NROS 310 Exam 3
Outline the signaling events that allow the elongation of a neuron.
NGF→phosphorylating RTK→PI3K→PIP3→CDC42GEF→CDC42→WASP & ARP2/3→actin polymerization draw!
What are the 4 steps necessary for cell movement? Outline the signaling pathways that underlie those 4 steps. Include an activation of WASP and increasing Ca++ levels.
1) extend lamelipod (actin jet pathway) 2) form attachment (focal adhesion 3) translocation using actin myosin interactions 4) break old attachemnts as translocation happens by breaking down actin
If you treat a moving neutrophil with cytochalasin (preventing actin polymerization) it loses its ability to follow a bacterium. A.) Why is this? B.) What is a signaling pathway that might lead from detection of a bacterial protein to changes in the cytoskeleton? C.)Are the cytoskeletal changes localized or do they take place in the entire cell? Why might this be important?
A) actin polymerization = actin jet = motility B) draw C) cytoskeleton changes are localized which is important for cell to move/ grow in particular directions
You have created a new transgenic mouse in which the troponin complex is nowindependent of calcium regulation. What would happen in the skeletal muscles of these mice if troponin was A.) never activated or B.) always activated.
A) contraction would never occur B) contraction would always occur given there was sufficient ATP
Beta catenin is a protein that functions both as part of the cytoskeleton and as a transcription factor. If beta catenin is mis-regulated, it can lead to colon cancer. Outline the role of beta catenin in junctional complexes, its role in regulation of transcription, and its regulation by the APC- GSK complex. Describe whether the following homozygous mutations or cellular treatments would lead to an increased or decreased risk of colon cancer? Explain why in a sentence or two. A. Beta catenin that could not function as a transcription factor B. Treatment of cells with a soluble cadherin. C. A non-functional APC D. Beta catenin that could not be phosphorylated by GSK
A) decreased risk because cell would not get the signal B) incresed risk because bet acatenin will be released C) increased risk because cell would keep dividing D) decreased risk because phosphorylaton will break down beta catenin and therefore wont act as a TF
Which microtubule-motor protein(s) is (are) most likely to mediate the following movements: A.) Movement of vesicles from the E.R. to the golgi. B.) Movement of vesicles from the golgi to the plasma membrane C.) Movement of vesicles from the cell body to the distal tip of an axon. D.) Movement of a vesicle from the cell body to the distal tip of the dendrite.
A) dynein B) kinesin and myosin C) kinesin D) Both kinesin and dynein
Predict the effects of adding soluble (not membrane attached) cadherins to cells that are held together using A.) adherens junctions B.) tight junctions C.) desmosomes
A) fall apart. Beta catenin complex falls apart and goes to the nucleus to act as TF and the whole thing falls apart B) nothing C) fall apart
Rigor mortis is the stiffening of the muscles that occurs after death. It results from an increase in the level of Ca++ and a lack of ATP in the dead muscle cells. (Assume that all of the protein components of the muscle remain intact.) A.) Why do the increased Ca++ levels and low ATP levels cause muscle stiffness? B.) What would happen if you tried to reanimate the tissue by decreasing calcium and increasing ATP levels in the muscle cells?
A) high calcium levels make it so that binding sites on actin are always available for myosin to attach but the lack of ATP makes it so that myosin in unable to be released B) This woudl just allow the muscles to relax because the ATP will make myosin detach and without sufficient Ca, troponin/ tropomyosin will cover up the binding sites indefinitely
Describe the functions and proteins that make up the following cell junctions: A.) Gap Junctions - B.) Desmosomes - C.) Tight Junctions - D.) Adherens Junctions - cadherins and actin, cell to cell interactions E.) Focal Adhesions F.) Hemi-Desmosomes -
A) highway to communicate- connexins B) caldherins and IF, cell to cell interactions C) claudin and occludin keeps tissues together, prevent anything from crossing D) cadherins and actin, cell to cell interactions E)Actin and integrin, anchors actin filaments in cell to ecm F) IF and integrin, anchors intermediate filalments to ecm
You are studying 2 different types (red and blue) of growing neurons in culture. You find that red neurons grow only on laminin stripes and blue neurons grow only on fibronectin stripes. A.) What is a possible difference between red neurons and blue neurons that accounts for their behavior in this assay. B.) What junctional complexes will be formed as the growth cones from the red neurons navigate on the laminin stripes.
