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exam 2 Flashcards

cell bio

1
Q

AB MT dimer structure

A
  • alpha tubulin permanently bound to GTP
  • Beta can bind GTP/GDP. hydrolyzes GTP during formation
  • alpha = - end
  • beta= + end
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2
Q

MTOC’s

A
  1. Spindle poles: duplicated centrosomes. found in mitosis
  2. Nerve cells:
    - axons: discontinuous. same polarity.
    - dendrites: not associated with MTOC. mixed orientation.
  3. centrosomes: main ones in animals
  4. basal body: cilia/flagella
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3
Q

Centrioles

A
  • a perpendicular set found in the pericentriolar matrix
  • 9 sets of triplet microtubules
  • yTurc initiates poly of MT’s. Nucleating site. negative end associated with yTurc
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4
Q

Augmin

A

provides nucelating site in cetrioles. Supports polymerizzation in yTurc. Centrioles

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5
Q

Dynamics of MT assembly- GTP/GDP tubulin

A 
GTP= strongly favoured to grow at + end 
GDP= rapid disassembly. weakens cohesion b/w protofilaments
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6
Q

Colchicine

A
  • induces depoly of cytoplasmic MT’s = singlets

- centrosomes remain b/c triplets = stable

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7
Q

Taxol

A

Stabilizes. inhibits mitosis

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8
Q

MAP’s

A

side binding proteins. + domain binds - MT side
MAP2: cross-bridges. Link MT’s to intermediate filaments. Dendrites
Tau: spacer. Axons/dendrites
+TIPs: associate with the growing + end. stabilize. Can attract hitchhikers

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9
Q

Regulation of MAP’s

A

via phosphorylation

cant bind to MT’s when phosphorylated

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10
Q

Kinesin 13 and Stathmin

A

K13: binds to dimers at the positive end. Promotes depoly. Requires ATP
Stathmin: binds sets of dimers. Promotes disassembly by promoting GTP hydrolysis. inactivated by Phosphorylation

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11
Q

Singlet, Doublet, Triplet examples

A

Singlet: Cytoplasm. Most common
Doublet: cilia/flagella
Triplet: basal body, centrosome (centrioles)

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12
Q

Cytoplasmic vs Axonemal MT’s

A

Cyto is more dynamic

Axonemal are more stable

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13
Q

MT formation diagram

A
  • Cc of tubulin dimers needed for MT assembly.
  • Cc reached, tubulin dimers are in equilibrium
  • above Cc, most of the mass is from MT’s
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14
Q

Kinesin

A

Motor proteins for MT’s

  • heavy chains: head, linker, stalk. Head binds ATP and MT’s
  • light chains: bind cargo
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15
Q

Examples of Kinesins

A

K1: conventional. 2 LC/HC. Anterograde movement
K2: 2 diff HC and 1 LC. Organelle transport. Anterograde
K5: bipolar. 4 HC=4 heads. Sliding of MT to + end
K13: short heavy chain. NOT a motor protein disassembly via ATP

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16
Q

Kinesin Movement

A
  1. head binds ATP, releases MT.
  2. Conf change in linker. Trailing head swings forward 16nm. Linker docks in head
  3. Leading head releases ADP , lagging head hydrolyses ATP. Linker undocked from head
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17
Q

Cytoplasmic Dynein

A
  • for organelle transport
  • Negative end retrograde transport
    Stem: links dynein to cargo through dynactin
    Head: ATPase
    Stalk: extension from head, has MT binding site on end
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18
Q

Dynactin Complex

A
  • links dynein to cargo and regulates movement
  • dynamititin regulates dynactin/cargo association. overexpression= separation of dynein and dynactin
  • p150 glued keeps system attached to MT
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19
Q

Post trasnlational modification of tubulin

A

acetylated lysine on alpha tubulin = stability and promoes Kinesin 1 movement

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20
Q

Axoneme

A
  • cilia/flagella structure
  • 9+2 array
  • doublets connected via Nexin
  • A tubule as axonemal dynein permanently bound
    stem attached to A, stalk attached t
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21
Q

Axoneme zones

A 
Continuous
Basal body: 9 sets of triplets (below PM). ABC
Transition Zone: 9 doublets. AB
Axoneme: 9+2. AB
- only doublets found above PM
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22
Q

Axoneme bending

A

= physical movement. Activtion of dynein:
With nexin linkers= bending
Wout nexin linkers: sliding. slide towards Negative end

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23
Q

Intraflagelular Transport

A
  • doesnt refer to movement
  • transport of organelles
  • cytoplasmic dynein and kinesin 1 utilized
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24
Q

Primary Cilium

A
  • interphase cell
  • sensory organelle
  • important in cell signalling
  • non-motile
  • NO axonemal dynein
  • 9+0 arrangement of MT
  • stabilized by acetylated alpha tubulin
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25
Dynamic Instability in Mitosis
- K13 is constitutively expressed. - XMAP215 inhibits the affect of K13. Enhances assembly at + end. activity fluctuates, decreases before mitosis and increases after, interphase. Can be inhibited by mitosis
26
Mitotic Apparatus
1. Astral: extend from poles towards to cortex, linked 2. Polar: towards cell centre. + end overlap= zone of interdigitation 3. Kinetochore: bind to KT's of MT
27
Spindle Formation
- all chromos must be aligned at meta plate - all must be captured - can attach at sides or end of MT - Bi-orientation: attachment to both sides of MT - chromos move towards furtherst pole
28
bi-orientation
- attachment to both sides of MT - ensures tension. Tension is required for movement - phosphorylation of Ndc80 by Aurora B= no tention - dephos by PP1= tension. Strong interaction b/w KT and MT - moves toward +end
29
Kinesin's in Spindle formation
Kinesin 13: depoly at both the -/+ end. Kinesin 4: on chromsome, + end movement. Results in ends pointing towards furthest pole Kinesin 7: keeps growing MT attached to KT
30
Anaphase
A: separtion of sister chromatids - shrinkage at both MT (+) and spindle pole (-) via kinesin 13 B: separation of spindle poles - B1: Sliding of MT's by Kinesin 5. push apart - B2: pulling to cell cortex by dynein/dynactin. astral MT's
31
Actin structure
Alpa: muscle Beta: cortex, networks Gamma: stress fibers (-) has ATP binding cleft
32
Actin assembly
1. nucleation 2. elongation. at both ends, preferentially the + end 3. steady phase. equilibrium b/w monomers and AF's. constant exchange
33
AF Cc
+: 0.12M -:0.60M the difference in Cc's results in treadmilling ATP-Actin needed for polymerization
34
Regulation of AF polymerization
Cellular G-actin in 400uM. Prevents constant poly Profilin: ADPactin--> ATPactin Cofilin: disassembly. binds to ADPactin in AF. Thymosin: Sequesters ATPactin, builds reservoir CapZ: binds to + end, prevents poly Tropomodulin: binds to- end, prevents depoly
35
Actin Disrupting Drugs
CytochalasinD: binds to AF at + end. Depoly Latrunculin: sequesters G-Actin. Depoly Phallodium: stabilizes AF's
36
Formins
- unbranched assembly - nucleating centre - activated by Rho-GTP, memrane bound protein - FH2 dimer, + end of Actin associated with dimer, - end extends
37
NPF
Wasp and Wave
38
Arp2/3
- nucleating centre - branched assembly - Activated by NPF's WASP acitvated by Cdc42 WAVE activated by Rac - can aid in endocytosis
39
Red Blood cell attachment to PM
Spectrin: links AF's together Ankyrin: binds spectrin units together and connect AF;s to PM via Glycophorin C
40
Microvilli attachment to PM
Via Ezrin
41
Muscle attachment to PM
Glycoprotein spans membrane into ECM | Dystrophin binds AF's and connects them to Glycoprotein
42
Myosin
- AF's motor protein - M2 is most abundant - Head: ATPase, actin and nucleotide binding site - Neck: essential and regulatory LC. - Tail: heavy chain. Binds cargo!!!!
43
myosin cleavage
Chymotrypsin: cleaves in middle of heavy chain. HMM (LC+Head and some heavy chain) and LMM (tail) Papain: cleaves at base of LC (head+LC) and HC
44
Classes of Myosin
1: monomer. membrane association and endocytosis 2: dimer. muscle conraction. bind AF's with tail domain 5. dimer. organelle transport
45
Myosin movement
1. head binds atp, releases from AF 2. ATP hydrolysis, elastic energy. cocked state 3. binds actin, releases Pi and elestic energy = powerstroke 4. head remains tightly bound until atp binds
46
Sarcomere components
- Zdisk: + end of AF associated with Z disk - myosin: attached to - end - I band: changes in size with contraction - A band: remains constant - CapZ/tropomodulin - Nebulin: binds actin subunits. determines length in sarcomere - Titin: binds myosin with the Z stack. prevents it from moving in the sarcomere
47
Sarco reticulum
- stores and regulates Ca2 levels | - is associated with transverse tubules
48
Steps in muscle contraction
1. Depolimerize. impule to transverse tubules 2. Ca channel opens, Ca release 3. Ca recoery via ATPase
49
Troponin/Tropomyosin
- calcium binding proteins - are bound to AF's, inhibit the binding of myosin - when calcium binds, changes conformation, allows myosin to find its bidning sites myosin only interacts with AF's in presence of Ca
50
Actin has roles in:
- phagocytosis | - cytokinesis: contractile ring
51
skeletal muscle
- regulated by Ca | - striated
52
Smooth muscle
- organs - regulated by phosphorylation - myosin LC phosphorylated = active myosin - much slower and persistant contraction
53
Cell migration
Stress fibers: made up of contriactile bundles (sarcomeres) - are associated with cell adhesion - associated with PM via Integrins (membrane proteins) Leading Edge: Pm pushed forward by Actin poly - treadmilling
54
integrins
- membrane proteins - connect stress fibers to plasma membrane - associated with the ECM - involved in cell migration
55
Migration requirements
- cells need receptors on surface | - Cdc42, Rac, Rho
56
migrating cells examples
- tissue and immune cells
57
Cell migration steps
1. extension: lamellipodium extends from the leading edge 2. adhesion: lamellipodium adheres to the substrave by focal adherens 3. translocation: bulk of cytoplasm in cell body flows forward due to contraction at the back of the cell 4. De-adhesion/Endocytic recycling: tail detaches, retracts. Endo cycle internalizes membrane and integrins at rear, transports them to the front
58
Cdc42-GTP
- filopodium - required for polarity - activates WASP (NPF), then Arp2/3 branching - activates Microtubule capture of lamellipodium - dynein activation - can activate Rac
59
Rac-GTP
- lamellipodium - activates WAVE--> Arp2/3 branching - can activate Rho
60
Rho-GTP
- stress fibers and contraction - activates formin--> unbranched - activates rho kinase--> active myosin
61
Rho proteins/GTPases
Cdc42, Rac, Rho
62
Dominant-Active GTPases
binds effector molecule consitituively | in absence of GTP
63
Dominant-Negative GTPases
inhibits activation of Rho proteins
64
Actin-binding proteins
- Fimbrin: microvilli, filopodia, focal adhesions - a-actinin: stress fibers, filopodia, muscle Z line - Spectrin: cell cortex - Filamin: leading edge, stress fibers, filopodia - dystrophin: linking membrane proteins to actin cortex in muscles
65
Intermediate Filaments
- not globular - don't bind nucleotide (don't need ATP/GTP - no polarity - no motor proteins - less dynamic - more stable - tetramer - add to pre-existing IF
66
Cytoplasm IF proteins
Keratins, Desmins, Vimentin, GFAP, Neurofilaments
67
Keratin
Class 1 and 2 - epithelial cells - associate with desmosomes and hemidesmosomes - tissue strength and integrity
68
Desmins
Class 3. - muscle cells. - muscle integrity - smooth muscle: link dense bodies - skeletal muscle: surround sarcomere. Encircle Z disk and are cross-linked with PM
69
Vimentins
Class 3 - mesenchymal cells - fibroblasts, migratory cells
70
Neurofilaments
Class 4 | - neurons, support the axon
71
Lamins
Class 5 - maintain the integrity of the nucleus - nuclear lamina links to the cytoplasm cytoskeleton via IF-associated proteins (found in LINC)
72
Lamins in mitosis
- need to be degraded to deplete nuc envelope - CDK's phosph lamins, promote IF disassembly and prevents reassembly - phosphatases remove phos, reassemble lamins. reform nuc env
73
IF-associated proteins
- organizational, no capping/motors/severing - can link nuclear lamina to the cytoplasm cytoskeleton via LINC - EX: plectin cross-links IF vimentin and MT
74
desmosomes
link epithelial cells
75
hemidesmosomes
link epithelial cells to ECM/connective tissue
76
IF function
- provide structural support for cell shape - can bind PM and Nuc mem - PM: vimentin links to ankyrin - NM: via lamins

Cell bio (7 decks)

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