approximately equal to 340-kD polypeptide, respectively. These antibodies have been used to purify molecules with cross-shaped structures in the electron microscope. The molecules, of approximately equal to 10(3) kD on nonreducing SDS gels, have subunits of approximately equal to 340, 220, and 160-180 kD. Attachment to the laminin-like molecules was sufficient to initiate sprouting by single isolated leech neurons in defined medium. This demonstrates directly a function for a laminin-related invertebrate protein. The mAbs directed against the approximately equal to 220-kD chains of the laminin-like leech molecule labeled basement membrane extracellular matrix in leech ganglia and nerves. A polyclonal antiserum against the approximately equal to 220-kD polypeptide inhibited neurite outgrowth. Vertebrate laminin did not mediate the sprouting of leech neurons; similarly, the leech molecule was an inert substrate for vertebrate neurons. Although some traits of structure, function, and distribution are conserved between vertebrate laminin and the invertebrate molecule, our results suggest that the functional domains differ.
Full Text
The Full Text of this article is available as a PDF (4.1 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- COGGESHALL R. E., FAWCETT D. W. THE FINE STRUCTURE OF THE CENTRAL NERVOUS SYSTEM OF THE LEECH, HIRUDO MEDICINALIS. J Neurophysiol. 1964 Mar;27:229–289. doi: 10.1152/jn.1964.27.2.229. [DOI] [PubMed] [Google Scholar]
- Chiquet M., Acklin S. E. Attachment of Con A or extracellular matrix initiates rapid sprouting by cultured leech neurons. Proc Natl Acad Sci U S A. 1986 Aug;83(16):6188–6192. doi: 10.1073/pnas.83.16.6188. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chiquet M., Fambrough D. M. Chick myotendinous antigen. I. A monoclonal antibody as a marker for tendon and muscle morphogenesis. J Cell Biol. 1984 Jun;98(6):1926–1936. doi: 10.1083/jcb.98.6.1926. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chiquet M., Nicholls J. G. Neurite outgrowth and synapse formation by identified leech neurones in culture. J Exp Biol. 1987 Sep;132:191–206. doi: 10.1242/jeb.132.1.191. [DOI] [PubMed] [Google Scholar]
- Dietzel I. D., Drapeau P., Nicholls J. G. Voltage dependence of 5-hydroxytryptamine release at a synapse between identified leech neurones in culture. J Physiol. 1986 Mar;372:191–205. doi: 10.1113/jphysiol.1986.sp016004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edgar D., Timpl R., Thoenen H. Structural requirements for the stimulation of neurite outgrowth by two variants of laminin and their inhibition by antibodies. J Cell Biol. 1988 Apr;106(4):1299–1306. doi: 10.1083/jcb.106.4.1299. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edgar D., Timpl R., Thoenen H. The heparin-binding domain of laminin is responsible for its effects on neurite outgrowth and neuronal survival. EMBO J. 1984 Jul;3(7):1463–1468. doi: 10.1002/j.1460-2075.1984.tb01997.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Engel J., Furthmayr H. Electron microscopy and other physical methods for the characterization of extracellular matrix components: laminin, fibronectin, collagen IV, collagen VI, and proteoglycans. Methods Enzymol. 1987;145:3–78. doi: 10.1016/0076-6879(87)45003-9. [DOI] [PubMed] [Google Scholar]
- Engel J., Odermatt E., Engel A., Madri J. A., Furthmayr H., Rohde H., Timpl R. Shapes, domain organizations and flexibility of laminin and fibronectin, two multifunctional proteins of the extracellular matrix. J Mol Biol. 1981 Jul 25;150(1):97–120. doi: 10.1016/0022-2836(81)90326-0. [DOI] [PubMed] [Google Scholar]
- Engvall E. Enzyme immunoassay ELISA and EMIT. Methods Enzymol. 1980;70(A):419–439. doi: 10.1016/s0076-6879(80)70067-8. [DOI] [PubMed] [Google Scholar]
- Fessler L. I., Campbell A. G., Duncan K. G., Fessler J. H. Drosophila laminin: characterization and localization. J Cell Biol. 1987 Nov;105(5):2383–2391. doi: 10.1083/jcb.105.5.2383. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Halfter W., Deiss S. Axonal pathfinding in organ-cultured embryonic avian retinae. Dev Biol. 1986 Apr;114(2):296–310. doi: 10.1016/0012-1606(86)90194-6. [DOI] [PubMed] [Google Scholar]
