1. Results from a field-theoretic approach to membrane fusion Michael Schick Department of Physics University of Washington

2. à mon cher maître

4. Maurice Ravel

6. Synaptic Vesicles They are small R~15-25 nm cf bilayer thickness t~4nm They dock: nothing happens They are triggered: fuse rapidly Fusion on demand

7. Synaptic vesicles in C. Elegans J.L. Bessereau: www.biologie.ens.fr

8. Questions Why does fusion occur and how? What are free energy barriers to fusion? What factors promote fusion?

9. Theoretical Procedure Consider fusion of vesicles of block copolymer: universality

10. Theoretical Procedure Consider fusion of vesicles of block copolymer: universality Polymers->Random walk->Diffusion Eq.-> Quantum Mech.

11. Theoretical Procedure Consider fusion of vesicles of block copolymer: universality Polymers->Random walk->Diffusion Eq.-> Quantum Mech. SCFT of interacting polymers-> Hartree Theory Advantages: microscopic model->architecture calculation of free energies

12. Bringing bilayers together removes water-> free energy per area increases-> bilayers are under tension WHY DOES FUSION OCCUR?

13. Bringing bilayers together removes water-> free energy per area increases-> bilayers are under tension Fusion eliminates area -> reduces the free energy Fusion is one possible response to induced tension Lysis (rupture) is another WHY DOES FUSION OCCUR?

14. HOW DOES FUSION OCCUR? one commonly accepted mechanism

15. Kozlov and Markin 1983 Profiles obtained by the theory

16. WHAT ARE THE FREE ENERGY BARRIERS TO FUSION?

17. Consider the fusion of two bilayers One component only A lamellar former

18. Kozlov and Markin 1983 Profiles obtained by the theory

19. One component, f = 0.4 (DOPC),  /  0 = 0.2, equilibrium H, stalk 1 bilayer =4.3 R g

20. One component, f = 0.4,  /  0 = 0.2, equilibrium H, stalk

23. One component, f = 0.4,  /  0 = 0.2, equilibrium H, pore

24. One component, f = 0.4,  /  0 = 0.2, equilibrium H, stalk & pore

25. In lipids, barrier would be 50 kT !

26. WHAT FACTORS AFFECT FUSION?

27. Effect of Tension

28. Effect of Two Components and Asymmetry in Leaves SMALL HEADS, LARGE TAILS FAVORED HERE IN PROXIMAL LEAF

29. Effect of Two Components and Asymmetry in Leaves Average concentration of hex-former is 0.35 0.35 in cis 0.40 in cis 0.43 in cis F/k b T f=0.4 (DOPC) and f=0.294 (DOPE)

30. Effect of Two Components and Asymmetry in Leaves Average concentration of hex-former is 0.35 0.35 in cis 0.40 in cis 0.43 in cis F/k b T Note that stalk becomes metastable. Its formation is now largest barrier

31. Effect of curvature Fusion of Bilayer and Vesicle: 1 bilayer =4.3 R g 60:40 mixture J Y Lee & M.S. BJ 2008

32. What should we expect the effect of curvature to be?

33. As vesicle radius decreases, effective tension increases, which decreases barrier

34. Fusion of two identical vesicles 60:40 mixture

35. H = 2.2 R_g, zero tension, 60:40 mixture Control Fusion by Controlling the Interbilayer Spacing

36. H = 2.2, 2.7 R_g, zero tension

37. H = 2.2, 2.7, 3.2 R_g, zero tension

38. H = 2.2, 2.7, 3.2, 3.7 R_g, zero tension

39. H = 2.2, 2.7, 3.2, 3.7, 4.0 R_g, zero tension

40. Conclusions Two barriers to fusion

41. Conclusions Two barriers to fusion Barrier to stalk formation linear in distance ->easy to prevent fusion

42. Conclusions Two barriers to fusion Barrier to stalk formation linear in distance ->easy to prevent fusion Second barrier reduced by composition and curvature

43. Conclusions Two barriers to fusion Barrier to stalk formation linear in distance ->easy to prevent fusion Second barrier reduced by composition and curvature Consequently, when brought close, stalk barrier is small, ~13kT, and fusion is quick Fusion on demand!

44. Acknowledgements Marcus Mueller Kirill Katsov Jae-Youn Lee NSF Grant DMR 0503752