Cell-Environment Interaction (inside-out)

1. Wagner Shin Nishitani 04/13/2009 Cell-Environment Interaction (inside-out)

2. Journal Papers Arnaout, M. A., Goodman, S. L. & Xiong, J.-P. Structure and mechanics of integrin-based cell adhesion. Curr. Opin. Cell Biol.19, 495–507 (2007) Meshel, A. S., Wei, Q., Adelstein, R. S. & Sheetz, M. P. Basic mechanism of three-dimensional collagen fibre transport by fibroblasts. Nat. Cell Biol.7, 157–164 (2005)

3. Outline • Introduction (integrins) • Structure • Ligand binding • Conformational regulation • Inside-out activation • Example

4. Introduction Adhesion by integrins Binds to extracellular matrix (ECM) through large ectodomain Binds to cytoskeleton through short cytoplasmic tail

5. Introduction • Adhesion by integrins • Major role in organization of tissue/organs • Cytoskeleton controls affinity of ectodomain to ECM (inside-out) • ECM binding changes cytoskeleton (outside-in) • Transmission of force across membrane

6. Structure Components 18 α-subunits 8 β-subunits Total of 24 integrins 2 groups Containing or Lacking von Willebrand factor type A domain (αA)

7. Structure • von Willebrand factor type A domain (αA) • GTPase-like domain with a metal-ion-dependent adhesion site (MIDAS) instead of the catalytic site • Closed (low affinity) or Open (high affinity) conformations

8. Structure • αVβ3 (αA-lacking): α - blue, β - red

9. Structure • Parts Head (formation of dimer) Knees Legs

10. Ligand binding • RGD binding (R – propeller, D – βA) Specificity Determining Loop

11. Conformational regulation • Similarly to αA, βA affinity is assumed to be controlled by conformational change

12. Conformational regulation • Switchblade model • High affinity of βA dependent on large Hybrid swing-out (~80°) • To provide space, knees fully extended • Allows access to binding site

13. Conformational regulation • Deadbolt model • High affinity of βA possible with small changes between βTD and βA/Hybrid • Ligand binding provides energy for Hybrid swing-out (maybe knees not fully extended)

14. Conformational regulation • Ligand-relay model • Active βA changes conformation of αA for high affinity • αA is the ligand for βA • Consistent with deadbolt model αA-containing αA-lacking

15. Inside-out activation • Talin • Actin binding protein • Localized early with high-affinity integrins • Interaction β • Tail • Close to membrane

16. Inside-out activation • Talin • Interaction close to membrane • Disrupts ionic bonds between α and β tails • Separation between tails for inside-out activation • Unclear!

17. Inside-out activation • Filamin • Binds competitively with talin to β tails • Negatively regulates talin-induced activation • Calcium and integrin binding protein 1 (CIB1) • Binds to αIIb peptide close to membrane • Potentially negatively regulates talin-induced activation

18. Example • Collagen fiber transport by fibroblast • Hand-over-hand cycle • Extension of lamellipodia • Collagen bound by integrin • Retraction of lamellipodia • Collagen released

19. Example

20. Example

21. Example • Collagen fiber transport by fibroblast • Integrin α2β1 involved • Blocking antibody inhibited spreading on collagen-coated substrate • Bind/release cycle • Inside-out control required