A) integrin can have specificity of what it can bind to B)focla adhesion would be the junctional complex
For either of the above pathways, predict the effects of the following treatments/mutations on the ability of the listeria or lammelipodium to move. Explain each answer in a sentence. What is the normal function of the affected protein? A.) Defective Arp2/3. B.) Defective Profilin. C.) Lots of extra alpha-actinin. D.) Lots of extra ADF. E.) Lots of extra gelsolin. F.) Nonhydrolyzable ATP
A) no nucleation, go nowhere B) compromised ATP so slows down C) not a whole lot to help it move any better D) destabilize the platform E) needs high levels of Ca to do anything F) You wouldnt have enough monomers
Outline the components necessary for the actin-polymerization based movement of listeria.
Act-A hijacks cell's actin jet machinery→ activates Arp ⅔→ capping of (+) end and nucleation→ filament grows and becomes stable→ capping of (-) end→ cofilin(ADF) breaks stable filament and produces actin-ADP monomers→ profilin converts actin-ADP to actin-ATP
What are the differences in structure and function between myosin V and myosin II?
Both have two heads but myosin II has an extended neck whereas myosin V has a shorter neck but two feet. II = contraction V = transportation of vesicles
Describe the basic properties of cadherins, IG molecules, selectins, and integrins.
Caldherins = homophilic interactions where caldherins on one cell interact with caldherins on another cell (cell to cell) - Adherins junctions: caldherins can hook up with actin (make a broader interaction) -Desmosomes: caldherins and intermediate filaments (making a broader interaction) IG molecules: Homophilic interaction; cell to cell interaction Inegrins = connects the cells to the extracellular matrix -focal adhesion: integrins and actin -hemidesmosome: integrins and intermediate filaments
Diagram the components of a sarcomere in skeletal muscle. Identify the proteins involved including actin, myosin,titin, CapZ, and tropomodulin. B.) If you looked at an electron micrograph of skeletalmuscle, could you tell if the muscle was contracted or relaxed? What would you see ifthere was a difference? C.) Predict the functional effects of a defect in titan
Cap Z binds the plus end of actin to the Z disc, tropomodulin binds the minus end of actin, titin binds myosin to the Z disc. B) you could tell if a mmuscle is contracted because you would see less lighter color thin filament sections and more darker sections where actin and myosin overlap C) a defect in titan would make it so that if the muscle was stretched too far, myosin would be out of reach to attach to actin and the cell would not be able to contract
What are the main functions of the different classes of actin associated proteins?
Capping proteins= prevents assembly and disassembly at plus end Tropomodulin= prevents assembly and disassembly at minus end Gelsolin= severs filaments and binds to plus end Cofilin= binds ADP-actin filaments and accelerates disassembly Profilin= bind subunits, prevents assembly Thymosin= binds subunits and prevents assembly WASP + ARP 2/3= nucleates assembly and remains associated with minus end
What are the basic components of the extracellular matrix?
Collagen, GAGs, elastinsn and cross linked proteins
What is meant by a critical period and why is it important when thinking about CNS injury repair?
Critical period is where developmental changes are more likely to happen. This is important in terms of injury to CNS because if myelination has already development regeneration can not tke place
Outline the signal transduction cascade that allows a neutrophil to follow a bacterium.
Draw
Dynamic instability is a characteristic of growing microtubules. What is it? What is the significance of GTP cap formation? How does the process of cap formation differ from that in actin filaments?
Dynamic instability of microtubles is the same concept as with actin polymerization but with GTP instead of
What properties of glycosaminoglycans help to lead to the proper functioning of cartilage?
GAGs bring lubrication and shock absorption
What is the role of nexin in cilia and flagella?
Holds microtubules in place through crosslinking
Describe the effects attractive factors have on the cytoskeleton. Include a likely signal transduction cascade for the attractive factor.
NGF→phosphorylating RTK→PI3K→PIP3→CDC42GEF→CDC42→WASP & ARP2/3→actin polymerization draw!
Why are there motor proteins associated with both actin filaments and microtubules but not intermediate filaments?
Intermediate filaments are stationary and are more for stable structure rather than movement
What components make up the MTOC (microtubule organizing center)? What is its main function? Do all MTs use MTOCs?
MTOC is made up of gama tubulin and centrioles. The function is to organize microtubules in such way that it creates a stable highway for things to be transported throughout the cell
Describe how intermediate filaments are generated. Why is there no polarity associated with intermediate filaments?
Made of multiple strands of fibrous proteins wound together. Assembled intermediate filaments have no polarity because individual monomers are oriented in both directions along the axis of the filament.