- Hauri H. P., Sterchi E. E., Bienz D., Fransen J. A., Marxer A. Expression and intracellular transport of microvillus membrane hydrolases in human intestinal epithelial cells. J Cell Biol. 1985 Sep;101(3):838–851. doi: 10.1083/jcb.101.3.838. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hynes R. O. Integrins: a family of cell surface receptors. Cell. 1987 Feb 27;48(4):549–554. doi: 10.1016/0092-8674(87)90233-9. [DOI] [PubMed] [Google Scholar]
- Imhof B. A., Marti U., Boller K., Frank H., Birchmeier W. Association between coated vesicles and microtubules. Exp Cell Res. 1983 Apr 15;145(1):199–207. doi: 10.1016/s0014-4827(83)80021-4. [DOI] [PubMed] [Google Scholar]
- Kramer A. P., Stent G. S. Developmental arborization of sensory neurons in the leech Haementeria ghilianii. II. Experimentally induced variations in the branching pattern. J Neurosci. 1985 Mar;5(3):768–775. doi: 10.1523/JNEUROSCI.05-03-00768.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Liesi P. Do neurons in the vertebrate CNS migrate on laminin? EMBO J. 1985 May;4(5):1163–1170. doi: 10.1002/j.1460-2075.1985.tb03755.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Liesi P. Laminin-immunoreactive glia distinguish regenerative adult CNS systems from non-regenerative ones. EMBO J. 1985 Oct;4(10):2505–2511. doi: 10.1002/j.1460-2075.1985.tb03963.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Macagno E. R., Muller K. J., DeRiemer S. A. Regeneration of axons and synaptic connections by touch sensory neurons in the leech central nervous system. J Neurosci. 1985 Sep;5(9):2510–2521. doi: 10.1523/JNEUROSCI.05-09-02510.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Martin G. R., Timpl R. Laminin and other basement membrane components. Annu Rev Cell Biol. 1987;3:57–85. doi: 10.1146/annurev.cb.03.110187.000421. [DOI] [PubMed] [Google Scholar]
- McCarthy R. A., Beck K., Burger M. M. Laminin is structurally conserved in the sea urchin basal lamina. EMBO J. 1987 Jun;6(6):1587–1593. doi: 10.1002/j.1460-2075.1987.tb02404.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Naidet C., Sémériva M., Yamada K. M., Thiery J. P. Peptides containing the cell-attachment recognition signal Arg-Gly-Asp prevent gastrulation in Drosophila embryos. Nature. 1987 Jan 22;325(6102):348–350. doi: 10.1038/325348a0. [DOI] [PubMed] [Google Scholar]
- Paulsson M., Aumailley M., Deutzmann R., Timpl R., Beck K., Engel J. Laminin-nidogen complex. Extraction with chelating agents and structural characterization. Eur J Biochem. 1987 Jul 1;166(1):11–19. doi: 10.1111/j.1432-1033.1987.tb13476.x. [DOI] [PubMed] [Google Scholar]
- Paulsson M., Deutzmann R., Timpl R., Dalzoppo D., Odermatt E., Engel J. Evidence for coiled-coil alpha-helical regions in the long arm of laminin. EMBO J. 1985 Feb;4(2):309–316. doi: 10.1002/j.1460-2075.1985.tb03630.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ruoslahti E., Pierschbacher M. D. New perspectives in cell adhesion: RGD and integrins. Science. 1987 Oct 23;238(4826):491–497. doi: 10.1126/science.2821619. [DOI] [PubMed] [Google Scholar]
- Shotton D. M., Burke B. E., Branton D. The molecular structure of human erythrocyte spectrin. Biophysical and electron microscopic studies. J Mol Biol. 1979 Jun 25;131(2):303–329. doi: 10.1016/0022-2836(79)90078-0. [DOI] [PubMed] [Google Scholar]
- Stuart D. K., Blair S. S., Weisblat D. A. Cell lineage, cell death, and the developmental origin of identified serotonin- and dopamine-containing neurons in the leech. J Neurosci. 1987 Apr;7(4):1107–1122. doi: 10.1523/JNEUROSCI.07-04-01107.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Timpl R., Wiedemann H., van Delden V., Furthmayr H., Kühn K. A network model for the organization of type IV collagen molecules in basement membranes. Eur J Biochem. 1981 Nov;120(2):203–211. doi: 10.1111/j.1432-1033.1981.tb05690.x. [DOI] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vaughan L., Huber S., Chiquet M., Winterhalter K. H. A major, six-armed glycoprotein from embryonic cartilage. EMBO J. 1987 Feb;6(2):349–353. doi: 10.1002/j.1460-2075.1987.tb04761.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wewer U. M., Tichy D., Damjanov A., Paulsson M., Damjanov I. Distinct antigenic characteristics of murine parietal yolk sac laminin. Dev Biol. 1987 Jun;121(2):397–407. doi: 10.1016/0012-1606(87)90176-x. [DOI] [PubMed] [Google Scholar]
- de StGroth S. F., Scheidegger D. Production of monoclonal antibodies: strategy and tactics. J Immunol Methods. 1980;35(1-2):1–21. doi: 10.1016/0022-1759(80)90146-5. [DOI] [PubMed] [Google Scholar]
