Lecture 16 Motor proteins and cytoskeletal-mediated cell behavior

1. Lecture 16 Motor proteins and cytoskeletal-mediated cell behavior

2. Myosin II and myosin II bipolar thick filament 2 heavy chains: Globular head domain (ATP) long tails (coiled coils) 2 light chains: Bipolar thick filaments: Tail-tail interactions Hundreds of myosin II molecules Thin filament: actin filament

3. Experimental evidence for the motor activity of the myosin head 0.6 sec apart 4 um/sec Myosin head S1: by chymotrypsin and papain Attached to the glass slide

4. Myosin superfamily Similar motor domains Move to plus end except VI Small insertion in the motor domain

5. Function of myosins Myosin II: contractile activity in muscle and nonmuscle cells, cytokinesis (pinching apart of a dividing cell into two daughters), forward translocation of cell body during migration); Myosin I: contain a second actin-binding or membrane-binding site in their tails- Intracellular organization and the protrusion of actin-rich structures at the cell surface; Myosin V: vesicle and organelle transport; Myosin VII: inner ear (deafness when mutated)

6. Microtubule motors Kinesins Dyneins: minus-end-directed Unrelated to the kinesin superfamily Cytoplasmic dyneins: vesicle trafficking and Golgi apparatus localization; Axonemal dyneins: sliding movements of MT(14um/sec) Third: cilia beating Kinesin-related proteins, KRPs Most of them have N-terminal motor domains walk toward plus end One family has C-terminal motor domains and move to minus end BimC forms bipolar motor

7. Cycles if structural changes in myosin Head-over-head movement of kinesin 5 nM

8. Attachment of dynein to a membrane-enclosed organelle Organization of Golgi by MT Large complex! Colchicine or nocodazole causes ER to collapse to center and Golgi to fragment and disperse Kinesin: motor protein receptors

9. Myofibrils under EM Skeletal muscle cells (muscle fibers) multinuclear sarcomere Myofibrils: 1-2 um diameter, made of sarcomeres, 2.2 um long Z-disc:plus end

10. Sliding-filament model of muscle contraction 300 heads Shorten by 10% in <1/50th sec 15 um/sec Titin: “molecular spring” Stable actin filament “Molecular ruler” Nebulin: repeating 35 aa actin-binding motif

11. The control of skeletal muscle contraction by troponin Troponin I-T complex pulls the the tropomyosin out of its normal into a position along the actin filament that blocks the the binding of myosin heads Upon Ca++ increase, troponin C causes troponin I to release its hold on actin, allowing tropomyosin to slip back so that myosin head can walk along the actin filament

12. Effect of the heart of a suble mutation in the cardiac myosin FHC: Inherited 2/1000: heart enlargement, abnormal small coronary vessels, disturbances of heart rhythm (cardiac arryhthmias)--mutations in myosins and Contractile proteins. Dilated cardiomyopathy: cardiac actin mutations--early heart failure Familial hypertrophic cardiomyopathy Frequent cause of sudden death in young athletes

13. The arrangement of MTs in a flagella or cilium Contrasting motions of flagella and cilia MT and dynein based motility structures “9+2” arrangement In sperm and protozoa Whip-like motion of cilia Move cells or liquid or other cells Base binds to A MT and heads bind to B MT

14. The bending of an axoneme The motor action causes a bending motion, creating waves of beating motion as in a sperm. Bacterial flagella don’t have MT or dynein and do not wave Or beat. Instead, long, rigid, helical Filaments of repeating subunits of flagellin. Move like propellers driven by rotary motor in the cell wall. The name is a mistake… Basal bodies Kartagener’s Syndrome: Male sterility (immotile sperm) Higher susceptibilty of long infections Left-right body axis defects

15. How basic cytoskeletal mechanisms produce complex cell behaviors Cell shape Migration Division

16. Polarity of actin patches and cables throuhgout the yeast cell cycle New patches Very few cytoplasmic MT Cables random Actin patches are concentrated at the growing tip of the bud Actin cables align and point toward them

17. Morphological polarization of yeast cells in response to mating factor Yeast cells can’t swim a-cell and a-cell, two mating types Shmoo tip grow toward the highest concentration of the signal molecule

18. The signaling pathway in the yeast mating factor response

19. A model of how forces from actin-rich cortex move a cell forward

20. Behavior of lamellipodial fragments Lamellipodia contain all of the machinery that is required for cell motility!

21. A model for protrusion of the actin meshwork at the leading edge

22. Lamellipodia and ruffles at the leading edge Lamellipodia that fail to attach to the substratum are swept backward-- ruffling

23. Neutrophil polarization and chemotaxis Peptide formyl Met-Leu-Phe New lamellipodium toward the tip Extends lamellipodium polarizes cytoskeleton and cell moves forward

24. The complex morphological specialization of nerrons depends on the cytoskeleton Anterograde transport Retrograde transport

25. Neuronal growth cone

26. Cytoskeleton provide the engine for construction of the entire nervous system As well as the supporting structures that strengthen, stabilize and maintain its parts

27. Summary Actin-based motors, microtubule motors, their structure and function Motility machines: myofibrils and flagella and cilia Complex cellular behaviors mediated by cytoskeleton Cell polarization: yeast budding and mating Cell crawling Morphology