1. Master course Molecular & Cellular Life Sciences Part 1: Intracellular Membrane Processes October 8, 2009 Paul van Bergen en Henegouwen

2. Mechanisms of clathrin-independent internalization Paul van Bergen en Henegouwen

3. Different endocytic portals

4. Characteristics

5. Endocytosis: whats the problem? bendingfission

6. Five ways to bend a membrane

7. Clathrin-dependent internalization

8. Clathrin-coated and vesicles: Ultrathin section of plastic-embedded material

9. Clathrin-coated pits and vesicles by EM Replica of freeze-dried plasma membranes

10. Clathrin-coated vesicles: Replica of freeze-dried plasma membranes

11. Clathrin: Heavy- and light chains

12. Clathrin makes a network

13. How clathrin is bending?

14. AP-2 Adaptor Protein-2 complex a b2b2 m2m2 s2s2 Ear Hinge Trunk 100 kDa 50 kDa 17 kDa a-adaptin b-adaptin

15. There are different AP-complexes on different sites in the cell Golgi-complex Plasma membrane Golgi-complex ab2b2 m2m2 s2s2 g b1b1 m1m1 s1s1 d b3b3 m3m3 s3s3 AP-1 AP-2 AP-3

16. Protein/protein interactions control CCV formation a b2 m2 s2 Receptor Yxxf Eps15 DPF Receptor LL L fxf D/E Clathrin AP-2 I EHs II COIL III DPF/UIM Epsin NPF Lipids DPW PIP 2

17. Epsin induces bending of liposomes

18. The netwerk forms a ball Electron tomography of a coated vesicle Adaptor protein (AP2) complex s chainm chain b adaptina adaptin Clathrin triskelion Smith et al. (1998) EMBO J. 17, 4943

19. Overview of clathrin structures Marsh and McMahon (1999) Science 285, 215-220

20. From bending to fission

21. Dynamin, a GTPase involved in fission PIP 2 Amphiphysin

22. Dynamin is forming a ring around the neck

23. Effect of pure dynamin on lipid vesicles

24. PIPK Membrane curvature generation AP2 Clathrin Eps15 Intersectin Amphiphysin Epsin Dynamin Endophilin Moleculen involved in the whole process

25. Possible role for actin filaments

26. PIPK Membrane curvature generation AP2 Clathrin Eps15 Intersectin Amphiphysin Epsin Dynamin Endophilin Actin Synaptojanin Auxilin Hsc70 Moleculen involved in the whole process

27. Overview of all interactions

28. EGFR internalization via a non-coated pathway!

29. Is the alternative route visible?

30. Caveolae

31. Caveolin

32. Caveolar internalization starting from cholesterol-rich domains

33. Trafficking of Caveolae

34. Phagocytosis: Internalization of large items

35. Overview of different internalization routes

36. Properties of different CI-routes

37. Classification of internalization routes

38. Questions What does the bending? What does the fission?

39. Interleukin-2 receptor complex

40. IL-2R internalization is Clathrin-independent clathrin-inhibition by ΔEps15

41. IL-2R internalization is Dynamin-dependent Dynamin-inhibition by Dyn-K44 TfR wt K44A

42. IL-2R is present in the Detergent Resistant Membrane (DRM) fraction