Jack Tuszynski Department of Physics University of Alberta Edmonton, Canada

1. The Dynamics of C-termini of Microtubules in Dendrites:A possible clue for the role of neuronal cytoskeleton in the functioning of the brain Jack Tuszynski Department of Physics University of Alberta Edmonton, Canada

2. Acknowledgements • Avner Priel, Bar Ilan U/Univ of Alberta • Nancy Woolf, UCLA • Eric Carpenter • Tyler Luchko • Horacio Cantiello, Harvard Med School • Stephanie Portet, U Toronto Funding: NSERC, MITACS, YeTaDel, Technology Innovations LLC

3. Motivation • Pyramidal cells collect O(105) inputs • Most of the computational tasks occur in the dendritic arbor • Dendrites are capable of non-trivial computation

4. Microtubules in neurons

5. Cytoskeletal structures in dendrites

6. Challenge: integration of various levels Building a bridge between • the molecular level (cytoskeleton) • the membranelevel (synaptic activity, AP)

7. Dendrites undergo significant structural plasticity during learning phases • Spine level changes • Cytoskeletal changes

8. A MT tree in a dendritic forest

9. Processivity kinesin ncd

10. MT Binding Site of Kinesin Role of C-termini

11. The Model MTN – Microtubule Network • A model of a single MT • Tubulin dimers interact electrically via dipole – dipole interactions • A model of the MAP • Elastic rod connected at both ends to adjacent MTs • MAP – MT interaction • Mechanical (elastic forces)

12. MTN Dynamics

13. Microtubules (MT) • lengths vary, but commonly reach 5–10 µm • usually 13 protofilaments in vivo • 12–17 protofilaments when self-assembled in vitro • protofilaments are strongly bound internally • protein filaments of the cytoskeleton

14. Microtubules in dendrites : • Self-assemble to extend dendrites (and axons) • Form synaptic connections • Linked to ion-channels and synaptic receptors • Organized in parallel structures – interconnected by MAP-2 • Appear in mixed polarity

15. Individual MT life story: dynamic instability • Catastrophes • Rescues • Growth phase • Shrinking phase Simulation using a recursive map

16. Molecular Dynamics (MD) • class of model system • point masses (atoms) • simple forces • bond stretching, • Coulomb, • van der Waals, etc. • Newtonian integration over time

17. Tubulin… • two major types of tubulin (a, b) pair in dimers • tubulin dimers are basic unit • dimers form protofilaments(vertical columns) • dimers form cylindrical microtubules (MTs)

18. Electric Potential • slice through dimer • red/blue by potential strength • green isopotential lines • not well replaced with multipole expansion

19. Protein Surface Potential • surface mix of positive (blue) and negative (red) charge regions • isopotential surface in yellow • note folded C-terminii

20. C-termini • highly variable sequences • strongly electro-negative • having up to 10 netnegative charges • highly mobile, andunstructured region • electrostatic interactionwith nearby charges

21. RMSF & RMSD (work in progress) • RMS fluctuations are a measure of the flexibility of a structure. • RMS deviations measure the change between two static structures. Created with VMD [Humphrey et al., 1996]

22. Reconstruction of tubulin isotype structures including C-termini Green=alphaTBA 2 human Yellow=alpha 1TUB Red=alpha TBA4 human

23. Effects of pH C-termini are antenna-like

24. MT2 MAP MT1 The C-termini are key degrees of freedom

25. Charge distribution of tubulin dimer including C-termini. Blue=negative Red=positive White=neutral Approx. 25 e per monomer or 50 per dimer

26. Blue=negative chargeRed=positive chargeGreen=MAP binding region

27. Conformational states of C-termini Up-up Up-down Down-down

28. C-terminal–BodyInteractions • section of MT with 9 dimers • C-termini (not shown) can fold down onto (red) groove on dimer • very fast in MD simulations

29. Local potential for C-termini orientations Well depth= several kT

30. The proposed model to be developed: • The C-termini as the key degrees of freedom • Each C- terminus can be in one of several states: • up – extending outward from the surface • down – bound to the surface of a MT • Transitions between the states are induced by thermal fluctuations, interaction with neighboring C-termini and interference from adjacent MT’s mediated by the MAP • Transitions may induce ionic waves between MT’s and down the dendrite

31. C-terminal Interactions • neighbouring C-termini • adjacent protein including • kinesin • MAPs (microtubule associated proteins) • adjacent surface of dimers

32. MD simulation of C-termini conformations

35. Collective states • Condition for an ordered state: zJ<kT => at T=300 K we get kT=25 meV Hence even J=5 meV with z=6 neighbors (hexatic lattice) leads to an ordered state where pair-wise interactions are below thermal noise This corresponds to f=6 10 12 Hz (MW) Possibly dipole-dipole or H-bond interactions

36. Ionic Wave Propagation • RLC representation of a biopolymer from the view point of counterions • Speeds of 1-100 m/s possible • Localized traveling waves • Singaling • C-termini may collapse