Chapter 16: Cytoskeleton

Indicate whether each of the following structures has actin organized mostly in a one- (1), two- (2), or three- (3) dimensional arrangement. Your answer would be a four-digit number composed of digits 1 to 3 only, e.g. 1333. ( ) Stress fibers ( ) Invadopodia ( ) Lamellipodia ( ) Filopodia

1321

A dimeric kinesin-1 molecule has 8-nm steps and can move at rates of about 1 µm/sec. Olympic 100-meter sprinters typically run at about 180 steps per minute and can reach speeds of about 42 km/h. With the same step size, if the Olympic runner had a step frequency of a kinesin-1 molecule, how fast could she run? Write down your answer in km/h, e.g. 52 km/h.

1750 km/h

You have used "optical tweezers" to study the mechanics of myosin movement in a newly discovered member of the myosin II subfamily. Your results indicate that each myosin exerts a force of about 3 pN along the length of the actin filament, and displaces the filament by about 10 nm in each cycle of ATP hydrolysis. Assuming that the free-energy change for ATP hydrolysis is -50 kJ/mol under your experimental conditions, what is the efficiency of the myosin motor in converting the free energy to mechanical work? Remember that under a constant force F (in newtons), work (W; in joules) is calculated as W = F × d, where d is displacement (in meters) in the direction of the force. Avogadro's number is approximately 6 × 1023 molecules/mole. Write down your answer as a percentage with no decimals, e.g. 99%.

36%

Indicate if each of the following descriptions applies to (1) EB1, (2) kinesin-1, (3) kinesin-13, or (4) katanin. Your answer would be a four-digit number composed of digits 1 to 4 only, e.g. 1432. ( ) This is a microtubule-severing protein that can release microtubules from the microtubule-organizing centers. ( ) It increases the frequency of catastrophe by deforming microtubule protofilaments. ( ) This is a conventional motor protein that moves toward the plus end of a microtubule. ( ) It recognizes the structure of a growing microtubule end and binds to it, helping other proteins to also bind to the plus end.

4321

A microtubule appears as a left-handed helix due to an approximately 0.9-nm stagger in the lateral contacts between adjacent protofilaments. In the lateral contacts, α- and β-tubulins in one protofilament interact with α- and β-tubulins, respectively, in the neighboring protofilament, except for a longitudinal discontinuity along the microtubule called the seam. Along the seam, lateral contacts have to be made between different tubulins (i.e. α-tubulin with β-tubulin). Which of the following do you think is acceptable as the repeat distance of tubulin monomers along a protofilament? A. 4 nm B. 5 nm C. 6 nm D. 7 nm E. 10 nm

A

Consider the structure of a sarcomere. Which of its features does NOT shorten during skeletal muscle contraction? A. The dark band B. The light band C. The distance from the M line to the Z disc D. The distance between two consecutive Z discs E. The extension of the titin molecules

A

In the polymerization in vitro of actin filaments and microtubules from their subunits, what does the "lag phase" correspond to? A. Nucleation B. Reaching steady state C. Nucleotide exchange D. ATP or GTP hydrolysis E. Treadmilling

A

In the presence of an ATP analog that can bind myosin normally but cannot be hydrolyzed, ... A. a myosin cannot bind tightly to an actin filament and cannot move on it. B. a myosin cannot be released from an actin filament and cannot move on it. C. a myosin performs the power stroke but cannot be released from an actin filament. D. a myosin performs the power stroke but cannot bind tightly to an actin filament. E. a myosin is able to perform normally.

A

In the structure of which of the following proteins are the actin-filament binding sites furthest apart in space? A. Spectrin B. Filamin C. α-Actinin D. Fimbrin E. Myosin II

A

The γ-tubulin ring complex is to microtubules what ... is to actin filaments. A. the Arp 2/3 complex B. the dynactin complex C. the troponin complex D. formin E. contractile ring

A

Unlike a myosin head, a kinesin head is tightly bound to its cytoskeletal track when bound to ... A. ATP. B. ADP. C. no nucleotide. D. GTP. E. GDP.

A

Indicate if each of the following structures is based on actin filaments (A), microtubules (M), or intermediate filaments (I). Your answer would be a five-letter string composed of letters A, M, and I only; e.g. AAAMM. ( ) The cell cortex ( ) The mitotic spindle ( ) The nuclear lamina ( ) Cilia ( ) Filopodia

AMIMA

Indicate if each of the following changes occurring during mitosis in a fibroblast is the result of the reorganization of actin filaments (A), microtubules (M), or intermediate filaments (I). Your answer would be a five-letter string composed of letters A, M, and I only; e.g. AIAAM. ( ) The cell rounds up. ( ) The endoplasmic reticulum collapses. ( ) The Golgi apparatus fragments. ( ) The primary cilium is resorbed (disappears). ( ) The contractile ring forms and constricts.

