Spectrin function will be analyzed in a model system, Drosophila melanogaster. Spectrin is a membrane-associated cytoskeletal protein that plays a key role in the determination of erythrocyte cell shape. spectrin also plays unknown, but essential, roles in non-erythroid cells. As a result of our recent work that has identified alpha-spectrin mutants, we are in the singular position of being able to directly assess the functions of spectrin in non-erythroid cells. Using a combination of biochemical, molecular genetic, and classical genetic analyses, we will test various hypotheses for spectrin function in non-erythroid tissue at the cellular, developmental and organismal level. Beta-spectrin mutants will be produced and these, together with recently identified alpha-spectrin mutants, will be utilized to investigate spectrin function in vivo. Phenotype analysis will determine the effects of the complete absence of functional spectrin; mosaic analysis will determine the effects of spectrin absence at selected developmental stages. This phenotype information will be used to design experiments that rescue null mutants with spectrin-DNA constructs that will test the cellular or developmental role of a specific aspect of spectrin organization, such as membrane binding, or spectrin chain- length. Although spectrin isoforms are present in nearly all human cells, the neurological and general health consequences of defective spectrin expression in non-erythroid cells are only beginning to be explored. The proposed experiments will provide an in-depth understanding of spectrin's role in non-erythroid cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM039686-07
Application #
2179961
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1988-06-01
Project End
1997-05-31
Budget Start
1994-06-01
Budget End
1995-05-31
Support Year
7
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Harvard University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
071723621
City
Cambridge
State
MA
Country
United States
Zip Code
02138
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Lue, R A; Brandin, E; Chan, E P et al. (1996) Two independent domains of hDlg are sufficient for subcellular targeting: the PDZ1-2 conformational unit and an alternatively spliced domain. J Cell Biol 135:1125-37
Deng, H; Lee, J K; Goldstein, L S et al. (1995) Drosophila development requires spectrin network formation. J Cell Biol 128:71-9
Zhang, P; Talluri, S; Deng, H et al. (1995) Solution structure of the pleckstrin homology domain of Drosophila beta-spectrin. Structure 3:1185-95
Lee, J K; Coyne, R S; Dubreuil, R R et al. (1993) Cell shape and interaction defects in alpha-spectrin mutants of Drosophila melanogaster. J Cell Biol 123:1797-809
Byers, T J; Brandin, E; Lue, R A et al. (1992) The complete sequence of Drosophila beta-spectrin reveals supra-motifs comprising eight 106-residue segments. Proc Natl Acad Sci U S A 89:6187-91
Dubreuil, R R (1991) Structure and evolution of the actin crosslinking proteins. Bioessays 13:219-26
Dubreuil, R R; Brandin, E; Reisberg, J H et al. (1991) Structure, calmodulin-binding, and calcium-binding properties of recombinant alpha spectrin polypeptides. J Biol Chem 266:7189-93
Winograd, E; Hume, D; Branton, D (1991) Phasing the conformational unit of spectrin. Proc Natl Acad Sci U S A 88:10788-91
Dubreuil, R R; Byers, T J; Stewart, C T et al. (1990) A beta-spectrin isoform from Drosophila (beta H) is similar in size to vertebrate dystrophin. J Cell Biol 111:1849-58

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