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The effect of ligand affinity on integrins’ lateral diffusion in cultured cells

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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.

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Abbreviations

SPT:

Single particle tracking

QDs:

Quantum dots

FRAP:

Fluorescence recovery after photobleaching

MSD:

Mean square displacement

LFA-1:

Lymphocyte function-associated antigen 1

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Acknowledgments

This work is supported by the National Science Foundation under Grant CHE-0845236. The authors thank Dr. Javier Vela and Ms. Yijun Guo for providing QDs for initial experiments and Mr. Aleem Syed for helpful discussions.

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  1. Department of Chemistry, Iowa State University, 1605 Gilman Hall, Ames, IA, 50011, USA

    Dipak Mainali & Emily A. Smith

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  1. Dipak Mainali
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  2. Emily A. Smith
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Correspondence to Emily A. Smith.

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Mainali, D., Smith, E.A. The effect of ligand affinity on integrins’ lateral diffusion in cultured cells. Eur Biophys J 42, 281–290 (2013). https://doi.org/10.1007/s00249-012-0873-x

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