AMMMA

Cofilin binds preferentially to ADP-containing actin filaments rather than to ATP-containing filaments. Consequently, this protein ... A. competes with profilin for actin binding. B. binds to older actin filaments. C. binds to the plus end of actin filaments. D. stabilizes actin filaments. E. All of the above.

B

Dynamitin is a subunit of the dynactin complex. Its overexpression leads to the disassembly of the complex, and it is therefore considered a dynactin inhibitor. Which of the following processes would happen if dynamitin is overexpressed in a cell? A. The motile cilia (if any) would stop beating because the axonemal dyneins could not attach to their neighboring microtubules. B. The Golgi apparatus would fragment and become dispersed because the cytoplasmic dyneins could not associate with the Golgi membranes. C. The endoplasmic reticulum would collapse because it could not recruit enough kinesins. D. The nuclear envelope would disintegrate because the nuclear lamins could not interact with the cytoplasmic cytoskeleton. E. The cortex would lose its integrity because the Arp 2/3 complex would also be inhibited.

B

In the presence of ATP in a flagellum, an axonemal dynein that is interacting through its tail with the A microtubule of a peripheral doublet can push this doublet toward the ...(1) of the flagellum, but due to the presence of linking proteins such as ...(2), this force is converted into a bending motion. A. (1) tip; (2) nexin B. (1) base; (2) nexin C. (1) tip; (2) nebulin D. (1) base; (2) nebulin E. (1) tip; (2) tau

B

Keratins are intermediate filaments that ... A. are localized inside the nucleus. B. are composed of 50% type I and 50% type II keratin proteins. C. are bundled in the epidermis to make cells more flexible. D. anchor the intermediate filament network at the adherens junctions. E. All of the above.

B

The actin-nucleating protein formin has flexible "whiskers" containing binding sites that help recruit actin subunits in order to enhance polymerization by this protein. What protein would you expect to bind to these sites? A. Thymosin B. Profilin C. Cofilin D. Gelsolin E. Tropomodulin

B

What is the major microtubule-organizing center in animal cells? A. The γ-tubulin ring complex B. The centrosome C. The cell cortex D. The primary cilium E. The spindle pole body

B

Which of the following proteins do you expect to be enriched near the plus end of microtubules? A. Dynein B. XMAP215 C. γ-Tubulin D. Katanin E. All of the above

B

"Headless" kinesin mutants only contain the stalk (middle) and tail domains and can therefore dimerize with their wild-type kinesin partners. However, since they lack the motor (head) domain, the resulting dimers are unable to carry out processive transport of their cargoes and the mutation thus behaves as "dominant negative," meaning that the mutant not only is nonfunctional, but can also interfere with the function of its wild-type counterparts. If a headless mutant of a kinesin heavy chain involved in melanosome movement is overexpressed in fish melanocytes, what would you predict happens in these cells? A. Pigment dispersion would be inhibited and there would be more tug-of-war between the motors. B. Pigment aggregation would be inhibited and there would be more tug-of-war between the motors. C. Pigment dispersion would be inhibited and there would be less tug-of-war between the motors. D. Pigment aggregation would be inhibited and there would be less tug-of-war between the motors.

C

Actin filaments that are held together by the cross-linking protein fimbrin are not contractile. This is probably because ... A. the very weak cross-linking by this protein cannot convert myosin II activity into a coherent contraction. B. fimbrin arranges the actin filaments in parallel bundles in which all the plus ends point to the same direction. C. the very tight packing of actin filaments by this small protein excludes myosin II filaments and other large proteins. D. fimbrin arranges the actin filaments into gel-like networks in which myosin II activity does not produce contraction. E. fimbrin is a large protein that binds to several actin filaments and resists contraction.

C

After an animal dies, its muscles start to stiffen before the decomposition of tissues relaxes the muscles again. Which of the following would you expect to explain this muscle stiffening (i.e. rigor mortis)? A. The myosin II heads in muscle fibers remain attached to actin filaments due to the absence of Ca2+ in these cells. B. The ATPase activity of muscle myosin II is inhibited by the elevated Ca2+. C. The myosin II heads in muscle cells remain attached to actin filaments due to the elevated Ca2+ in the muscle fibers. D. Titin molecules unfold, preventing muscle relaxation. E. The myosin II heads in muscle cells remain attached to actin filaments due to covalent cross-linking.

