1. Cell Motility Lecture 17

2. Cell Motility Includes: –Changes in Cell Location –Limited Movements of Parts of Cells Occurs at the Subcellular, Cellular, and Tissue Level Is Dependent Upon: –The Cytoskeletal Scaffold –Motor Proteins –ATP

3. Motor Proteins and The Cytoskeleton Microtubule-based Movements: Kinesins and Dyneins (MAPS); Fast Axonal Transport Movement of Cell Appendages Movement of Internal Membranes Movement of Chromosomes during Mitosis Actin-Based Movements: Myosins Muscle Contraction Cell Contraction Whole Cell Movement All Motor Proteins use ATP as an energy source.

4. Microtubule Motor Proteins Two Main Families Kinesins- Primarily Plus End Directed -Away from Centrosome Dyneins- Minus End Directed -Toward Centrosome -Largest Motor Proteins 2 Globular Heads -MT Binding -ATP Binding/Hydrolyzing Middle Flexible Domain -Mediates Dimerization Neck- Tail For Binding Cargo/Adaptors

5. Microtubules Function to Maintain Internal Cell Organization and Cell Shape Cell Polarization is a Reflection of the Polarized System of Microtubules in the Cell Interior MT can be Capped to Stabilize

6. Microtubule Motor Proteins Help Arrange Membrane Enclosed Organelles in Eukaryotes Microtubules Golgi Depolymerize MT- breakdown of Golgi

7. The Tail of the MT Motor Proteins Determines What Cargo Is Transported Kinesin- Processive -Fast Axonal Transport -Arrange ER Cytoplasmic Dynein- Processive -Localize Golgi Axonemal Dynein Motion of Cilia and Flagella Cargo Binding: Often Involves Adaptor Proteins

8. Kinesin Walking to the Plus End of MTs Leading Head (1) Attaches to a  Tubulin (No ATP) Head 1 Binds ATP Head 2 Propelled Forward Past Leading Head ADP Release From Head 2 and Hydrolysis Of ATP by Head 1 Cycle Can Begin Again Moved One Step (1) (2) (1) (2) (1) (2) Processive: At Least One Head is Always Attached

9. Kinesin in vitro Motility Assays

10. Cilia from Rabbit Trachea Axonemal Dynein: The Motor that Moves Cilia and Flagella MT are Stabilized- No Longer Demonstrate Dynamic Instability Cilia of Rabbit Trachea Flagellum Of Sperm + + + + Flagella Drive Movement of Individual Cell Longer than Cilia 1-2 per cell S shaped wave Stationary Cells- Allows Movement over Tissue Shorter than Flagella 100+ per cell Generate force by bending at base

11. Organization of the Axoneme “9 + 2” Pattern Nexin Connects Adjacent Doublets Dynein -Head and Tail Bound to Different MT

12. Model for Dynein-Mediated Sliding of MTs (No Nexin)

13. Actin Based Movements: The Myosin Superfamily 18 Classes of Myosins Move toward Plus(Barbed) End of Actin (except one Class) 4 Broad Groups Based on Function: 1)Power Muscle and Cellular Contraction Striated Muscle Contractile Ring-Will cover later lecture 2) Power Membrane and Vesicle Transport Short Range Endocytic and Phagocytic Vesicles 3)Cell Shape and Polarity 4) Signal Transduction and Sensory Photoreceptors and Hearing Head- binds/hydrolyzes ATP and Actin Regulatory Sites Tail-

14. In vitro Motility Assays with Myosin Purified S1 Heads Attached to Glass + ATP Filaments Glide Along Surface

15. Membrane and Vesicle Transport: The Role of Myosin I in Cells All Cell Types Bind Cortical Actin Filaments- Lead to Change in Cell Shape Transport a Vesicle or Organelle

16. Two Globular Heads Coiled Coil Tail Assemble into Bipolar Myosin Filament Muscle Contraction: The Role of Myosin II

17. Tight binding No ATP Myosin II Movement to the Plus End of Actin Filaments ATP Binding Releases Head ATP Hydrolysis Occurs Head is Cocked Head Rebinds Filament Pi Release Trigger Power Stroke ADP released Bind ATP Again and Release Cycle Starts Over Not Processive Like Kinesin

18. Organization of Muscle Fiber

19. Organization of the Skeletal Muscle Fiber Skeletal Muscle Cell (Muscle Fiber) Myofilaments make up Myofibrils Thick- Myosin Thin- Actin

20. Hexagonal Arrangement of Actin Filaments Around the Myosin

21. Skeletal Muscle Banding Pattern Light Band= I Band Thin Actin Fibers Dark Band=A Band Thin Actin and Thick Myosin Fibers Overlap

22. Structural Proteins of the Sarcomere Actin Filament Stability Requires: Cap Z- Cap Plus End of Actin Filament Tropomodulin- Cap Minus Ends of Actin Filament Nebulin –Stabilize Filament/Role in Length  Actinin- Component of Z Disc - Actin Bundling

23. Sarcomeres Shorten During Skeletal Muscle Contraction No Shortening of Thin or Thick Filaments Sliding over one another

24. Myosin Walks Along Actin Filament Attached Released Cocked Force Generating Attached Nucleotide Free Form ATP Binding ATP Hydrolysis Conformational Change Pi release-Initiates Power Stroke Strong Binding- Power Stroke (Conformational change to Original Shape ) and ADP Release

25. The Role of Ca2+ and Regulatory Proteins in Skeletal Muscle Contraction

26. The Cytoskeleton in Moving Fibroblast Thin, Sheetlike Thin,Stiff Protrusions Meshwork

27. Crawling Cell

28. Actin-Rich Cortex Moves a Cell Forward Myosin II functions in Cell Contraction Extension of lamellipodium Lamellipodium anchors to substrate Retraction of the opposite end of the cell

29. Formation of Leading Edge of a Lamellipodia Nucleation of Actin at the Plasma Membrane Size of Lamellipodia Remains Constant As Assembly at Leading Edge is Balanced By Disassembly At Rear

30. Lamellipodia Filopodia Extracellular Signals Activate Small G proteins That Regulate ARP Complex