Chapter 11 Flashcards
What are the 5 functions of cytoskeleton of eukaryotic cells?
a. enables motion of organelles inside the cell
b. allows chromosomes to be properly partitioned to progeny cells
c. enables cells to move or move things along their surface
d. organizes the internal structure of the cell
e. determines the overall shape of the cell
What is a collection of proteins that form the roadways of the cell’s transportation system and motors that run on them?
cytoskeleton
What 3 major structural proteins is the cytoskeleton composed of?
a. microtubules
b. microfilaments
c. intermediate filaments
Microtubules are formed from what proteins?
tubulin proteins
Structure of microtubules
hollow tube
What 4 drugs disrupt microtubule polymerization?
a. taxol (paclitaxel)
b. colchicine
c. zoxamide
d. griseofulvin
Which drug prevents tubulin subunits from dissociating?
Taxol
Which drug blocks mitosis and are used to treat cancers?
Taxol
Which drug causes microtubules to dissociate and disappear?
colchicine
Which drug binds to fungal tubulins and prevents fungal growth?
zoxamide
What drug is used to control fungal blight in potatoes?
zoxamide
When does GTP never hydrolyze nor exchange with nucleotides in solution?
when the molecule of GTP bound to alpha-tubulin is next to the beta-tubulin
When is GTP hydrolyzed to GDP and can exchange with nucleotides in solution?
when the molecule of GTP is bound to beta-tubulin exposed at the end
About how many of what assemble parallel to one another to form a hollow microtubule tube?
11-15 protofilaments
How do tubulin heterodimers assemble into microtubules?
self-assemble
Microtubules are arranged in what fashion?
polar fashion
Why are microtubules arranged in a polar fashion?
all the tubulin heterodimers have the same orientation within each protofilament and all the protofilaments run in the same direction within a microtubule, so one end of the microtubule has only beta-tubulin exposed (plus end) while the other end has only alpha-tublin exposed (minus end)
What allows microtubules to act as directional tracks for molecular motor proteins, and is essential for organizing the interior of a cell?
polarity of microtubules
In fibroblasts, where are the minus ends and the plus ends?
minus ends in the interior of the cell
plus ends at the periphery
In epithelial cells where are plus and minus ends?
plus ends are basal
minus ends are apical
What happens if microtubules are depolymerized?
cells tend to lose their shape, forming round balls
Golgi complex fragments, disperse through the cell, the ER collapses around the nucleus
What does the instability of microtubules allow?
allows for dramtic rearrangement of microtubules to take place
In vitro, studies with purified tubulin and GTP in an appropriate buffer enable what?
observation of polymerization kinetics
In vitro studies, the amount of polymerization can be tracked by what?
light scattering by microtubules but not by tubulin heterodimers
How does the speed of polymerization progress change?
progress is slow at first, then speeds up after small subunits form because they are more likely to grow than depolymerize
In vivo, how do cells avoid the slow nucleation step?
by having specialized protein complexes to accelerate the nucleation step and direct where it will occur.
How do microtubules elongate?
by the addition of subunits onto both plus and minus ends
What is the model of microtubule polymerization where microtubule exist in persistent phases of either growth or shortening, with abrupt transitions between them?
Dynamic instability
What is the abrupt switch from growing to shortening?
Catastrophe
What is the abrupt switch from shortening to growth?
Rescue
What end of microtubules undergo catastrophe more frequently?
plus ends
What end of microtubules grow faster?
plus ends
What documents dynamic instability, found persistent elongation and shortening phases, abrupt transitions between them?
video microscopy
If an end is shortening, individual protofilaments peel away from what?
polymer lattice
Individual curling protofilaments are only held together by ______, resulting in _____.
longitudinal bonds; more rapidly disassembled
What is a non-equilibrium process?
Dynamic instability
Dynamic instability is possible because of what?
GTP hydrolysis by tubulin
During assembly, ______ stimulated to hydrolyze _____ to _____.
beta tublin; GTP to GDP
The hydrolysis of beta tubulin is slightly slower than polymerization, so growing microtubules have mostly what?
GDP-beta tubulins with a cap at the end of GTP-beta tubulins
Having a cap at the end of GTP-beta tubulins helps what?
regulate dynamic instability
________ dissociates from an end 50 X faster than _________
GDP tubulin; GTP tubulin
Exposure of GDP tubulins at the end results in what?
rapid depolymerization
________ result of a growing microtubule losing its GTP cap
Catastrophe
______ requires that GTP tubulins re-cap the end of a shortening microtubule
Rescue
What holds the protofilaments straight?