C

If myosin II heads are attached to a glass slide and actin filaments are allowed to bind to them, the filaments will glide on the surface ... A. toward their plus end in the presence of ATP. B. toward their plus end in the presence of GTP. C. toward their minus end in the presence of ATP. D. toward their minus end in the presence of GTP. E. toward their plus end in the presence of ADP.

C

In contrast to growing microtubules, shrinking microtubules ... A. have a GTP cap at their plus end. B. have strong lateral interactions at their plus ends. C. have curved protofilaments at their plus ends. D. cannot be rescued unless microtubule-stabilizing proteins bind and inhibit depolymerization. E. All of the above.

C

The duty ratio for a motor protein is defined as the fraction of time in each cycle of activity of the motor in which the head is bound with high affinity to its cytoskeletal track. Which of the following is reasonable given this definition? A. Myosin V has a higher duty ratio than myosin II, because Pi release is the rate-limiting step in its cycle. B. Myosin II has a higher duty ratio than myosin V, because Pi release is the rate-limiting step in its cycle. C. Myosin V has a higher duty ratio than myosin II, because ADP release is the rate-limiting step in its cycle. D. Myosin II has a higher duty ratio than myosin V, because ADP release is the rate-limiting step in its cycle.

C

Indicate whether each of the following descriptions applies to cilia (C), flagella (F), or both (B). Your answer would be a four-letter string composed of letters C, F, and B only, e.g. CCFB. ( ) They are short and present at high numbers per cell. ( ) They have a whiplike motion that resembles breaststroke in swimming. ( ) They are based on the axoneme structure. ( ) They are found in the epithelial cells of the human respiratory tract.

CCBC

Bacteria contain homologs of cytoskeletal filament subunits ... A. except those of the intermediate filaments. B. but these homologs are incapable of nucleotide binding and hydrolysis. C. that are less diverse in their function relative to their eukaryotic counterparts. D. that can have different functions to those of their eukaryotic counterparts. E. that are dispensable for cell growth and proliferation.

D

Consider an actin subunit that has just been incorporated into an actin filament at the leading edge of a lamellipodium. Before its ATP is hydrolyzed, how does its distance from the leading front edge of the plasma membrane change over time? How does its distance from the F-actin minus end change over time? A. Decreases; decreases B. Decreases; remains constant C. Decreases; increases D. Increases; decreases E. Increases; remains constant

D

How is membrane protrusion by blebbing different from that by lamellipodia? A. Blebbing usually occurs on a rigid substratum such as glass, whereas lamellipodia form on pliable substrata. B. Blebbing is mostly observed in vitro, whereas lamellipodia are observed both in vitro and in vivo. C. Blebbing does not involve myosin II activity, but formation of lamellipodia does. D. Blebbing requires loss of membrane interaction with actin filaments, whereas lamellipodia require a close interaction between the two.

D

If a certain isoform of myosin II is depleted from a cell, stress fibers are lost and focal adhesions disappear. If these cells are placed on a surface coated with an array of flexible pillars to measure traction forces, would you expect the traction to increase (I) or decrease (D) in these cells compared to wild-type cells? Write down I or D as your answer.

D

If the concentration of free subunits is C, under which condition does the growth of a cytoskeletal filament proceed spontaneously? A. C > 1/kon B. C < 1/koff C. C > kon/koff D. C > koff/kon E. C > Cc×kon/koff

D

In lamellipodia, actin polymerization is nucleated by ...(1), while depolymerization is catalyzed by ...(2). A. (1) formin; (2) gelsolin B. (1) Arp 2/3 complex; (2) gelsolin C. (1) formin; (2) cofilin D. (1) Arp 2/3 complex; (2) cofilin E. (1) formin; (2) capping protein

D

In the dendritic networks of actin filaments in lamellipodia, nucleation of actin polymerization is mostly performed by ... A. ERM proteins B. WASp proteins C. Formin D. Arp 2/3 complex E. γ-TuRC

D

Persistence length for a cytoskeletal filament is the minimum filament length at which random thermal fluctuations are likely to cause it to bend. Which of the following comparisons are true, considering the persistence lengths of (a) an actin filament, (b) a bundle of cross-linked actin filaments, and (c) a microtubule? A. (a > b) and (a > c) B. (a > b) and (a < c) C. (a < b) and (a > c) D. (a < b) and (a < c) E. (a > b) and (b > c)

