The goal of this research is to develop assays that can be used to study the interactions of integrins, a class of cell receptor proteins implicated in some forms of tumor metastasis, with extracellular matrix proteins, and also for the study of integrin clustering in the cell membrane. The assay will first be developed using integrin expressing S2 cells, and then tissue expressing mutant integrins will be studied in situ. The mutant integrins to be studied were obtained from genetic screens performed on Drosophila, and will be used to elucidate the molecular mechanism of integrin action. Drosophila is used as a model organism for studying vertebrate integrins due to the considerable structural homology between vertebrate and invertebrate integrins. In situ fluorescent labeling techniques will be utilized to tag the integrin and ligand so that the cells do not need to be fixed to a substrate, and to enable both time-dependent and distance-dependent binding data to be obtained. The general assay format will use fluorescence resonance energy transfer to quantitatively study the interactions between donor and acceptor tagged integrins and/or ligands, and total internal reflection fluorescence will be used as the detection scheme to limit autofluorescence from the bulk cellular material.
| Smith, Emily A; Bunch, Thomas A; Brower, Danny L (2007) General in vivo assay for the study of integrin cell membrane receptor microclustering. Anal Chem 79:3142-7 |
