Module Four Research Proposal
Project Overview: "Integrin adhesion receptors have emerged as central regulators of cell-biomaterial interactions" - Andrés J. Garcia (reference (1)) We want to mediate cell adhesin by engineering changes in integrins in hopes of better understanding cell response to binding materials. We want to make integrins that bind more or less strongly to materials. Natural ligands for integrins are fibronectin, collagen, and laminin. We could engineer a novel integrin to bind to a different ligand, perhaps a sythetic material or scaffold.
Background Information: Integrins are membrane bound proteins important for cell adhesion to ECM (extra cellular matrix) and also for signal transduction. They are involved in signaling that leads to differentiation, migration, cell growth, and apoptosis. Integrins are also connected to cancer as mutated integrin genes have been found in some types of cancer and integrins may be involved in metastasis. The integrin protein is a heterodimer composed of alpha and beta subunits (in mammals, there are 18 alpha subunits and 8 beta subunits, which can be combined in various ways to create 24 unique integrin receptors). Ligand binding is dependent on divalent cations - Mg2+ and Mn2+ promote binding.
Statement of Research Proposal and Goals: We propose to create novel integrins using a targeted mutagenesis approach. Our goals are to create integrins with different binding properties and then study their interactions with synthetic scaffolds used in tissue engineering. Or we could use the mutated integrins to study differentiation - we could examine how a very weakly or tightly binding integrin affects differentiation of stem cells.
Project Details and Methods: We will create a library of cells expressing different integrins, each with a different single amino acid change. We can find which of these integrins allow cells to bind better or worse to a certain material, and then sequence the integrin to find out how the integrin differs from the naturally occurring integrin. We can perform assays to see if the mutated integrin affects important cell signaling processes.
Predicted Outcomes: We predict that changing structure of integrin will change the way the cell can interact with a material. Some engineered integrins will bind tighter to a certain surface than their naturally occurring counterparts and some engineered integrins will bind less tightly than their naturally occurring counterparts. I am not sure how this will effect cell signaling processes mediated by integrins.
Necessary Ressources: tools to mutate amino acid sequences of proteins, materials of interest to test cells on (a material used for tissue engineering)
Societal Impact: tissue engineering, we could create cells with integrins that allow these cells to bind better to synthetic scaffolds.
1)Get a Grip: integrins in cell-biomaterial interactions Andrés J. García, Biomaterials Volume 26, Issue 36 (This is an overview of how integrins interact with materials and how one can engineer changes in these interactions)
2)Integrin - Wikipedia, the free encyclopedia URL:http://en.wikipedia.org/wiki/Integrin (This is a good overview of what integrins are)
3)Integrin Structure: Heady advances in ligand binding but activation still makes the knees wobble Martin J. Humphries et al, Trends in Biochemical Sciences, Volume 28 No.6 June 2003 (Contains information on the structure of Integrins and how structure may be related to functionality)
4)Protein engineering - Wikipedia, the free encyclopedia URL:http://en.wikipedia.org/wiki/Protein_engineering (Outlines the different strategies for protein engineering)
5)Signal transduction through chondrocyte integrin receptors induces matrix metalloproteinase synthesis and synergizes with interleukin-1