Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1995 Jun 1;129(5):1355–1362. doi: 10.1083/jcb.129.5.1355

Calsensin: a novel calcium-binding protein expressed in a subset of peripheral leech neurons fasciculating in a single axon tract

PMCID: PMC2120460  PMID: 7775579

Abstract

The mAb lan3-6 recognizes a cytosolic antigen which is selectively expressed in the growth cones and axons of a small subset of peripheral sensory neurons fasciculating in a single tract common to all hirudinid leeches. We have used this antibody to clone a novel EF-hand calcium- binding protein, calsensin, by screening an expression vector library. A full-length clone of 1.1 kb identified by the antibody was isolated and sequenced. In situ hybridizations with calsensin probes and antibody staining using new polyclonal antisera generated against calsensin sequence demonstrate that calsensin indeed corresponds to the lan3-6 antigen. Calsensin consists of 83 residues with a calculated molecular mass of 9.1 kD that contains two helix-loop-helix domains. The calcium-binding domains are likely to be functional in vivo since a fusion protein derived from the calsensin clone binds 45Ca2+ in vitro. Immunoaffinity purification experiments with the lan3-6 antibody shows that a large 200,000 M(r) protein selectively copurifies with calsensin in two different leech species. These results suggest that calsensin may be functioning as a trigger protein which interacts with the larger protein. These data are consistent with the hypothesis that calsensin may mediate calcium-dependent signal transduction events in the growth cones and axons of this small group of sensory neurons which fasciculate in a single axon tract.

