Our goal is to elucidate the regulatory mechanisms of integrin function. As transmembrane receptors, integrins physically connect a cell to another cell or to the extracellular matrix and biochemically transduce signals bidirectionally across the membrane. Understanding the inner workings and regulations of integrins is important because their dysfunction and dysregulation contribute to a wide variety of diseases. The adhesive and signaling activities of integrins depend on their conformations. Applied force can regulate adhesion, signaling, and conformational changes of integrins. In the past grant cycle, we investigated the mechanically-regulated interaction kinetics of single-bonds of integrins 1L22, 1421 and 1521 with their respective ligands, intercellular adhesion molecule 1, vascular adhesion molecule 1 and fibronectin. In this renewal application, we will first directly visualize and characterize the bending and unbending conformational change dynamics of three integrins, 1L22, 1421, and 1521, at the single-molecule level, under controlled forces on living cells in real-time. Using (un)bending measurements as readouts, we will next examine the conformational coupling of unbending with leg separation and hybrid domain swing-out. Thus, our proposal will fill the gaps of crystallography and electron microscopy as these approaches can only observe static conformations in the absence of force. In addition to innovation in the research topics, our approaches are also innovative, as we will combine single-molecule experiments and molecular dynamics simulations with mutagenesis to dissect the structural determinants of integrin conformational change dynamics. Finally, we will bridge biophysics with cell biology by relating the structural- and force-dependent conformational changes to the integrin-ligand binding kinetics. The three specific aims are to: 1) characterize the integrin bending and unbending conformational changes and their regulations;2) examine coupling among three conformational changes and dissect their structural determinants;3) relate integrin conformational changes and ligand binding to intracellular signaling. The results of this study will advance our understanding of the inner workings of integrins as molecular nano machines and will serve as a foundation for a better understanding of physiological and pathological processes mediated by integrins.

Public Health Relevance

We propose to directly visualize and characterize global conformational changes of single integrins under controlled forces on living cells in real-time and relate these to the integrin-ligand binding kinetics. Integrins are transmembrane receptors that integrate the cytoskeleton with points of attachment in the extracellular environment to mediate a wide variety of cellular processes by providing adhesion and signals. Understanding the inner workings and regulations of integrins is important because their dysfunction and dysregulation contribute to a wide range of diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI044902-11
Application #
8245904
Study Section
Intercellular Interactions (ICI)
Program Officer
Leitner, Wolfgang W
Project Start
2000-08-01
Project End
2017-01-31
Budget Start
2012-02-03
Budget End
2013-01-31
Support Year
11
Fiscal Year
2012
Total Cost
$480,804
Indirect Cost
$134,167
Name
Georgia Institute of Technology
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
097394084
City
Atlanta
State
GA
Country
United States
Zip Code
30332
Chen, Yunfeng; Lee, Hyunjung; Tong, Haibin et al. (2016) Force regulated conformational change of integrin αVβ3. Matrix Biol :
Lee, Cho-Yin; Lou, Jizhong; Wen, Kuo-Kuang et al. (2016) Regulation of actin catch-slip bonds with a RhoA-formin module. Sci Rep 6:35058
Li, Quhuan; Wayman, Annica; Lin, Jiangguo et al. (2016) Flow-Enhanced Stability of Rolling Adhesion through E-Selectin. Biophys J 111:686-99
Li, Zhenhai; Lee, Hyunjung; Zhu, Cheng (2016) Molecular mechanisms of mechanotransduction in integrin-mediated cell-matrix adhesion. Exp Cell Res 349:85-94
Li, Zhenhai; Kong, Fang; Zhu, Cheng (2016) A model for cyclic mechanical reinforcement. Sci Rep 6:35954
Li, Kaitao; Xiang, Xue; Sun, Jie et al. (2016) Imaging Spatiotemporal Activities of ZAP-70 in Live T Cells Using a FRET-Based Biosensor. Ann Biomed Eng 44:3510-3521
Liu, Baoyu; Chen, Wei; Zhu, Cheng (2015) Molecular force spectroscopy on cells. Annu Rev Phys Chem 66:427-51
Chen, Yunfeng; Liu, Baoyu; Ju, Lining et al. (2015) Fluorescence Biomembrane Force Probe: Concurrent Quantitation of Receptor-ligand Kinetics and Binding-induced Intracellular Signaling on a Single Cell. J Vis Exp :e52975
Rosetti, Florencia; Chen, Yunfeng; Sen, Mehmet et al. (2015) A Lupus-Associated Mac-1 Variant Has Defects in Integrin Allostery and Interaction with Ligands under Force. Cell Rep :
Mallis, Robert J; Bai, Ke; Arthanari, Haribabu et al. (2015) Pre-TCR ligand binding impacts thymocyte development before αβTCR expression. Proc Natl Acad Sci U S A 112:8373-8

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