GTP tubulin cap
What prevents GDP-tubulins within core from relaxing to their preferred curved conformation?
GTP tubulin cap
MTOCs
microtubule organizing center
What is used by cells to nucleate microtubules?
MTOCs
MTOCs remain associated with ______ of nucleated microtubules, dictating _____ and _______.
minus ends; position; orientation
What is the most common animal cell MTOC?
centrosome
What are centrosomes composed of?
pair of centrioles at right angles to each other plus some pericentriolar material
What are centrioles constructed of?
triplet microtubules, 9 arranged in a circle to form the walls of a barrel-like structure
What are triplet microtubules made from?
alpha tubulin and beta tubluin plus delta and epsilon tubulin
What are pericentriolar matrix composed of?
about 100 different proteins, including gamma tubulin as part of the gamma tubulin ring complex
γTuRC
γ-tubulin ring complex
How is γ tubulin arranged?
as one turn of a very shallow helix with the shape of a lock washer
Arrangement of γ tubulin resembles what?
one turn of the helix made from the microtubule protofilaments as they form the hollow tube, in a spiral fashion
γTuRC nucleates microtubules from ________ so if the MTOC is near the nucleus, the ______ will be at the cell periphery
minus end; plus ends
What gets larger during interphase?
centrosomes
What dupulicates in S phase at right angle toward themselves?
centriole
Motile animal cells have additional MTOCs called ____.
basal bodies
Basal bodies serve as a template for what?
assembly of axoneme
What is axoneme?
bundle of microtubules that forms the core of cilia and flagella
Cilia and flagella formed by axoneme is essential for what?
movement
What uses structure called the spindle pole body, embedded in the nuclear envelope?
fungi
What 3 cells lack centrosomes so use other type of MTOC to nucleate and organize microtubules?
fungi, plant cells, and epithelial cells
What several microtubule nucleating sites distributed throughout the cell cortex?
plant cells
What have several microtubule nucleation sites near the apical end of the cell, thus microtubule plus ends grow out from the MTOC toward the basal cell end?
Epithelial cells
The dynamics of the growing and shrinking of microtubules can be visualized with what?
fluorescent tubulin
What 2 ways can the fluorescent tubulin be generated?
a. by either expressing tubulin fused to a fluorescent protein
b. by injecting cells with purified tubulin tagged with a fluorescent dye
What is used to obtain images of microtubule turnover?
FRAP
What does FRAP demonstrate about half time of interphase and mitotic microtubules?
interphase microtubules have a half time of 5-10 minutes
mitotic microtubules have a half time of 0.5-1 minutes
How does the dynamic instability of microtubules assembled in vitro differ from that in living cell? (3)
a. plus ends grow 5-10X faster in cells than in vitro
b. microtubules in cells switch between growth and shortening more frequently
c. pauses more common in living cells
Where is the dynamic instability switches are more frequent?
toward the edges of the cell near the plasma membrane
What microtubules contain more modified tubulins and capped at their plus ends?
stable microtubules
In what cells are stable microtubules more abundant?
nonmitotic, differentiated cells
What happened to free microtubules in cells?
broken off from anchored microtubules or have been released from the centrosome
What happens if free microtubules are not stabilized at their minus end?
rapidly disassemble
What undergoes treadmilling?
free microtubules
What is treadmilling?
dynamic instability of the microtubule plus end is biased toward net growth while the minus end shortens
Why do cells need dynamic molecules?
because they need to adapt to new situations or new stages of the cell division cycle
Example of microtubules searcing interior of a cell
During prophase, they need to find and connect their plus ends to kinetochores
What happens to microtubules that does not hit a kinetochore?
rapidly fall apart
What happens to microtubules that hit a kinetochore?
they are stabilized
Each kinetochore is connected with up to how many microtubules?
40
Another function of dynamic microtubules related to signal.
allow cells to detect a signal at their plasma membrane, become polarized, and begin to change shape
Some proteins and organelles have the ability to hold onto what, allowing them to be _____?
tip of a growing or shrinking moelcule; transported
What generated most movement in cells?
molecular motors
MAPs
microtubules associated proteins
3 functions of MAPs
a. bind to microtubules
b. modify dynamic instability by slowing down or speeding up tubulin addition or subtraction
c. linkers between the microtubule tip or sides and membrane vesicles or other structures
What are MAPs that bind to microtubules only at their plus ends?
+TIPs
How long are +TIPs bound to microtubules?
for a short time, continually falling off and being added
An example of +TIP
CLIP-170
2 functions of CLIP-170
a. stabilizes microtubules by promoting rescues
b. links endosomes to microtubules
Other than TIPs, other MAPs do what?
speed up microtubule turnover by making microtubules less stable
How do other MAPs hasten catastrophe and make rescues less likely? (3)
a. by disrupting the GTP cap to stimulate catastrophe
b. by cutting microtubules into pieces to make more ends that can shorten
c. binding free tubulin subunits to decrease the amount of tubulin available
What are 2 examples of destabilizing MAPs?
a. katanin
b. MCAK = mitotic centromere associated kinesin
2 function of katanin
a. cuts microtubules by binding to their walls
b. disrupts contact between tubulin subuntis
MCAK
mitotic centromere associated kinesin
A molecular motor that disrupts the GTP cap by binding at microtubule ends
MCAK
2 functions of MCAK
a. disrupts the GTP cap by binding at microtubule ends
b. destabilizes the tip structure
How do MCAK destabilize the tip structure?
By favoring formation of protofilaments that curve away from the microtubule wall
Which destabilizing MAPs require ATP?
Katanin
MAP activity is regulated by?
phsophrorylation/dephosphorylation
Regulation of MAP activity by phosphorylation/dephosphorylation changes what?
the affinity fo the MAP protein for microtubules
What proteins are molecular motors?
microtubule binding protein
What does molecular motors use to power continuous movement along the side of a microtuble?
use repeated cycles of ATP hydrolysis
What moves vesicles and other organelles throughout the cell?
molecular motors
Which motors disperse pigment throughout the cell?
plus end directed motors
Which motors collect pigments at the center of the cell?
minus end directed motors
What are the 2 families of molecular motors that move on microtubules?
a. kinesins
b. dyneins
Kinesins move towards?
plus ends of microtubules
Dyneins move towards?
minus ends of microtubules
For radial array of microtubules found in a typical fibroblast cell, where does dyneins transport cargo towards?
towards the center of the cell
For radial array of microtubules found in a typical fibroblast cell, where does kinesins transport cargo towards?
towards the plasma membrane
What contributes to the direction of movement and navigation by the motor?
polarity of the polymer
What lacks polarity, so they have no motors?
intermediate filaments
Motor proteins can _______ anchored microtubules and transport cargo; or motor proteins can ____ anchored, so______.
walk along; be; motor protein moves the microtubule
Shape of all molecular motors
a pair of identical large globular domains are at the end of a long rod-shaped domain
Other than the large globular domain, some molecular motors have what?
second pair of smaller globular domains at the other end
What 2 sites do large globular domains have?
polymer binding sites and ATP binding sites
What are these large globular domains referred to as other than dynein?
head or motor domains
How is the large globular domains of dynein different?
it has an extra stalk protruding from its head the stalk binds to microtubules
What does the tail domain binds to?
cargoes
Each motor molecule is made from what?
several polypeptides
What is a homodimer held together by coiled-coil interactions along the rod-shaped region?
bulk
Kinesin superfamily with what move vesicles toward the plus end of microtubules?
N-terminal motor domain
Kinesin superfamily with what move towards the minus ends of microtubules?
C-terminal motor domain
Kinesin superfamily with what are used to regulate microtubule dynamics by using ATP hydrolysis to weaken the microtbule cap?
motor domain near the middle
Kinesin superfamily have four-headed motor called
bipolar motor
How does bipolar motors help kinesin superfamily?
allow them to bing 2 microtubules at once and slide them past each other
When are bipolar motors important?
during mitosis, necessary to rearrange the cytoskeleton
What are cytoplasmic dyneins?
homodimer with 2 motor domains
What are axonemal dyneins?
heterodimers or heterotrimers with 2-3 motor domains per molecule
Where are axonemal dyneins found in?
cilia and flagella
Motor proteins to bind to what 2 and do what with them?
a. bind ATP, hydrolyze it, and undergo a very large conformational change
b. bind and let go of the surface
What are the 2 possible mechanisms of bidirectional movement?
a. head-over-head mechanism (walking)
b. inchworm mechanism (sliding)
What bidirectional movement mechanism is used by 2 head motors?
head-over-head mechanism
Steps of kinesin’s sequence of events as it walks along a microtubule (6)
- Head 2 unattached and head 1 tightly bound to microtubule
- ATP binding to head 1 causes head 2 to swing forward
- Head 2 now positioned over the next binding site and will bind to the microtubule
- head 2 weakly binds to the microtubule and release ADP
- ATP hydrolysis at head 1, resulting in strengthening of interactions of head 2 to microtubule and releases head 1 from microtubule
- returns kinesin to starting position, but head 2 leading head 1
Kinesin’s two heads must be ______ to walk long distances without falling off the microtubule
coordinated
Kinesin and dynein walk along microtubule in _____ steps
8-nm
the length of steps that kinesin and synein walk along the microtubules is equal to what?
length of a single tubulin heterodiemr
What kind of motors do not always keep one head bound to the microtubule, act in large arrays, and spend more time NOT bound?
cilia and flagella
Motor heads not bound to microtubules allow other motors to do what?
to generate force on the same microtubules
Explain tug of war between motors
motors on both sides active; vesicle moves in direction with greater number of motors
Explain coordination of motor activity
motors on one side active; vesicle move in direction of active motor
Binding cargo to the correct motor is mediated by
motor’s tail domain
For the large kinesin family, what are similar and what are different?
motor domains are similar and tail domains are very different
What binds indirectly to cargo?
tail domain
Tail domain binds indirectly to cargo via what?
adaptor proteins
What does adaptor AP-1 link? (2)
a. links cytoplasmic domain of the M6P-receptor with the tail of a kinesin
b. links clathrin to regions of the TGN where vesicles bud
Where is the M6P receptor located?
within a vesicle from trans-Golgi network
What is vesicle budding linked to?
loading of motor proteins
What is an adaptor that links cytoplasmic dynein to membranes?
dynactin complex
Strucutre of dynactin complex
7 polypeptides and a short filament composed of Arp1
Arp1 is similar to what?
actin
Using Arp1 what does dynactin link?
dynein to spectrin
Spectrin is attached to the cytoplasmic face of a vesicle via what?
ankyrin
What is an important property of highly shaped and specialized cell types?
definite, intentional asymmetry
What two things of microtubules are crucial for asymmetry within cells?
a. dynamic instability
b. movement of microtubule-dependent motor proteins
When are the dynamic instability and movement of microtubule-dependent motor proteins crucial for asymmetry within cells?
in conjunction with actin and intermediate filaments of the cytoskeleton
How does growth cones of two cells encounter each other and react to the contact signal?
by extending microtubules towards the point of contact
Steps of growth cones of two cells encountering each other and reacting to the contact signal (6)
- 2 growth cones move toward one another (actin at the edge and microtubule inside the cell)
- actin moves to the inside of the cell
- edges of 2 growth cones make contact in small region
- signaling pathway is initiated
- microtubules reorient towards the point of contact
- both cones focus microtubules to the point of contact
Other than the microtubules, what are centered on the same point of contact?
polymerization of actin
Why does the orientation of the spindle during the cell division must be carefully chosen?
for daughter cells to have the required orientation in a tissue
Steps of orientation of spindles (4)
- spindle form with random orientation
- astral microtubules search interior of the cell and are captured by dynein
- dynein pulls on the astral microtubules, rotating the spindle
- because of the arrangement of dynein, the spindle rotates until it is oriented across the cell
What is crucial to ensure the budding daughter cell receives an equal share of chromosomes?
spindle orientation in budding baker’s yeast (saccharomyces cerevisiae)
Steps of the budding process from mother cell (4)
- budding starts
- spindle pole bodies duplicate and separate
- spindle forms with random orientation
- spindle orients and moves into the opening between the mother cell and bud
Steps of spindle moving from mother cell to bud (6)
- Kip2 and Kar9 load onto microtubule at one of the spindle pole bodies
- Kip2 transports Kar9 to + end of microtubules
- Bim1 anchors Kar9 at the end of microtuble, which binds to myosin
- myosin walks along actin cable, so microtubule grows into the bud
- microtubule attached to dynein
- dynein pulls the spindle into bud
What does microtubules function as?
cytoskeletal directors
Microtubules functioning as cytoskeletal directors determine what?
where actin should assemble and where it should contact
Actin and microtubules interact through what two linkers?
MAP and motor proteins
What binds to both actin and microtubules in neurons?
MAP2c
MAP2c function
help form and send out long projections in neurons
Linking microtubules to actin filaments guide what?
growing microtubules to specific sites in the cell
How can actin filaments and microtubules work together without being physically connected?
By relaying signals to each other via signaling pathways, and be controlled and respond to events from outside or inside the cell via signaling pathways
Many of the signals between actin and microtubules involve what?
G proteins (GTP-binding proteins)
Polymerization state of microtubules affects the dynamics and organization of actin filaments via what?
G protein Rac1
Rac1 function
responsible for actin filament growth and lamellipodia formation
What are cilium and flagellum composed of?
long bundle of microtubules surrounded by an extension of the plasma membrane
Interactions among the microtubules inside causes what to the structures?
to bend and to beat back and forth, moving fluid past the surface of the cell
Cilia or flagella on the surface of unicellular eukaryotes or sperm cells enable cells to exhibit what?
motility
What 3 functions does the cilia covering the apical domains of some epithelial cells enable?
a. clearing mucus and debris from respiratory tract
b. transporting eggs from the ovary to the uterus
c. circulating the cerebrospinal fluid
What is generally shorter and more numerous?
cilia
How does each cilium or flagellum generate force?
by bending near its base with a power stroke and then a recovery stroke
What does recovery stroke do? (2)
a. propagates the bend from base to tip
b. readies the cilium or flagellum for the next power stroke
What continues to beat if removed from the cell, even if the plasma membrane is removed? What is the only requirement?
Flagella; having ATP
What is the core that is made of at least 250 different types of polypeptides?
axoneme
What arrangement does the cross-section illustrate?
9 + 2 arrangement with variety of proteins to bind and stabilize the microtubules
Where are the plus and minus ends in microtubules?
plus ends at the tip of the axoneme and minus ends at its base
What protein is responsible for connecting adjacent double microtubules around the circumference of the axoneme?
Nexin
What connects the double microtubules to the central pair of microtubules?
Radial spokes and spokeheads
What connect adjacent doublet microtubules generate force within the axoneme?
Axonemal dyneins
How is the beating motion of cilia and flagella generated?
by propagating a bend in the axoneme from the base toward the distal tip
What are active only within a small region of the axoneme and activated sequentially?
dyneins
Dyneins are regulated via
the central pair of microtubules and radial spokes
How can the rapid rotation of the central pair microtubules regulate/activate dynein activity?
spin my transmit signals to the radial spokes
Central pair microtubules rotate rapidly to regulate/activate dyenin activity via what?
kinases and phosphatases
Where is the basal body?
at the base of flagellum or cilium
Structure of basal body
9 triplet microtubuels
What is the structure of the basal body similar to?
centrioles
What serves as a template for assembly of the 9 outer double microtubules of the axoneme?
basal body
What remains connected as an anchor to the cell and what is cut off a cell?
basal body remain connected while flagellum is cut off
How fast does the new flagellum grow and when will it be functional?
grow in less than an hour and functional during the regeneration process
During the assembly process, necessary axonemal components are transported to the tip via what?
intraflagellar transport (IFT)
IFT
intraflagellar transport
What powers IFT of aconemal components to the tip?
kinesin
What powers IFT towards the cell body?
cytoplasmic dynein
What are found on nearly all vertebrate cells?
nonmotile cilia (primary cilia)
Structure of vertebrate cells with one primary cilium
9 + 0
A cell has a 9 + 0 structure when its axoneme lacks what?
central pair of microtubules
In some highly differentiated cell types, what is highly expanded and elaborated into a specialized domain?
distal tip of the primary cilium
How large can the expanded distal tip of the primary cilium be?
large as the cell body
What cells are an example of expanded tip of cilium into a large domain?
rods and cone cells
Large domain that is created by an expansion of the tip of the cilium
outer segment
What does the outer segments contain?
stacks of membrane disks filled with rhodopsin
What is a rhodopsin?
photoreceptor protein
What moves membrane vesicles containing rhodopsin from the cell body to the outer segment?
IFT-type transport
