This revised application is designed to investigate the role of c-abl in integrin-mediated signaling. It is based on the published observations made by this investigator that adhesion of 10T1/2 fibroblasts to a fibronectin matrix results in the transient shuttling of c-abl from the nucleus to the cytoplasm where it co-localizes with integrins in focal adhesions. This is concomitant with transient activation of c-abl kinase prior to c-abl returning to the nucleus. Over-expression of a kinase-dead c-abl blocked integrin-dependent MAP kinase activation. The hypothesis to be tested is that integrin molecules regulate c-abl activity and that c-abl may participate in the transduction of signals from focal adhesions to the nucleus.
In Aim 1 in vitro binding experiments utilizing c-abl fusion proteins are proposed to identify c-abl binding proteins. It is anticipated that some proteins, such as paxillin, will be identifiable by their size and available antibodies. Novel proteins will be purified by affinity chromatography in sufficient quantities to permit microsequencing and subsequent cloning.
Aim 2 will utilize a set of existing c-abl mutants in transfection experiments into c-abl null cells to delineate regions of the molecule that are important in the integrin-mediated translocation, association with proteins identified in Aim 1 and activation.
Aim 3 will use the same mutants to map the regions necessary for MAP kinase activation.
Aim 4 is a new Aim designed to address the possible physiologic role of c-abl in integrin signaling. C-abl mutants will be transfected into c-abl null cells and their potential effects on cell growth, apoptosis and transformation measured using standard procedures.
| Lewis, Jean McArthur; Truong, Tony Nguyen; Schwartz, Martin Alexander (2002) Integrins regulate the apoptotic response to DNA damage through modulation of p53. Proc Natl Acad Sci U S A 99:3627-32 |
| Renshaw, M W; Lewis, J M; Schwartz, M A (2000) The c-Abl tyrosine kinase contributes to the transient activation of MAP kinase in cells plated on fibronectin. Oncogene 19:3216-9 |
| Lewis, J M; Schwartz, M A (1998) Integrins regulate the association and phosphorylation of paxillin by c-Abl. J Biol Chem 273:14225-30 |
