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

2. 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. 3 Outline  Introduction (integrins)  Structure  Ligand binding  Conformational regulation  Inside-out activation  Example

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

5. 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. 6 Structure  Components  18 α -subunits  8 β -subunits  Total of 24 integrins  2 groups  Containing or Lacking von Willebrand factor type A domain ( α A)

7. 7 Structure  von Willebrand factor type A domain ( α A)  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. 8 Structure  αVβ3 ( α A-lacking): α - blue, β - red

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

10. 10 Ligand binding  RGD binding (R – propeller, D – β A) Specificity Determining Loop

11. 11 Conformational regulation  Similarly to α A, β A affinity is assumed to be controlled by conformational change

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

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

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

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

16. 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. 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 α IIb peptide close to membrane  Potentially negatively regulates talin-induced activation

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

19. 19 Example

20. 20 Example

21. 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