Answer Key: Cell Motility

BIOL230W Week 12 – Cell Motility During Week 12, we will be looking at how motile animal cells move using the ECM. Not many cells in animals are motile. The three common examples ameboid of motility (not flagellar)include: cells migration during wound repair, motility of white blood cells and growth of neuronal cells during synapse formation. We will explore these forms of motility. Day 1 Learning Objectives 1. Describe the role of cell motility, ECM remodeling, cell-cell interactions in wound repair. 2. Analyze the role of the epithelial to mesenchymal transition (EMT) in wound repair. 3. Describe the roles of fibroblasts and chemo-attractants in wound repair. 4. Compare and contrast the functions and components of the two categories of cell junctions: occluding and anchoring. Mini-Lecture: Cell mobility, cell-to-cell interactions and adhesion in wound repair 1. What cells are motile in the image depicting the first stage of wound repair? Fibroblasts and white blood cells 2. What attracts the fibroblasts? Chemo-attractants 3. What is the function of the fibroblasts? Reconstruct the extracellular matrix in the wound. 4. Why does the action of the fibroblast come before the movement of the epithelial cells into the wound? Provides structure for new epithelial cell movement. Small Group Activity: Epithelial to Mesenchymal Transition (EMT) 1 BIOL230W Week 12 – Cell Motility 5. What do the purple, cyan and blue lines between the plasma membranes of neighboring cells near the #1 and #2 on image represent? All of these lines represent cell junctions, or interactions between cells. 6. What happens to the structures mentioned above as the cell transitions to a mesenchymal cell? As this transition takes place, structures that hold the 2 cells go away to allow the cell to move away from the tissue. 7. Describe what happens to the basement membrane as the EMT progresses. As EMT progresses, the basement membrane needs to degrade (temporarily) so the cells can move. 8. Use Google or Wikipedia to look up “matrix metalloproteinases” (MMPs). What structures do you think represent these enzymes and what is their function in the EMT? MMPs degrade collagen in the basement membrane (collagen 4). They are proteinases that break covalent bonds between amino acids to break up basement membrane. 9. As the epithelial specific gene expression becomes repressed and the mesenchymal specific gene expression is activated, export of which ECM component would you expect to increase? Fibronectin 10. What is the result of the signaling pathway transduced by focal adhesions between integrin and fibronectin, remember from last week? It will promote motility. 2 BIOL230W Week 12 – Cell Motility Minilecture: Cell-cell-interactions 11. Name the two types of cell-to-cell junctions we discussed. Tight junctions and anchoring junctions 12. What effect does each cell junction type have on motility? Desmosomes (E-cadherin)- Promote strength and polarity within epithelial cells. Hemidesmosomes (N-cadherin) – Interact with ECM which allows increased motility. Tight junctions (Occludin) – Force regulation of what enters and exits the cell. 13. How might a loss of E-cadherin function and a gain of N-cadherin function affect a cell and why? It would make the cell more motile because it is interacting with the ECM instead of other cells. 14. Why is disruption of occludin function required for motility? Occludin does not interact with the cytoskeleton of other cells, but if it is degraded things can move around the cells instead of through them. Day 2: Learning Objectives 1. Describe how chemicals can attract or repel cells. 2. Compare and contrast actin and mictotubule structure and function in cell motility. 3. Outline the molecular events occurring at the five stages of amoeboid crawling, including the role of cytoskeletal proteins: polarization, cell protrusions, cell adhesion, cell contraction, cell detachment. 4. Identify and/or describe filopodia vs. lamellipodia. Minilecture: Motility 15. How is the formation of the two types of protrusions similar? They’re dependent on actin formation and driven by forces made by actin polymerization. 16. What are the two functions of a focal adhesion? Attachment forms stable interactions with the extracellular matrix and signaling deals with moving and cell division. 17. What type of enzyme is required for focal adhesion detachment? Proteases (calpain) 18. Which two steps of ameboid movement require the establishment of ligand/receptor binding? What are the ligands? 1 and 3, the ligands bind to the extracellular matrix 3 BIOL230W Week 12 – Cell Motility Small Group Activity: Ameboid Movement- Use the following images to draw in the components for each stage. = actin = microtubules The cell to the left is non-motile. It could be a homeostatic fibroblast or a non-cancerous ductal epithelial cell. Draw the actin and microtubules in this cell. The cell to the left has undergone polarization in response to a negative or repelling signal. Indicate the location and organizational pattern of actin and microtubules, and the location of the signal. Indicate the leading and trailing edges. The cell to the right is starting to move by extending protrusions. Indicate the type of protrusion, the leading and trailing edges, the actin and the microtubule location and organizational pattern. 4 BIOL230W Week 12 – Cell Motility The cell to the left has formed focal adhesions (FA) at the tips of its protrusions. Indicate their location. As the cell goes through the process of contraction the cell will pull itself across the focal adhesions until FAs that were at the front of the cell are now at the back of the cell. Indicate the location of the same focal adhesions as the cell moves. Finally, indicate the location of a FA that needs to undergo detachment. Day 3: Metastasis Learning Objectives 1. Compare and contrast processes of wound repair and metastasis. 2. Compare and contrast the epithelial cell and the mesenchymal cell with respect to the EàM transition. Include cadherins, cell junctions, actin and the ECM components collagen and fibronectin. 3. Describe the two roles of ECM degradation and remodeling in tumor invasion. 4. Analyze data relating the EMT or motility to metastasis. 5 BIOL230W Week 12 – Cell Motility Small Group Activity: Metastasis 19. Based on what you learned about wound repair, what do the small orange rectangles on the plasma membrane of the epithelial cells represent? Cell junctions 20. What does the absence of those structures suggest about how the E à M transition enables metastasis? Junctions hold cells together, therefore when they disappear the cell gains motility, so cell can invade other areas. 21. Recalling the role of the cytoskeleton in motility, what might be happening to the actin in the transitioning cells? Anchoring promoted by actin stress fibers. Mini-Lecture Metastasis 22. What are two effects of ECM degradation? Increased cell motility and removal of basement membrane 23. Targeting of what two molecules by MMPs promotes invasiveness in carcinomas? 6 BIOL230W Week 12 – Cell Motility Collagen 4 and E-cadherin Small Group Activity: Metastasis Data Analysis The following image is measuring the levels of protein in two different tissue samples. One is from a ductal breast cancer tumor and one is from non-cancerous tissue. Examine the data and then answer the following questions. 24. Based on this data, which cells have lost cell-to cell connections (A or B)? B 25. Based on what we now know about the role of cell junctions and the extracellular matrix in metastasis, which of the cell lines in this image (A or B) were taken from a metastatic tumor and which was taken from non-cancerous tissue? B is metastatic tumor, A is non-cancerous 26. Vimentin inhibits the transport of one of the cadherins to the plasma membrane. Which cadherin do you think it would be (E or N)? E-cadherin