The structural protein tubulin is involved in a variety of cellular processes important in morphogenesis, including cell division and flagellar assembly. In many cases, tubulin involved in different processes is found in structurally different microtubule arrays. The regulation of tubulin assembly into microtubule arrays with different functions may utilize discrete structural domains of the tubulin molecule and may be mediated by interactions between tubulin and different sets of microtubule associated proteins. The goal of our research is to determine how tubulin structure regulates tubulin function in vivo and to identify gene products which interact with tubulin during its several functions and which may regulate its assembly. As a model system, we propose a genetic, biochemical, and ultrastructural analysis of the function of tubulin and the molecules with which it interacts during spermatogenesis in Drosophila. We propose to identify genes encoding proteins which interact with the testis-specific Beta tubulin (Beta2) by isolation and characterization of mutations which fail to complement mutant alleles of Beta2 tubulin but which map to other sites on the genome. We have already identified two such """"""""second-site"""""""" recessive male sterile mutations. We plan to identify the mutant gene products by 2D gel analysis or recombinant DNA techniques. We will investigate the function of the mutant gene products in spermatogenesis and the nature of their interaction with Beta2 tubulin by light and electron microscopic analysis of defects in spermatogenesis caused by """"""""second-site"""""""" mutations both as homozygotes and in heterogyzous combination with Beta2 tubulin mutants. We also propose to analyze changes in tubulin structure caused by mutations in the Beta2 tubulin gene which affect some, but not all, of the functions of Beta2 tubulin during spermatogenesis. Changes in mutant tubulin structure will be assayed using monoclonal antibodies specific for different determinants on the tubulin molecule as probes of molecular structure. Structural changes will be correlated with changes in function of mutant tubulins in vivo and in vitro to determine if discrete structural regions of the tubulin molecule function uniquely in different kinds of microtubule dependent processes.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD018127-03
Application #
3315106
Study Section
Genetics Study Section (GEN)
Project Start
1983-12-01
Project End
1986-11-30
Budget Start
1985-12-01
Budget End
1986-11-30
Support Year
3
Fiscal Year
1986
Total Cost
Indirect Cost
Name
University of Colorado at Boulder
Department
Type
Schools of Arts and Sciences
DUNS #
City
Boulder
State
CO
Country
United States
Zip Code
80309
Johnstone, Oona; Deuring, Renate; Bock, Ronald et al. (2005) Belle is a Drosophila DEAD-box protein required for viability and in the germ line. Dev Biol 277:92-101
Brill, J A; Hime, G R; Scharer-Schuksz, M et al. (2000) A phospholipid kinase regulates actin organization and intercellular bridge formation during germline cytokinesis. Development 127:3855-64
Mounkes, L C; Fuller, M T (1999) Molecular characterization of mutant alleles of the DNA repair/basal transcription factor haywire/ERCC3 in Drosophila. Genetics 152:291-7
Mounkes, L C; Fuller, M T (1998) The DUG gene of Drosophila melanogaster encodes a structural and functional homolog of the S. cerevisiae SUG1 predicted ATPase associated with the 26S proteasome. Gene 206:165-74
Hime, G R; Brill, J A; Fuller, M T (1996) Assembly of ring canals in the male germ line from structural components of the contractile ring. J Cell Sci 109 ( Pt 12):2779-88
Hays, T S; Porter, M E; McGrail, M et al. (1994) A cytoplasmic dynein motor in Drosophila: identification and localization during embryogenesis. J Cell Sci 107 ( Pt 6):1557-69
Mounkes, L C; Jones, R S; Liang, B C et al. (1992) A Drosophila model for xeroderma pigmentosum and Cockayne's syndrome: haywire encodes the fly homolog of ERCC3, a human excision repair gene. Cell 71:925-37
Regan, C L; Fuller, M T (1990) Interacting genes that affect microtubule function in Drosophila melanogaster: two classes of mutation revert the failure to complement between haync2 and mutations in tubulin genes. Genetics 125:77-90
Green, L L; Wolf, N; McDonald, K L et al. (1990) Two types of genetic interaction implicate the whirligig gene of Drosophila melanogaster in microtubule organization in the flagellar axoneme. Genetics 126:961-73
Fuller, M T; Regan, C L; Green, L L et al. (1989) Interacting genes identify interacting proteins involved in microtubule function in Drosophila. Cell Motil Cytoskeleton 14:128-35

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