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Trinity Term 1 > Mitosis > Flashcards

Mitosis Flashcards

1
Q

Describe prophase

A
  • cdk triggers entry
  • nuclear envelope is intact
  • mitotic spindle is forming, radiating from the centrosome
  • sister chromatids, attached the kinetochore, are condensing
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2
Q

Describe pro-metaphase

A
  • nuclear membrane is breaking down into fragments
  • centrosomes are migrating
  • kinetochores and associated proteins assemble at and form on the constricted centromeres
  • checking mechanisms ensure attachments remain type IV
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3
Q

Describe kinetochore assembly in pro-metaphase

A
  • organised layer
  • capture bundles of condensed microtubules via ndc80 long tethered proteins
  • leave a physical separation between kinetochore and microtubules
  • plus end is left free for subunit exchange
  • correct attachment of the microtubules to the chromosomes allow for stability
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4
Q

Describe type III attachment

A
  • if both kinetochores are attached to one microtubule, the attachment is unstable
  • syntelic
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5
Q

Describe type II attachement

A
  • unstable structure
  • attachment of one kinetochore to two microtubules
  • merotelic
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6
Q

Describe type IV attachment

A
  • stable
  • amphitelic attachment structure
  • both centromeres are attached to different microtubules
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7
Q

Describe type I attachement

A

Unattached

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8
Q

Describe type V attachment

A

laterally attached

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9
Q

Describe the type IV checking mechanisms of the cell

A
  • rely upon tension forces that only occur to the correct degree on type IV attachment
  • help of the Aurora-B kinase protein that is attached to inner kinetochore
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10
Q

Describe the congression to metaphase

A
  • relies upon both plus- and minus- end motors stabilising the microtubules
  • driven by plus end motors
  • minus-end motors are essential in causing tension that pulls the microtubules towards the poles
  • chromokinesins are necessary on the chromosome arms to generate polar ejection forces
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11
Q

Describe metaphase

A
  • sister chromatids align along the metaphase spindle and ultimately metaphase plate
  • kinetochores under tension
  • plus end motors on the chromosome arms generate a polar ejection force
  • tubulin subunits (during their poleward flux towards the periphery through the spindle) are added to the plus end of the kinetochore microtubules
  • creates a dynamic system of microtubule flux underlain by continuous dynamic catastrophe and rescue
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12
Q

List some plus end motors

A
  • centromere protein E
  • kinesin-7
  • kinesin-10
  • kinesin-1
  • kinesin-5
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13
Q

List some minus ends motors

A
  • dyenins
  • kinesin 14 (previously termed non-claret disjunction)
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14
Q

List some chromokinesins

A
  • XKLP1
  • kinesin-4
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15
Q

Describe dynamic instability in metaphase

A
  • facilitated by motor attachment to the kinetochore microtubules
  • tubulin subunits are added at the kinetochore and lost at the centrosome constricted region
  • visualised using caged-fluorescein staining on the tubulin and blue-stained chromosomes under photoactivation
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16
Q

Describe the tension between kinetochores and kinesins

A
  • tension created by pulling kinetochores counteracted by a multiplicity of plus-end directed motor proteins such as pushing interpolar microtubules away from the kinetochore
  • creates a stable chromosomal state on the metaphase plate between the two poles
  • proven under laser ablation of a kinetochore from a spindle arm, resulting in arm movement away from the pole, and kinetochore-connected arms moving towards the pole
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17
Q

Interpolar microtubules

A

astral microtubules

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18
Q

Describe the metaphase-anaphase transition

A
  • must be achieved synchronously
  • chromosome separation triggered by destruction of cohesin between sister chromatids
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19
Q

Describe the destruction of cohesin between sister chromatids

A
  • by separase
  • allows detachment
  • mediated by securin
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20
Q

Describe securin

A
  • usually binds separase
  • destroyed by ubiquitination
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21
Q

Describe the ubiquitination of securin

A
  • mediated by the APC
  • degraded by the 26S proteasome
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22
Q

APC

A
  • multi-subunit anaphase promoting complex
  • protein targeting for degradation
  • synchronisation of mitotic exit
  • checkpoint in the mitotic cycle
  • activated by cdk, amongst other kinases
  • manages a binary system: sensitive to signals from chromosomes indicating incorrect attachments via incorrect tension levels
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23
Q

What happens if there are any unattached kinetochores at the metaphase-anaphase transition

A
  • APC ubiquitination of securin is prevented by Mad2p by remaining bound
  • mitotic checkpoint to prevent mature activation
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24
Q

Mad

A

mitosis arrest deficient

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25
Q

Describe the APC in meiosis

A
  • ubiquitylates cyclin B, separating it from Cdc2 for proteasomic degradation
  • irreversible step that contributes to mitotic exit
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26
Q

Cdc2

A

cyclin-dependent kinase 2

27
Q

Describe anaphase

A
  • microtubules shorten, pulling the kinetochore, and the centrosome outwards
  • separates the sister chromatids towards opposite poles of the cell
  • several redundancy mechanisms to ensure appropriate chromosomal separation during anaphase
28
Q

Give a redundant anaphase mechanism

A
  • minus-end pulling upon microtubules corresponding with tethered dyneins pulling the kinetochores towards the pole
  • even though the microtubules depolymerise at both ends, shortening the kinetochore microtubules, the motors remain attached
29
Q

Describe anaphase B

A

CENP-E is found at the mid-zone, having undergone dephosphorylation at the metaphase-anaphase boundary

30
Q

CENP-E

A
  • centrosome protein E
  • plus-end motor acting upon overlapping microtubules
31
Q

Describe CENP-E dephosphorylation

A

activates the binding to the microtubules, and pushes them apart.