What are the three main components of the cytoskeleton and what are their main functions?
Microfilaments: movement microtubules: transport of cellular materials intermediate: support
What are neurofibrillary tangles and where do you find them?
Neurofibrillary tangles are insoluble twisted fibers found inside neurons. They are composed of highly phosphorylated forms of the microtubule associated protein tau
What are the purposes of profilin and thymosin?
Profilin: phosphorylates ADP to ATP/ GDP to GTP Thymosin: Lowers monomer concentration by pumping out monomers from cell
Describe how the assembly of actin filaments is regulated. Include both the role of associated protein and monomer concentration.
Rac/Rho/CDC43 trigger Arp2/3 to nucleate→ capping of (+) end→ grow filament via actin-ATP monomers→ stable filaments→ cap (-) end→ ADF breaks filament to actin-ADP monomers→ profilin phosphorylates
What is main function of intermediate filaments? Why does their structure made them well suited for this function?
Rope-like, flexible, support only. They are specialized to bear tension, and their jobs include maintaining the shape of the cell and anchoring the nucleus and other organelles in place.
Describe the effects repulsive factors have on the cytoskeleton. Include a likely signal transduction cascade for the repulsive factor.
Sema3A causes collapse of growth cone vis phosphorylation of RTK to activate CDC42GAP which inhibits CDC42
Describe how an increase in calcium causes the contraction of skeletal muscle and smooth muscle. What are the key differences?
Skeletal: Calcium binds to troponin which then pushes tropomyosin out of the way so that the binding sites on actin are exposed and myosin attaches. Smooth: Calcium and CaM turn MLCK active which then phosphorylates Myosin LC and contraction takes place
What is the clinical significance of slow axonal transport?
This is the rate limiting step of axon regeneration
Define critical concentration.
The concentration where the probability of addition happens just as easily as falling off
Why is it important that kinesin and dynein always have one motor head group attached to the microtubule?
To make sure it continues to go in the right direction and doesnt fall off
Why are PNS neurons able to regenerate while CNS neurons are not?
They myelination in PNS and CNS are different in such a way that the proteins in the CNS myelin shut down growth cones
What would be the effect of incorporating a non hydrolyzable analog of ATP in the assembly process?
You would not produce any more actin monomers to feed the actin jet
What is the basic structure of GAGs? What features of the structure of hyaluronan lead to the proper functioning of cartilage?
a lot of sugars with a littl ebit of protein attached. The hyaluronan is very charged so it brings a lot of water along to provide shock absorption
Why are there motor proteins associated with both actin filaments and microtubules but not intermediate filaments?
actin filaments and microtubules construct a highway for motor proteins while intermediate filaments are used for structural support
What are the main similarities and differences between the three components of the cytoskeleton?
all are important for cytoskeleton structure differ in size
Both skeletal muscles and cilia and flagella use motor proteins and cytoskeletal elements to move cells. What is similar and what is different about these two processes?
both are processes of movement. skeletal = actin and myosin cilia/ flagella = kinesin and dynein
Colchicine inhibits microtubule formation while taxol stabilizes microtubules. Why do both drugs cause problems for cells?
both drugs regulate micrtubule growth/ formation that can have negative effects
How can the differential strength of binding between cells be used to organize tissues?
caldhrins prefer to link with other caldherins of the same type and same level, which sorts them out on their own
What is the function of cilia and flagella?
cell movement
What properties of collagen lead to the proper functioning of cartilage?
collagen is super strong which is able to keep cartilage together
What are the functions of nogo and MAG? Why are they a problem for CNS regeneration?
components present in CNS myelin that shut down cone growth
A.) Outline the components necessary for the extension of lamellipodium in a moving cell.
draw?
What is the motor protein that drives movement is cilia?
dynien
What motor protein(s) are thought to drive fast axonal transport? What motor protein(s) are thought to drive slow axonal transport? What is the evidence that supports this idea?
fast = kinesin down axon, dynein back up slow = kinesin only evidence = if you knock out dynein you lose fast transport from axon to cell body, if you knock out kinesin you lose fast transport from celly body to axon and all of slow transport
What types of molecules are transported by fast axonal transport? What types of molecules are transported by slow axonal transport?
fast = neuropeptide in vesicles, mitochondria, naything packaged in vesicels slow = everything else like kinases, phosphatases, soluble enzymes
Why are focal adhesions more common in moving cells and hemidesmosomes more common in stationary epithelial cells?
