): An early event in the development of colon cancer is loss of heterozygosity for the normal adenomatous polyposis coli (APC) gene. APC is normally expressed in the proliferative zone of the crypt border in the colon. It is felt to have important functions in cell migration, anchorage and cell-cell interactions. One of APC's known binding partners is EB1, product of a gene found by the yeast two hybrid interaction trap using APC as bait. EB1 has no known function, but it is expressed in many tissues and conserved from humans to the highly homologous yeast protein Yer0l6p. In unrelated work, EB1 was independently isolated in the lab of Barbara Bierer by immunoprecipitation with a specific form of tubulin, tyrosinated tubulin, thought to represent the dynamic pool of tubulin. The interaction between EB1 and tyrosinated tubulin was established in T-cells, suggesting a role for EB1 in lymphocytes. This proposal outlines the approach to determining the function of EB1. To test the hypothesis that EB1 coordinates the binding of APC with the cytoskeleton, members of the macromolecular complex involving APC, EB1, and tubulin, will be analyzed for direct interactions. Mutational analysis will be used to dissect the domains of EB1 which interact with each specific partner. Changes in EB1 expression and cellular localization during cell division and differentiation will be investigated. The yeast homolog of EB1, Yer016p, has been deleted in S. cerevisiae by homologous recombination. Haploid yeast lacking Yer016p are viable. Their phenotype is currently under study. The yeast model system will allow rapid analysis of the phenotype of Yer016p deletion, assessment of the capacity of the human EB1 gene to replace the function of the yeast homolog, and mutational analysis of human EB1. To assess the role of EB1 within the intact colon and in mammalian development, a murine model of targeted germline deletion of EB1 will be generated. These studies bear directly on the understanding of mitosis, cell cycle progression, growth control, and tumorigenesis. Determining the function of EB1 will elucidate how an early lesion in colon cancer positions the cell to undergo malignant transformation.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Clinical Investigator Award (CIA) (K08)
Project #
7K08DK002578-03
Application #
6132583
Study Section
Subcommittee G - Education (NCI)
Program Officer
Podskalny, Judith M,
Project Start
1998-02-26
Project End
2002-11-30
Budget Start
1999-02-01
Budget End
1999-11-30
Support Year
3
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Harvard University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
082359691
City
Boston
State
MA
Country
United States
Zip Code
02115
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Carvalho, Pedro; Tirnauer, Jennifer S; Pellman, David (2003) Surfing on microtubule ends. Trends Cell Biol 13:229-37
Tirnauer, Jennifer S; Grego, Sonia; Salmon, E D et al. (2002) EB1-microtubule interactions in Xenopus egg extracts: role of EB1 in microtubule stabilization and mechanisms of targeting to microtubules. Mol Biol Cell 13:3614-26
Tirnauer, Jennifer S; Canman, Julie C; Salmon, E D et al. (2002) EB1 targets to kinetochores with attached, polymerizing microtubules. Mol Biol Cell 13:4308-16
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Tirnauer, J S; O'Toole, E; Berrueta, L et al. (1999) Yeast Bim1p promotes the G1-specific dynamics of microtubules. J Cell Biol 145:993-1007