D

What are the typical consequences of mutations in keratins and their associated proteins? A. Neurodegeneration as a result of interference with normal axonal transport. B. Muscle development defects as a result of sarcomere disorganization. C. Skeletal and cardiac abnormalities as a result of a weakened nuclear envelope. D. Cornea disorders as a result of cell rupture from mechanical trauma. E. Developmental defects as a result of abnormal signaling related to the primary cilia

D

Which of the following cytoskeletal filaments are abundant in an animal cell nucleus? A. Microfilaments B. Microtubules C. Septins D. Intermediate filaments E. Spectrin filaments

D

Sort the following events to show the sequence in the mechanochemical cycle of myosin II, following ATP binding by the head domain. (A) Myosin head binds tightly to actin. (B) Power stroke is triggered. (C) Cocking occurs and the head is displaced along the actin filament. (D) The binding affinity of myosin for actin is reduced.

DCAB

Indicate whether each of the following descriptions applies to myosins (M), kinesins (K), or dyneins (D). Your answer would be a five-letter string composed of letters M, K, and D only, e.g. MMMDD. ( ) They have larger structures than the other two. ( ) They are generally faster than the other two. ( ) They are structurally unrelated to the other two. ( ) They walk on a different cytoskeletal filament than the other two. ( ) They are all minus-end directed.

DDDMD

Sort the following events to reflect the sequence during smooth muscle contraction upon delivery of muscle stimulation. Your answer would be a five-letter sequence composed of letters. (A) Activation of the myosin light-chain kinase (B) Interaction of myosin head with actin (C) Phosphorylation of myosin (D) Calcium release into the cytoplasm (E) Activation of calmodulin

DEACB

Skeletal muscle cells consume enormous amounts of ATP. From the following list, choose the two proteins that directly hydrolyze most of this ATP. Your answer would be a two-letter string composed of letters A to F only, in alphabetical order, e.g. AF. A. Troponin complex B. Ca2+-release channel C. Myosin light chains D. Myosin heavy chain E. Myosin light-chain kinase F. Sarcoplasmic reticulum Ca2+-pump

DF

A small unique insert found near the end of the motor domain in myosin VI has been linked to the singular ability of this myosin to move toward the pointed end of actin filaments. If the unique insert is removed by genetic engineering, the resulting mutant myosin VI is a plus-end directed motor. This observation indicates that ... A. the unique insert is necessary and sufficient for minus-end directionality. B. the unique insert is necessary for motor activity. C. the unique insert is necessary for plus-end directed directionality. D. the unique insert is not sufficient for minus-end directionality. E. None of the above.

E

F-actin is not a straight polymer but can be considered a double-helical assembly. The two protofilaments twist around each other and cross over approximately every 13 actin subunits, and the subunits in each protofilament are repeated every 2.8 nm along the helix. F-actin is almost 8 nm wide. Different myosin motors have different "step lengths" on actin depending on their function. Which of the following step lengths would you expect to belong to a processive myosin motor that carries large cellular cargoes such as endocytic vesicles? A. About 2.8 nm B. About 8 nm C. About 13 nm D. About 27 nm E. About 36 nm

E

In which of the following structures are actin-myosin II bundles regulated by the troponin complex? A. Stress fiber B. Circumferential belt C. Contractile ring D. Adherens junction E. None of the above

E

SUN and KASH proteins embedded in the nuclear envelope provide connections between the organization of the nucleus and the cytoplasm. Which of the following pairs of proteins DO NOT bind directly to each other in these connections? A. Nuclear lamina and SUN proteins B. Plectins and KASH proteins C. KASH proteins and microfilaments D. KASH proteins and SUN proteins E. SUN proteins and cytoplasmic intermediate filaments.

E

The proteins of the ERM family such as moesin ... A. bind to and organize the cortical actin cytoskeleton. B. interact with transmembrane proteins. C. affect cortical stiffness and cell shape. D. affect the localization and activity of cell signaling molecules. E. All of the above.

E

These proteins are found in the budding yeast Saccharomyces cerevisiae; they form "neck filaments" between a mother cell and its growing bud, and help polarize protein distribution between the two. These proteins ... A. can polymerize to form filaments and sheets. B. bind GTP. C. are also involved in contractile ring formation during cytokinesis in animal cells. D. form filaments that are thought to be nonpolar. E. All of the above.

E

Which of the following actin-binding proteins cannot bind to the same actin filament simultaneously? A. Gelsolin and tropomodulin B. Tropomyosin and tropomodulin C. Profilin and tropomodulin D. Cofilin and CapZ E. Formin and CapZ

E

Which of the following drugs is toxic for our cells? A. Cytochalasin B, which caps the plus end of actin filaments and prevents actin polymerization. B. Phalloidin, which binds along actin filaments and stabilizes them. C. Nocodazole, which binds to tubulin subunits and prevents microtubule polymerization. D. Colchicine, which caps microtubule ends and leads to their depolymerization. E. All of the above.