focal adhesions form an attachment point that allows the cell to start to move and better supports dynamic actin hemidesmosomes contain intermediate filaments which are not dynamic
What features of collagen lead to its high tensile strength? Why does scurvy cause connective tissue problems? Describe the process of collagen synthesis.
glycines and prolines are hydroxylated which forms a tight weave of the collagens. They are made by making individual chains, hydroxylating some of the prolines and lysines forming procollagen which is secreted out of the cell and used to make collagen fibril. Scurvy causes problems because there is no vitamin C which is necessary cofactor allowing for modification of proline to hydroxyproline so collagen is much less fragile
What is the role of gamma tubulin?
helps with initiating nucleation of microtubules and binds to the minus end
What is the evidence that vesicles need to be transported by more than one type of motor protein during exocytosis?
if you knock out myosin exocytosis doesnt happen but even if you actually look at the microtubule, it doesnt reach the plasma membrane so something else much help it get there
How does high calcium trigger the destruction of old attachments?
increased calcium triggers gelsolin which breaks down ctin and destorys attachments
What are the main functions of kinesin and dynein? How are these motor proteins different from myosin II?
kinesin = plus end directed motor dynein = minus end directed motor these motors are always attached to microtubule whereas myosin II is not always attached to actin
Name and describe the functions of two microtubule associated proteins.
kinesin, dynein are both motor proteins that differ in their direction
Describe how cadherins mediate cell interactions and how those interactions can be regulated.
mediates interactions by binding to other caldherins. They are regulated by calcium, which binds to the hinge regions and allows the caldherin domains to extend. If you lower the extracellular calcium the cells will eventually fall apart
Why is the nucleation of microtubules more difficult than nucleation of actin filaments? How is this difficulty reflected in the organization of the microtubule network?
microtubules have much more complicated and organized structure which makes nucleation that much more difficult because the rate limiting step of monomers randomly adding in the exact right orientation has to be even more intentional than in actin polymerization
How does the hydrolysis of ATP cause myosin to move along the actin filament?
myosin heads hydrolyze ATP and become reoriented and energized
Would you expect components of the desmosome such as the adapter proteins plakoglobin and desmoplakin to have signaling functions such as seen for beta catenin? Why or why not? What is the point of having a protein function both as an adapter in junctional complexes and as a transcription factor?- essentially its used as back up if one dies
no because beta catenin is a specialized protein specific to WNT pathway The point of having a protein function as an adapter and TF is so that if one cell dies the other can take over
Nerve growth factor can cause neurons to move and it also helps them to survive by preventing apoptosis. How can the same molecule perform both functions?
not on exam
Outline the steps necessary for leukocyte extravasation.
not on exam
What is inside out integrin signaling and outside in integrin signaling?
not on exam
What is the rate-limiting step in actin filament assembly?
nucleation
How do the ends of the actin filament differ?
plus end vs minus end each have their own concentration and probability of adding or taking off monomers Also establishes a sense of direction for motor proteins to act on
What is the function of elastin?
provides elasticity within connective tissues, especially in vessels
Why is it important to both form and destroy cytoskeletal attachments to the membrane and the substrate?
so the cell isnt stuck in any one place or direction
How do the properties of actin subserve both stable and dynamic functions in the cell?
the ability to be phosphorylated to make stable polymers and networks but also the ability to be broken down
What is dynamic instability? How does the regulation of ATP hydrolysis figure into the process?
the actin-ATP monomers within the polymer eventually become hydrolyzed that makes it very difficult to add on and an overall unstable structure to hold together
What are the main functions of laminin and fibronectin?
they are crosslinking proteins fibronectin = hold ecm together and provides a wy for cells to hold onto it laminin = in the basal lamina holds things in the sheet-like complex
Why is it important that each individual myosin head group is only transiently attached to the actin filament?
to allow the muscle to be relaxed/ stretch/ not be contracted all the time
Why is it important for cells to regulate the activity of integrins? How do cells accomplish this?
to help determine how sticky the cell is by secreting inegrins that the cell can bind to and dissociate from the ecm
Describe how the formation of a focal adhesion triggers the contraction of a neutrophil.
translocation uses actin and myosin interactions by increasing calcium, which activates MLCK which allows myosin to cause contraction toward extension
What is the difference between collagen IV and the other collagen isoforms? Why is collagen IV particularly well suited to the basal lamina?
type IV = sheet like network. They form a basal lamina that forms under epithelial tissue. isoforms = collagen fibril rope like structure