title: The effect of ligand affinity on integrins' lateral diffusion in cultured cells.
abstract: The role of ligand affinity in altering αPS2CβPS integrins' lateral mobility was studied using single particle tracking (SPT) with ligand-functionalized quantum dots (QDs) and fluorescence recovery after photobleaching (FRAP) with fluorescent protein tagged integrins. Integrins are ubiquitous transmembrane proteins that are vital for numerous cellular functions, including bidirectional signaling and cell anchorage. Wild-type and high ligand affinity mutant (αPS2CβPS-V409D) integrins were studied in S2 cells. As measured by SPT, the integrin mobile fraction decreased by 22% and had a 4× slower diffusion coefficient for αPS2CβPS-V409D compared to wild-type integrins. These differences are partially the result of αPS2CβPS-V409D integrins' increased clustering. For the wild-type integrins, the average of all diffusion coefficients measured by SPT was statistically similar to the ensemble FRAP results. A 75% slower average diffusion coefficient was measured by SPT compared to FRAP for αPS2CβPS-V409D integrins, and this may be the result of SPT measuring only ligand-bound integrins, in contrast all ligand-bound and ligand-unbound integrins are averaged in FRAP measurements. Specific binding of the ligand-functionalized QDs was 99% for integrin expressing cells. The results prove that the ligand binding affinity affects the lateral dynamics of a subset of integrins based on the complementary SPT and FRAP data.
authors.affiliation: Department of Chemistry, Iowa State University, 1605 Gilman Hall, Ames, IA 50011, USA.
authors.forename: Dipak
authors.initials: D
authors.lastname: Mainali
authors.forename: Emily A
authors.initials: EA
authors.lastname: Smith
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004058:Diffusion / Q000187:drug effects; D036681:Fluorescence Recovery After Photobleaching; D016023:Integrins / Q000378:metabolism*; D008024:Ligands; D009068:Movement / Q000187:drug effects; D011485:Protein Binding; D045663:Quantum Dots; D013379:Substrate Specificity
chemical_list: D016023:Integrins; D008024:Ligands