Full Text

The Full Text of this article is available as a PDF (1.7 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Atashi J. R., Klinz S. G., Ingraham C. A., Matten W. T., Schachner M., Maness P. F. Neural cell adhesion molecules modulate tyrosine phosphorylation of tubulin in nerve growth cone membranes. Neuron. 1992 May;8(5):831–842. doi: 10.1016/0896-6273(92)90197-l. [DOI] [PubMed] [Google Scholar]
  3. Baimbridge K. G., Celio M. R., Rogers J. H. Calcium-binding proteins in the nervous system. Trends Neurosci. 1992 Aug;15(8):303–308. doi: 10.1016/0166-2236(92)90081-i. [DOI] [PubMed] [Google Scholar]
  4. Bandtlow C. E., Schmidt M. F., Hassinger T. D., Schwab M. E., Kater S. B. Role of intracellular calcium in NI-35-evoked collapse of neuronal growth cones. Science. 1993 Jan 1;259(5091):80–83. doi: 10.1126/science.8418499. [DOI] [PubMed] [Google Scholar]
  5. Briggs K. K., Johansen K. M., Johansen J. Selective pathway choice of a single central axonal fascicle by a subset of peripheral neurons during leech development. Dev Biol. 1993 Aug;158(2):380–389. doi: 10.1006/dbio.1993.1196. [DOI] [PubMed] [Google Scholar]
  6. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dizhoor A. M., Ray S., Kumar S., Niemi G., Spencer M., Brolley D., Walsh K. A., Philipov P. P., Hurley J. B., Stryer L. Recoverin: a calcium sensitive activator of retinal rod guanylate cyclase. Science. 1991 Feb 22;251(4996):915–918. doi: 10.1126/science.1672047. [DOI] [PubMed] [Google Scholar]
  8. Doherty P., Ashton S. V., Moore S. E., Walsh F. S. Morphoregulatory activities of NCAM and N-cadherin can be accounted for by G protein-dependent activation of L- and N-type neuronal Ca2+ channels. Cell. 1991 Oct 4;67(1):21–33. doi: 10.1016/0092-8674(91)90569-k. [DOI] [PubMed] [Google Scholar]
  9. Fernández J., Stent G. S. Embryonic development of the hirudinid leech Hirudo medicinalis: structure, development and segmentation of the germinal plate. J Embryol Exp Morphol. 1982 Dec;72:71–96. [PubMed] [Google Scholar]
  10. Goodman C. S., Shatz C. J. Developmental mechanisms that generate precise patterns of neuronal connectivity. Cell. 1993 Jan;72 (Suppl):77–98. doi: 10.1016/s0092-8674(05)80030-3. [DOI] [PubMed] [Google Scholar]
  11. Gumbiner B. M. Proteins associated with the cytoplasmic surface of adhesion molecules. Neuron. 1993 Oct;11(4):551–564. doi: 10.1016/0896-6273(93)90068-3. [DOI] [PubMed] [Google Scholar]
  12. Heizmann C. W., Braun K. Changes in Ca(2+)-binding proteins in human neurodegenerative disorders. Trends Neurosci. 1992 Jul;15(7):259–264. doi: 10.1016/0166-2236(92)90067-i. [DOI] [PubMed] [Google Scholar]
  13. Hynes R. O. Integrins: versatility, modulation, and signaling in cell adhesion. Cell. 1992 Apr 3;69(1):11–25. doi: 10.1016/0092-8674(92)90115-s. [DOI] [PubMed] [Google Scholar]
  14. Jessell T. M. Adhesion molecules and the hierarchy of neural development. Neuron. 1988 Mar;1(1):3–13. doi: 10.1016/0896-6273(88)90204-8. [DOI] [PubMed] [Google Scholar]
  15. Johansen J., Johansen K. M., Briggs K. K., Kopp D. M., Jellies J. Hierarchical guidance cues and selective axon pathway formation. Prog Brain Res. 1994;103:109–120. doi: 10.1016/s0079-6123(08)61131-0. [DOI] [PubMed] [Google Scholar]
  16. Kobayashi T., Takagi T., Konishi K., Cox J. A. The primary structure of a new Mr 18,000 calcium vector protein from amphioxus. J Biol Chem. 1987 Feb 25;262(6):2613–2623. [PubMed] [Google Scholar]
  17. Kozak M. Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell. 1986 Jan 31;44(2):283–292. doi: 10.1016/0092-8674(86)90762-2. [DOI] [PubMed] [Google Scholar]
  18. Kretsinger R. H. Structure and evolution of calcium-modulated proteins. CRC Crit Rev Biochem. 1980;8(2):119–174. doi: 10.3109/10409238009105467. [DOI] [PubMed] [Google Scholar]
  19. Levine B. A., Dalgarno D. C. The dynamics and function of calcium-binding proteins. Biochim Biophys Acta. 1983 Sep 15;726(3):187–204. doi: 10.1016/0304-4173(83)90005-8. [DOI] [PubMed] [Google Scholar]
  20. Lin C. S., Shen W., Chen Z. P., Tu Y. H., Matsudaira P. Identification of I-plastin, a human fimbrin isoform expressed in intestine and kidney. Mol Cell Biol. 1994 Apr;14(4):2457–2467. doi: 10.1128/mcb.14.4.2457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Macagno E. R., Stewart R. R., Zipser B. The expression of antigens by embryonic neurons and glia in segmental ganglia of the leech Haemopis marmorata. J Neurosci. 1983 Sep;3(9):1746–1759. doi: 10.1523/JNEUROSCI.03-09-01746.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Maruyama K., Mikawa T., Ebashi S. Detection of calcium binding proteins by 45Ca autoradiography on nitrocellulose membrane after sodium dodecyl sulfate gel electrophoresis. J Biochem. 1984 Feb;95(2):511–519. doi: 10.1093/oxfordjournals.jbchem.a134633. [DOI] [PubMed] [Google Scholar]
  23. Nemoto Y., Ikeda J., Katoh K., Koshimoto H., Yoshihara Y., Mori K. R2D5 antigen: a calcium-binding phosphoprotein predominantly expressed in olfactory receptor neurons. J Cell Biol. 1993 Nov;123(4):963–976. doi: 10.1083/jcb.123.4.963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Palczewski K., Subbaraya I., Gorczyca W. A., Helekar B. S., Ruiz C. C., Ohguro H., Huang J., Zhao X., Crabb J. W., Johnson R. S. Molecular cloning and characterization of retinal photoreceptor guanylyl cyclase-activating protein. Neuron. 1994 Aug;13(2):395–404. doi: 10.1016/0896-6273(94)90355-7. [DOI] [PubMed] [Google Scholar]
  25. Persechini A., Moncrief N. D., Kretsinger R. H. The EF-hand family of calcium-modulated proteins. Trends Neurosci. 1989 Nov;12(11):462–467. doi: 10.1016/0166-2236(89)90097-0. [DOI] [PubMed] [Google Scholar]
  26. Schuch U., Lohse M. J., Schachner M. Neural cell adhesion molecules influence second messenger systems. Neuron. 1989 Jul;3(1):13–20. doi: 10.1016/0896-6273(89)90111-6. [DOI] [PubMed] [Google Scholar]
  27. Shure M., Wessler S., Fedoroff N. Molecular identification and isolation of the Waxy locus in maize. Cell. 1983 Nov;35(1):225–233. doi: 10.1016/0092-8674(83)90225-8. [DOI] [PubMed] [Google Scholar]
  28. Strynadka N. C., James M. N. Crystal structures of the helix-loop-helix calcium-binding proteins. Annu Rev Biochem. 1989;58:951–998. doi: 10.1146/annurev.bi.58.070189.004511. [DOI] [PubMed] [Google Scholar]
  29. Szebenyi D. M., Moffat K. The refined structure of vitamin D-dependent calcium-binding protein from bovine intestine. Molecular details, ion binding, and implications for the structure of other calcium-binding proteins. J Biol Chem. 1986 Jul 5;261(19):8761–8777. [PubMed] [Google Scholar]
  30. Tautz D., Pfeifle C. A non-radioactive in situ hybridization method for the localization of specific RNAs in Drosophila embryos reveals translational control of the segmentation gene hunchback. Chromosoma. 1989 Aug;98(2):81–85. doi: 10.1007/BF00291041. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. Yamagata K., Goto K., Kuo C. H., Kondo H., Miki N. Visinin: a novel calcium binding protein expressed in retinal cone cells. Neuron. 1990 Mar;4(3):469–476. doi: 10.1016/0896-6273(90)90059-o. [DOI] [PubMed] [Google Scholar]
  33. Yamanaka M. K., Saugstad J. A., Hanson-Painton O., McCarthy B. J., Tobin S. L. Structure and expression of the Drosophila calmodulin gene. Nucleic Acids Res. 1987 Apr 24;15(8):3335–3348. doi: 10.1093/nar/15.8.3335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zipser B., McKay R. Monoclonal antibodies distinguish identifiable neurones in the leech. Nature. 1981 Feb 12;289(5798):549–554. doi: 10.1038/289549a0. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES