Ch.17 - Cytoskeleton Flashcards Preview

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Flashcards in Ch.17 - Cytoskeleton Deck (98)
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1

Summarize the general structure and function of intermediate filaments (ifils).

Ifils are polymers of proteins—wh vary according to cell type—that combine to form strong, cable-like fils that provide animal cells w mech strength.

  • 100+ diff types, e.g. keratin (epithelial cells), vimetins (fibroblasts), and neurofils (neurons)
  • Greatest tensile strength/flexibility (comp to mtubs/mfils) → bend instead of rupture.
  • Typ anchored to cyto-side of pmem via desmosomes interconn adj cells → form meshwork of struc support (e.g. nuclear lamina).
  • Durable - if cells are treated w salt solutions and nonionic detergents, most of cytoskeleton is destroyed, but ifils survive.

2

Ifil proteins are fibrous subunits, ea w an elongated central rod domain w distinct unstructured terminal domains at either end. Describe how these features influence their function.

Central rod domains of diff ifils are all similar in size/AA seq → enable close assoc/coiled-coil; by contrast, terminal domains (unstructured) of diff ifils vary greatly in size/AA seq → interact w specific components of cytoplasm.

3

Ifil proteins are fibrous subunits, ea w an elongated central rod domain w distinct unstructured terminal domains at either end. The central rod domain consists of an extended _______ region that enables pairs of ifil proteins to form stable ______ by wrapping around ea/o into _________ config. Two of these structures run in opp directions and assoc to form a staggered, antiparallel _______ → assoc side-by-side into helical array of __ (#) strands → final rope-like ifil

Ifil proteins are fibrous subunits, ea w an elongated central rod domain w distinct unstructured terminal domains at either end. The central rod domain consists of an extended α-helical region that enables pairs of ifil proteins to form stable dimers by wrapping around ea /o into coiled-coil config. Two of dimers run in opp directions and assoc to form a staggered, antiparallel tetramer → assoc side-by-side into helical array of 8 strands → final rope-like ifil

4

T/F: due to opposed dimer orientation of ifil proteins, tetramer ends are the same.

True

Tetramer ends are the same (structurally nonpolar) - due to opposed dimer orientation; distinguishing feature comp to mtubs/fils.

5

Ifil-ifil conns dep solely on noncovalent bonding. How can such weak bonds evoke such high tensile strength characteristic of ifils?

Ifil-ifil conns dep solely on noncovalent bonding → tensile strength fr combo of overlapping lateral interactions along length of ifil.

6

Central rod domains of diff ifils are all similar in size/AA seq, while terminal domains (unstructured) of diff ifils vary greatly in size/AA seq. How does this impact their role in the cell?

Central rod domains of diff ifils are all similar in size/AA seq → enable close assoc/coiled-coil; by contrast, terminal domains (unstructured) of diff ifils vary greatly in size/AA seq → interact w specific components of cytoplasm.

7

What are the four classes of ifils? Where are they located, and how are they assembled?

Four classes of ifils; all cytoplasmic except nuclear lamins (nucleus), and all assembled via polymerization of subunits:

  • Keratin fils in epithelial cells; most diverse class.
  • Vimentin and vimentin-related fils in CT, muscle, and supporting cells of NS (e.g. glial cells).
  • Neurofils in nerve cells.
  • Nuclear lamins of the nuclear envelope.

Note that ea class incl many subtypes, e.g. humans have 50+ keratin genes.

8

Many ifils are further stabilized and reinforced by accessory proteins. Describe one such protein.

Plectin - cross-link ifils into bundles → link to mtubs/fils and desmosomes.

  • Plectin mutations → epidermolysis bullosa simplex, muscular dystrophy, neurodegen.
  • Plectin may not be reqd for initial formation of ifils, but its cross-linking action is reqd to provide cells w necessary strength to handle mech stress.

9

Which of the following types of cells would you expect to contain a high density of ifils in their cytoplasm? Explain.

a. Amoeba proteus (a free-living amoeba)

b. skin epithelial cell

c. smooth muscle cell in digestive tract

d. Escherichia coli

e. nerve cell in spinal cord

f. sperm cell

G. Plant cell

Cells that migrate rapidly from one place to another, such as amoebae (A) and sperm cells (F), do not in general need intermediate filaments in their cytoplasm, since they do not develop or sustain large tensile forces.

Plant cells (G) are pushed and pulled by the forces of wind and water, but they resist these forces by means of their rigid cell walls rather than by their cytoskeleton.

Epithelial cells (B), smooth muscle cells (C), and the long axons of nerve cells (E) are all rich in cytoplasmic intermediate filaments, which prevent them from rupturing as they are stretched and compressed by the movements of their surrounding tissues.

All of the above eukaryotic cells possess at least intermediate filaments in their nuclear lamina. Bacteria, such as Escherichia coli (D), have none whatsoever

10

The nuclear envelope is supported by a meshwork of ifils called the ___________, wh is composed of _______.

The nuclear envelope is supported by a meshwork of ifils called the nuclear lamina, wh is composed of lamins.

  • Lamins - form 2-D meshwork instead of rope-like strucs like cytoplasmic ifils → comprise nuclear lamina, wh dis/assembles at ea cell division; also provides attachment sites for chromos.
  • Caution: don't confuse w laminin - an ECM protein.

11

Dis/assembly of nuclear lamina is controlled by de/phos of lamins. Explain.

Lamins are phos by protein kinases → conform change weakens binding b/w lamin tetramers → lamins dissoc.

Lamins dephos by protein phosphatases at end of mitosis → tetramer binding becomes energ favorable → lamins reassoc.

12

Microtubules (mtubs) are long, straight, hollow cylinders of tubulin protein; typ w one end attached to single mtoc/centrosome. Describe several other features of their structure and function.

  • More rigid than mfils/ifils; rupture when stretched.
  • Dynamic instability - rapidly dis/assemble.
  • Organizing euk cells - radiate out fr mtoc to cell periphery → created system of tracks along wh vesicle/organelles/other components can be xprtd.
  • Mitosis - mtubs dis/reassemble into mitotic spindle → helps segregate chromos equally.
  • Form stable core of cilia/flagella - rhythmically beating, hair-like structures; extend fr surface of many euks → swim/sweep fluid over their surface.
    • Bac flagella have entirely diff struc → swim by v diff mechanism.

13

Mtubs are hollow, cylindrical tubes composed of ___________ subunits, ea composed of ____ (2/3/4) v similar globular proteins bound tightly by ___________ (non/covalent) interactions.

Mtubs are hollow, cylindrical tubes composed of tubulin dimer subunits, ea composed of two v similar globular proteins (α- and β-tubulin) bound tightly by noncovalent interactions.

14

__________ are a linear, alternating chain of tubulin dimers, ___ (#) of wh form a mtub.

Protofilaments are a linear, alternating chain of tubulin dimers (heterodimer/αβ-dimer = .mtub subunit); 13 parallel protofils form mtub

15

T/F: Ea protofil—and mtub as a whole—has struc polarity

True

Ea protofil—and mtub as a whole—has struc polarity, w α-tubulin exposed at one end ("minus" end), β-tubulin at other ("plus" end).

  • Think: Beta = Bigger = plus end.
  • Protofils align w same orientation → cross-section of mtub would reveal ring of all α- OR all β-tubulin.
  • Caution: struc polarity and +/- ends have nothing to do w elec charge; rather, refers to chem/functionally distinct ends.
  • Struc polarity is critical for IC xprt.

16

In concentrated solution of pure tubulin (in vitro), protofils/mtubs polymerize at both ends, but more rapidly at which end?

Polymerization - in concentrated solution of pure tubulin (in vitro), protofils/mtubs polymerize at both ends, but more rapidly at β/plus end.

17

The ________ is the major mtub-organizing center in animal cells

The centrosome is the major mtub -organizing center in animal cells.

MTOC (centrosome in animal cells) - specialized organizing centers that control location, #, and orientation of mtubs; typ close to nucleus (except during mitosis).

18

A centrosome consists of pair of ________ surrounded by a matrix of proteins that incl hundreds of ring-shaped strucs of ________ → ea ring complex serves as a starting point ("____________") for growth of one mtub. _________ dimers add to ea ring complex in specific orientation → mtub's _____ (+/-) end is embedded in centrosome and ____ (+/-) end extends into cytoplasm.

A centrosome consists of pair of centrioles surrounded by a matrix of proteins that incl hundreds of ring-shaped strucs of γ-tubulin → ea γ-tubulin ring complex serves as starting point ("nucleation site") for growth of one mtub. αβ-tubulin dimers add to ea γ-tubulin ring complex in specific orientation → mtub's minus end embedded in centrosome and plus end extends into cytoplasm.

19

T/F: centrioles have no role in nucleation of mtubs fr centrosome.

True

Centrioles are strange - ea centriole—sitting perpendicular to its partner—is a cylindrical array of short mtubs, yet centrioles have no role in nucleation of mtubs fr centrosome (γ-tubulin ring complex alone is sufficient).

  • Besides serving as organizing centers (basal bodies) for mtubs in cilia/flagella, key function still a mystery.
  • Note: plant lack centrioles.

20

Why do mtubs need nucleating sites such as those provided by γ-tubulin rings in centrosome?

Much harder to start new mtub fr scratch (i.e. new ring of αβ-tubulin dimers) than to add dimers to pre-existing γ-tubulin ring.

Despite spont assembly in vitro (w high concen of free tubulin), free/cytoplasmic αβ-tubulin concen in vivo is too low.

  • By contrast, free actin monomers are at high enough concen in vivo to spont assemble.

21

Once a mtub has been nucleated, typ grows outward fr _________ for many minutes by polymerization of ________ to _____ end.

Once a mtub has been nucleated, typ grows outward fr MTOC/centrosome for many minutes by polymerization of αβ-dimers to β/plus end.

22

Dynamic instability of mtubs has been likened to a fisherman casting a line. Explain.

Dynamic instability occurs w/o warning; mtub can suddenly shrink rapidly inward (depoly) of β/plus end → suddenly start growing again, or may disappear completely, to be replaced by new mtub that grows fr same γ-tubulin ring complex/nucleation site.

  • Centrosome/mtoc continually shoots out new mtubs in diff directions in exploratory fashion → many retract, but can be stabilized by attachment to another molecule (capping protein) or cell struc.
  • Like a fisherman casting a line - if no bites (stabilized conns) → reels line (mtub) back in → casts in diff direction (fr same nucleation site).
  • Used to position organelles relative to one/an.

23

Describe how selective stabilization can polarize a cell.

Selective stabilization of mtubs can polarize a cell.

Newly formed mtub will persist only if both ends are protected fr depoly: α/minus end typ anchored in mtoc, but β/plus end initially free → must be stabilized by binding specific proteins, e.g. "capping" proteins in cell cortex.

24

Dynamic instability is driven by ____ hydrolysis

Dynamic instability is driven by GTP hydrolysis.

GTP is tightly bound to tubulin dimers: GTP hydrolysis ↓ affinity of dimers for their neighbors → promote mtub disassembly.

25

Ea free αβ-dimer contains one GTP tightly bound to β-tubulin → dimer added to growing mtub → quickly hydrolyzes GTP, and GDP remains tightly bound to β-tubulin.

How does this intrinsic capacity of tubulin dimers to hydrolyze GTP contrib to dynamic stability and formation of GTP or GDP 'caps'?

GTP cap - β/plus end composed entirely of GTP-dimers → favors growth; occurs when poly proceeds faster than GTP hydrolysis.

  • GTP-assoc dimers bind more strongly/eff to ea/o than GDP-dimers → mtub continues to poly/grow.

GDP cap - β/plus end composed of GDP-dimers → favors shrinkage; occurs when dimers 'randomly' hydrolyze their GTP before next dimers added, e.g. when growth is too slow.

  • Recall: rest of mtub is composed of GDP-tubulin → depoly/shrinkage tends to persist once started → may disappear completely → new mtub started fr same nucleation site.
  • GDP-tubulin freed during depoly join pool of free dimers in cytosol → exchange GDP for GTP → ready for poly.

26

Why do you suppose it is much easier to add tubulin to existing mtubs than to start a new mtub from scratch? Explain how γ-tubulin in centrosome helps to overcome this hurdle.

Two αβ-dimers have a limited # of possible interaction sites → lower affinity for ea/o → unlikely that sequential dimers will assoc for long enough to spont form new mtub.

By contrast, αβ-dimers have many possible interaction sites w end of a mtub: end-to-end or side-to-side w protofil.

The preassembled rings of γ-tubulin (centrosome) are held t/g in much tighter side-to-side interactions than αβ-dimers, so binding additional αβ-dimers is similar to adding to existing mtub; thus γ-tubulin rings can be thought of as permanently preassembled nucleation sites.

27

Colchicine and Taxol are drugs that assoc w free dimers and mtubs (resp) and result in chromosomes being unable to segregate during mitosis.

Briefly describe how such mechanisms might work.

Colchicine - tightly binds free dimers → prevents poly into mtubs; e.g. cell in mitosis stalls as mitotic spindles rapidly disappear, unable to sep chromos.

  • Indicates that mitotic spindle is normally maintained by continuous poly/depoly.

Taxol - opp effect as colchicine; binds tightly to mtubs → prevents depoly, but poly still permitted → same net effect: chromos can't sep → cell in mitosis stalls.

  • Cancer cells divide 'uncontrollably' → may be killed preferentially by mtub-de/stabilizing 'antimitotic' drugs, e.g. colchicine, Taxol, vincristine, and vinblastine

28

Cells can modify dynamic instability of mtubs for partic purposes. Describe how this applies to mitosis and differentiated cells.

Cells can modify dynamic instability of mtubs for partic purposes:

E.g. mitosis - mtubs become more dynamic → enables rapid dis/reassembly into mitotic spindle

E.g. differentiated cells - dynamic instability of mtubs often suppressed by proteins that bind/stabilize mtub → maintain specialized function.

29

T/F: Most differentiated animal cells are polarized

True

Most differentiated animal cells are polarized

  • E.g. nerve cells - axons extend fr one end of soma (cell body), dendrites fr other; all mtubs in axon have β/plus end pointed toward axon terminals (away fr soma) → directional xprt of organelles, vesicles, and macros.
  • E.g. secretory cells - Golgi positioned toward site of secretion.

30

Dynamic instability causes mtubs either to grow/shrink rapidly. Consider an individual, shrinking mtub:

a.  What must happen at the end of the mtub in order for it to stop shrinking and to start growing again?

b.  How would a change in the tubulin concen affect this switch?

c.  What would happen if only GDP, but no GTP, were present in the solution?

d.  What would happen if the solution contained an analog of GTP that cannot be hydrolyzed?

A. The mtub is shrinking bc it has lost its GTP cap (dimers on end in GDP-bound form). While shrinking, free GTP-loaded dimers fr solution will still add to this end; if added quickly enough to overcome depoly ('cover up' GDP-loaded dimers), a new GTP cap can form and regrowth is favored.

B. As free (cytosolic) GTP-dimer concen ↑ → rate of addition to mtub ↑. Creates self-balancing system: mtubs shrink, releasing GDP-dimers into cytosol → dimers swap GDP for GTP, thus ↑ free GTP-dimer concen → rate of addition to mtub ↑; and v-v.

C. If only GDP present, mtubs would continue to shrink and eventually disappear, bc dimers w GDP have v low affinity for ea/o and will not add stably to mtubs.

D. If GTP is present but cannot be hydrolyzed, mtubs will continue to grow until all free dimers have been used up.