Adhesion of cells to an extracellular matrix (ECM) is involved in a number of biological phenomena including wound healing, embryogenesis, blood coagulation, and metastasis. The long term goal of this project is to elucidate the molecular events that commence with the cell attachment stage of cell-substrate adhesion and regulate the ensuing cell spreading and cell migration stages. We have demonstrated that during attachment of cells to collagen of fibronectin the cognate receptors become clustered and activate cytosolic phospholipase A2 by phosphorylation and translocation to the membrane where arachidonic acid (AA) is release from phospholipids. AA can be oxidized by lipoxygenase into metabolites that turn on the production of diacylglycerol followed by the translocation of protein kinase C to the membrane to activate actin polymerization and the cell spreading stage of adhesion. This information and preliminary work leads to the following hypothesis: Cell spreading and cell migration are regulated respectively by two lipid second messenger pathways which are branches of AA metabolism; lipoxygenase (LOX) and cyclooxygenase (COX) metabolites separately activate sequences of kinases and small GTP-binding proteins leading respectively to the polymerization of actin essential for spreading and bundling of actin filaments essential for migration. Pharmacological, biochemical and molecular biological strategies will be used to evaluate the hypothesis by exploring the following specific aims: 1. Determine which COX metabolites upregulate cAMP to increase PKA activity for actin bundling and migration. 2. Determine whether the amount of COX that is ectopically expressed or expressed in response to a calcium signal determines the level of cAMP and the rate of migration. 3. Establish if and where the GTP-binding proteins, Cdc42, Rac and Rho are involved in the LOX and COX pathways. 4. Ascertain how the metabolites of the three lipoxygenases might regulate the production of diacylglycerol which is essential for PKC activation in the cell spreading stage of adhesion.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM029127-17
Application #
6018533
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1981-04-01
Project End
2002-06-30
Budget Start
1999-07-01
Budget End
2000-06-30
Support Year
17
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Massachusetts Amherst
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
153223151
City
Amherst
State
MA
Country
United States
Zip Code
01003
Tsai, Irene Y; Green, J Angelo; Kimura, Masahiro et al. (2007) Novel transparent nano- to micro-heterogeneous substrates for in-situ cell migration study. J Biomed Mater Res A 80:509-12
Tsai, Irene Y; Kimura, Masahiro; Stockton, Rebecca et al. (2004) Fibroblast adhesion to micro- and nano-heterogeneous topography using diblock copolymers and homopolymers. J Biomed Mater Res A 71:462-9
Green, J Angelo; Stockton, Rebecca A; Johnson, Christopher et al. (2004) 5-lipoxygenase and cyclooxygenase regulate wound closure in NIH/3T3 fibroblast monolayers. Am J Physiol Cell Physiol 287:C373-83
Glenn, Honor L; Jacobson, Bruce S (2003) Cyclooxygenase and cAMP-dependent protein kinase reorganize the actin cytoskeleton for motility in HeLa cells. Cell Motil Cytoskeleton 55:265-77
Roberts, Louis A; Glenn, Honor L; Whitfield, Rebecca A et al. (2002) Regulation of cell-substrate adhesion by the lipoxygenase and cyclooxygenase branches of arachidonic acid metabolism. Adv Exp Med Biol 507:525-9
Stockton, R A; Jacobson, B S (2001) Modulation of cell-substrate adhesion by arachidonic acid: lipoxygenase regulates cell spreading and ERK1/2-inducible cyclooxygenase regulates cell migration in NIH-3T3 fibroblasts. Mol Biol Cell 12:1937-56
Jacobson, B S (2000) Hereditary hemorrhagic telangiectasia: A model for blood vessel growth and enlargement. Am J Pathol 156:737-42
Whitfield, R A; Jacobson, B S (1999) The beta1-integrin cytosolic domain optimizes phospholipase A2-mediated arachidonic acid release required for NIH-3T3 cell spreading. Biochem Biophys Res Commun 258:306-12
Crawford, J R; Jacobson, B S (1998) Extracellular calcium regulates HeLa cell morphology during adhesion to gelatin: role of translocation and phosphorylation of cytosolic phospholipase A2. Mol Biol Cell 9:3429-43
Chun, J; Auer, K A; Jacobson, B S (1997) Arachidonate initiated protein kinase C activation regulates HeLa cell spreading on a gelatin substrate by inducing F-actin formation and exocytotic upregulation of beta 1 integrin. J Cell Physiol 173:361-70

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