E

Which of the following is NOT an example of a crawling cell? A. Macrophage B. Osteoclast C. Keratocyte D. Neural crest cell E. Sperm

E

Which of the following is an actin homolog? A. Arp2 B. Arp3 C. MreB D. ParM E. All of the above

E

Indicate true (T) and false (F) statements below regarding cytoskeletal motor proteins. Your answer would be a five-letter string composed of letters T and F only, e.g. TTTFF. ( ) All myosin motors move toward the plus end of actin filaments. ( ) All myosin motors move toward the minus end of actin filaments. ( ) All kinesin motors move toward the plus end of microtubules. ( ) All kinesin motors move toward the minus end of microtubules. ( ) All dynein motors move toward the minus end of microtubules.

FFFFT

The pathogen Listeria monocytogenes can hijack the actin cytoskeleton in human cells and spread inside the host. Indicate true (T) and false (F) statements below about this phenomenon. Your answer would be a four-letter string composed of letters T and F only; e.g. TTFF. ( ) The movement can be reconstituted in vitro by placing the bacteria in a mixture of actin, formin, gelsolin, and capping protein. ( ) Cofilin counteracts the movement by depolymerizing actin filaments. ( ) The actin filaments grow with their minus ends pointed toward the bacterium. ( ) The movement depends on myosin activity to transport the bacteria on the actin filaments.

FFFT

Indicate whether each of the following descriptions matches Cdc42 (C), Rac (R), or Rho (H) from the Rho family of monomeric GTPases. Your answer would be a three-letter string composed of letters C, R, and H only, e.g. HRR. ( ) It activates formin. ( ) It inhibits myosin II activity. ( ) When constitutively active, it induces the formation of many prominent stress fibers

HCH

Heart conditions known as cardiomyopathies include two main subtypes, namely the hypertrophic and dilated cardiomyopathies, in which some portions of the cardiac muscle become stiff or dilated, respectively. Drugs such as the calcium channel blocker verapamil are used to treat the former, while the Na+-K+ ATPase inhibitor digoxin (which elevates intracellular calcium concentration in cardiac muscle cells) is sometimes used for the latter. Mutations in the subunits of the troponin complex are among the genetic causes of these diseases, and can be grouped into mutations that result in either decreased (D) or increased (I) calcium-ion sensitivity of actin-myosin contraction. Which of these mutations would you expect to be associated with hypertrophic cardiomyopathy? Write down D or I as your answer.

I

Indicate whether each of the following descriptions matches actin filaments (A), microtubules (M), or intermediate filaments (I). Your answer would be a three-letter string composed of letters A, M, and I only, e.g. MMA. ( ) They have the smallest persistence length. ( ) They lack polarity. ( ) They have the highest tensile strength.

III

Indicate if each of the following descriptions matches actin filaments (A), microtubules (M), or intermediate filaments (I). Your answer would be a four-letter string composed of letters A, M, and I only; e.g. AAMM. ( ) They form hollow structures with multiple lateral interactions. ( ) They form strong structures that are more resilient than the other two cytoskeletal filaments. ( ) Their subunits bind GTP and hydrolyze it. ( ) They form coiled-coil interactions between the subunits.

MIMI

Indicate true (T) and false (F) statements below regarding the microtubule cytoskeleton in neurons. Your answer would be a five-letter string composed of letters T and F only, e.g. TTFFF. ( ) All microtubules in an axon are normally oriented in the same direction. ( ) All microtubules in a dendrite are normally oriented in the same direction. ( ) Each microtubule in an axon has one end at the cell body and its other end at the axon terminal. ( ) Anterograde transport in axons is exclusively carried out by kinesins. ( ) Anterograde transport (toward the tip) in dendrites is exclusively carried out by kinesins.

TFFTF

Indicate true (T) and false (F) statements below regarding the primary cilia. Your answer would be a four-letter string composed of letters T and F only, e.g. TFFF. ( ) Primary cilia are found on the surface of almost all cell types in our bodies. ( ) Primary cilia are motile. ( ) Primary cilia are made in interphase at basal bodies. ( ) Primary cilia are thought to function in sensing and responding to external signals.

TFTT

Fill in the blank: Each microtubule is typically made of thirteen parallel .... that associate laterally to form a hollow tube.

protofilaments