Regular Article
Structural Features of Collapsin Required for Biological Activity and Distribution of Binding Sites in the Developing Chick

https://doi.org/10.1006/mcne.1997.0636Get rights and content

Abstract

We have used Fc-chimeras of collapsin-1/Sema III to study the structure–function activity of this recently identified repulsive axonal guidance molecule and to map the distribution of its binding sites during chick development. Our results show that the biological activity of the collapsin-Fc in anin vitrocollapse assay is independent of both the Ig-domain and the positive charged carboxy terminus. Collapsin binding sites were found on a number of neuronal fiber tracts, and in two instances (DRG tracts and the retinotectal projection) this expression is both highly dynamic and consistent with them playing a role in axonal growth and guidance. Collapsin-1 binding sites were also found on a number of nonneuronal structures that do not produce collapsin-1 mRNA. We postulate that these sites may act to localize or concentrate collapsin-1 released from growing axons and in this way allow for an autocrine axonal guidance mechanism to function during development.

References (41)

  • E.K. Messersmith et al.

    Semaphorin III can function as a selective chemorepellent to pattern sensory projections in the spinal cord

    Neuron

    (1995)
  • L. Puelles et al.

    Differentiation of neuroblasts in the chick optic tectum up to eight days of incubation: A Golgi study

    Neuroscience

    (1978)
  • I. Shepherd et al.

    The distribution of collapsin-1 mRNA in the developing chick nervous system

    Dev. Biol.

    (1996)
  • O. Behar et al.

    Semaphorin III is needed for normal patterning and growth of nerves, bones and heart

    Nature

    (1996)
  • B.M. Davis et al.

    Development of central projections of lumbosacral sensory neurons in the chick

    J. Comp. Neurol.

    (1989)
  • R.G. DeLong et al.

    Development of the retinotectal topographic projection in the chick embryo

    Exp. Neurol.

    (1965)
  • J. Fawcett et al.

    Molecular tools for the analysis of cell adhesion

    Cell Adhes. Com.

    (1994)
  • S. Goldberg

    Studies on the mechanics of development of the visual pathway in the chick embryo

    Dev. Biol.

    (1974)
  • C.S. Goodman

    Mechanism and molecules that control growth cone guidance

    Annu. Rev. Neurosci.

    (1996)
  • A. Inoue et al.

    Lamina-specific connectivity in the brain: Regulation by N-cadherin, neurotrophins, and glycoconjugates

    Science

    (1997)
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