Abstract

We have completed all of the aims of the original proposal, as well as discovering brain protein 4.1, demonstrating the identity of brain 4.1 as synapsin I, and beginning the study of brain spectrin expression in mouse neuroblastoma cells and primary cultures of dorsal root ganglion neurons. In this application we describe experiments which are designed to give us a better understanding of brain spectrin structure, function, location, and interaction with synapsin I.
The aims of this proposal are (1) to determine the domain structure of brain spectrin by combining peptide mapping with monoclonal antibody technology; (2) Quantitate the number of phosphate groups on the 235k Dal Beta subunit of brain spectrin. Localize these phosphates within the brain spectrin domain structure, and determine the function of the phosphate groups; (3) Quantitate the amount of the two brain spectrin subtypes within mouse brain with a quantitative immunodot assay; (4) Determine the precise location of brain spectrin subtypes, within specific neurons and glial cells by immunoelectronmicroscopy utilizing monoclonal antibodies; (5) Determine the domain structure of synapsin I, localizing the spectrin and synaptic vesicle binding domains. Quantitate the affinity and stoichiometry of the brain spectrin-synapsin I interaction and determine whether this interaction is regulated by synapsin I phosphorylation. Study the role of synapsin I in the brain spectrin-actin interaction; (6) Study the rate and stoichiometry of spectrin subunit synthesis and assembly within cultured S20Y mouse neuroblastoma cells and dorsal root ganglion (DRG) primary neuronal cultures; (7) Study the function of brain spectrin by microinjection of monoclonal antibodies which recognize only the brain spectrin (240/235) or brain spectrin (240/235E) subtypes, into neuroblastoma or primary DRG neurons. Observe the effect of spectrin antibody microinjection upon neuronal shape and neurite extension. Determine the effect upon microtubule, neurofilament, and actin microfilament organization.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
2R01NS019357-04
Application #
3399417
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1983-04-01
Project End
1991-07-31
Budget Start
1986-08-01
Budget End
1987-07-31
Support Year
4
Fiscal Year
1986
Total Cost
Indirect Cost

  Institution

Name
Pennsylvania State University
Department
Type
Schools of Medicine
DUNS #
129348186
City
Hershey
State
PA
Country
United States
Zip Code
17033

  Publications

Goodman, S R; Zimmer, W E; Clark, M B et al. (1995) Brain spectrin: of mice and men. Brain Res Bull 36:593-606
Lengeling, A; Zimmer, W E; Goodman, S R et al. (1994) Exclusion of two candidate genes, Spnb-2 and Dcd, for the wobbler spinal muscular atrophy gene on proximal mouse chromosome 11. Mamm Genome 5:163-6
Clark, M B; Ma, Y; Bloom, M L et al. (1994) Brain alpha erythroid spectrin: identification, compartmentalization, and beta spectrin associations. Brain Res 663:223-36
Ma, Y; Zimmer, W E; Riederer, B M et al. (1993) The complete amino acid sequence for brain beta spectrin (beta fodrin): relationship to globin sequences. Brain Res Mol Brain Res 18:87-99
Bloom, M L; Lee, B K; Birkenmeier, C S et al. (1992) Brain beta spectrin isoform 235 (Spnb-2) maps to mouse chromosome 11. Mamm Genome 3:293-5
Zimmer, W E; Ma, Y; Zagon, I S et al. (1992) Developmental expression of brain beta-spectrin isoform messenger RNAs. Brain Res 594:75-83
Zimmer, W E; Zagon, I S; Casoria, L A et al. (1992) Identification of an amelin isoform located in axons. Brain Res 582:94-100
Zimmer, W E; Ma, Y P; Goodman, S R (1991) Identification of a mouse brain beta-spectrin cDNA and distribution of its mRNA in adult tissues. Brain Res Bull 27:187-93
Sikorski, A F; Goodman, S R (1991) The effect of synapsin I phosphorylation upon binding of synaptic vesicles to spectrin. Brain Res Bull 27:195-8
Isayama, T; Goodman, S R; Zagon, I S (1991) Spectrin isoforms in the mammalian retina. J Neurosci 11:3531-8

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