. Tyrosine phosphorylation of proteins in cell-ECM adhesions is a major factor driving the proliferative growth, motility, and invasion of cells. Integrin-mediated adhesion activates a signaling complex formed by the association of two tyrosine kinases: FAK and Src. A major substrate of the FAK/Src complex is the docking protein p130CAS, which undergoes phosphorylation on numerous tyrosine residues in its """"""""substrate domain"""""""". Recent studies demonstrated the importance of FAK/Src/CAS complex signaling in the related processes of cell motility and invasion. The broad long-term objective of this project is to fully understand the molecular interactions necessary for FAK/Src/CAS complex activation and how downstream signaling events impact the dynamic events that underlie cell motility and invasion.
Three specific aims are proposed.
Aim 1 will use live cell imaging of fluorescent-tagged CAS variants to determine the dynamics and mechanism of CAS adhesion site targeting. The identification of CAS-interacting proteins important for its localization will also be pursued.
Aim 2 will employ time-lapse microscopy to determine the impact of CAS substrate domain phosphorylation on the dynamic processes of lamellipodial protrusion and adhesion assembly/disassembly that drive cell motility, and on the dynamic assembly of podosomes-invadopodia that drive invasion. In related studies, the capacity of CAS substrate domain phosphorylation to promote actin polymerization will be investigated. While the first two Aims make use of CAS-null mouse fibroblast models, Aim 3 extends the work to model invasive human breast cancer cell lines through the use RNA interference to disrupt CAS expression. The impact of the CAS knockdown on cell proliferation and invasion in vitro and on tumor growth and metastasis in vivo will be assessed. Reconstitution experiments with wild-type vs. CAS mutational variants will reveal the important functional domains. Relevance. The proposed studies will fill key gaps in our understanding of the basic signaling machinery that drives the complex dynamic events important for cell motility and invasion. These events have a central role in normal tissue development/regeneration and in many pathological processes including arthritis, atherosclerosis, inflammation, osteoporosis, and cancer progression. The work has potential to reveal new therapeutic targets and prognostic indicators for diseases that arise from aberrant motility and invasion.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM049882-15
Application #
7779412
Study Section
Tumor Progression and Metastasis Study Section (TPM)
Program Officer
Maas, Stefan
Project Start
1994-12-01
Project End
2011-06-28
Budget Start
2010-03-01
Budget End
2011-06-28
Support Year
15
Fiscal Year
2010
Total Cost
$315,902
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Luo, Weifeng; JanoŇ°tiak, Radoslav; Tolde, Ond?ej et al. (2017) ARHGAP42 is activated by Src-mediated tyrosine phosphorylation to promote cell motility. J Cell Sci 130:2382-2393
Meenderink, Leslie M; Ryzhova, Larisa M; Donato, Dominique M et al. (2010) P130Cas Src-binding and substrate domains have distinct roles in sustaining focal adhesion disassembly and promoting cell migration. PLoS One 5:e13412
Dumbauld, David W; Michael, Kristin E; Hanks, Steven K et al. (2010) Focal adhesion kinase-dependent regulation of adhesive forces involves vinculin recruitment to focal adhesions. Biol Cell 102:203-213
Donato, Dominique M; Ryzhova, Larisa M; Meenderink, Leslie M et al. (2010) Dynamics and mechanism of p130Cas localization to focal adhesions. J Biol Chem 285:20769-79
Cunningham-Edmondson, Anna C; Hanks, Steven K (2009) p130Cas substrate domain signaling promotes migration, invasion, and survival of estrogen receptor-negative breast cancer cells. Breast Cancer (Dove Med Press) 2009:39-52
Michael, Kristin E; Dumbauld, David W; Burns, Kellie L et al. (2009) Focal adhesion kinase modulates cell adhesion strengthening via integrin activation. Mol Biol Cell 20:2508-19
Constancio-Lund, Sabata S; Brabek, Jan; Hanks, Steven K (2009) Src transformation of colonic epithelial cells: enhanced anchorage-independent growth in an Apc(+/min) background. Mol Carcinog 48:156-66
Ricono, Jill M; Huang, Miller; Barnes, Leo A et al. (2009) Specific cross-talk between epidermal growth factor receptor and integrin alphavbeta5 promotes carcinoma cell invasion and metastasis. Cancer Res 69:1383-91
Luo, Weifeng; Slebos, Robbert J; Hill, Salisha et al. (2008) Global impact of oncogenic Src on a phosphotyrosine proteome. J Proteome Res 7:3447-60
Siesser, Priscila M F; Meenderink, Leslie M; Ryzhova, Larisa et al. (2008) A FAK/Src chimera with gain-of-function properties promotes formation of large peripheral adhesions associated with dynamic actin assembly. Cell Motil Cytoskeleton 65:25-39

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