32
Q

Describe telophase

A
  • nuclear envelopes reform around the chromosomes
  • contractile ring that will facilitate cytokinesis begins to form
  • can be observed via light-sheet imaging of EB1 and MT tracking
33
Q

Describe the centrosome

A
  • contains two centrioles angled at 90 degrees to one another
  • divide during prophase in order to duplicate
34
Q

Describe the centrosome cycle

A

an essential prerequisite for mitotic spindle formation.

35
Q

Describe the centrosome post-cytokinesis

A
  • one centrosome has been distributed to each daughter cell
  • in G1 there is one centrosome present
  • in late G1, the centrioles split, forming new pro-centrioles at the right angles to their parent centrioles
  • in prophase stage, the duplicated centrosomes separate, with the spindle microtubule array forming and separating between them
36
Q

Describe the spindle apparatus

A
  • composed of dynamically unstable microtubules.
37
Q

Describe microtubules

A
  • compounds of antiparallel alpha and beta tubulin dimers
  • bound either to GTP or GDP
  • forming 24nm hollow rings
38
Q

Describe the minus ends of microtubules

A

there is a section of gamma tubulin, capped by proteins and embedded into the centromere to ensure association

39
Q

Describe the plus ends of the microtubules

A

polymerisation and depolymerisation occurs, resulting in either a growing or shrinking molecule depending on their relative, regulated rates of catastrophe and rescue.

40
Q

What causes microtubule catastrophe?

A

tubulin dimers bound to GDP, which is more unstable

41
Q

What causes microtubule rescue?

A

tubulin dimers bound to GTP, which is more stable

42
Q

Describe the motor proteins

A
  • spindle organisation
  • driving chromosomal movement
43
Q

Describe the dyenins

A

positive end towards the negative end of the microtubules using ATP

44
Q

Describe the kinesins

A

negative end towards the positive end of the microtubules using ATP

45
Q

Describe spindle formation

A
  • dependent upon double-headed minus end motors localising to the centrosomes, linking microtubules and focussing them at one pole
  • can be observed via mutant, RNAi or immunolocalisation studies of Drosophila using GFP tagging at the cleavage stage
46
Q

Describe the role of + end motor proteins

A

may help to anchor the spindle from the centrosome by attaching to the negative ends of the attached microtubules.

47
Q

Describe pole separation

A
  • achieved by double-headed plus end motors such as kinesin-5 pushing the growing microtubule plus-ends against each other,
  • pulling of cytoplasmic dynein anchored in the cell periphery
48
Q

Describe the chromosome dynamics in prophase

A
  • condense
  • rounding of nucleotides to form filaments, and the looping of filaments to form solenoids
  • histone phosphorylation trigger by cdk, which activates the motor proteins, as well as SMC proteins
49
Q

Describe chromosome condensation

A

1400nm of DNA occupies the same amount of space previously taken up by 2nm.

50
Q

SMC proteins

A
  • stable maintenance of chromosomes
  • such as condensin, cohesin, smc1 and smc3
  • conserved across the domains of life
51
Q

Describe condensin

A

collects chromosomes together

52
Q

Describe cohesin

A

sticks chromosomes together

53
Q

Describe smc1 and smc3

A
  • form a hinged ring-like structure alongside scc3 and scc1 surrounding both sister chromatids
  • established in the S phase
54
Q

Describe hinged smc2 and smc4

A

stabilise the 300nm fibre alongside CAP-G, CAP-H and CAP-D2 proteins.

55
Q

Describe the chief function of cdk in the mitotic context

A
  • synchronising spindle formation, chromosome condensation, and localisation of motor protein localisation for mitotic initiation
  • phosphorylate the nuclear lamins supporting the large nuclear envelope, resulting in dissociation of the meshwork of intermediate filaments, destabilising the membrane and therefore causing NEB
  • allows vesicles to dissociate and disperse, and the microtubules to invade the nucleus
56
Q

Describe the perception of SMC action

A

can be perceived under phase contrast images, as condensed chromosomes exhibit a different refractive index.

57
Q

NEB

A

nuclear envelope breakdown

58
Q

Describe microtubule dynamic regulation in mitosis

A

by KRPs

59
Q

KRPs

A
  • kinesin related proteins
  • migrate towards the plus end, increasing catastrophe frequency
  • rate of replacement of microtubules more rapid
  • creating visualisable comet trails of microtubule caps
60
Q

Describe the observations of dynamic microtubule regulation in mitosis

A
  • observed in mutant studies
  • XKCM1 (kinesin-12) mutant
  • very long microtubules (even during mitosis) relative to a control spindle
  • in vivo using EB1-GFP
61
Q

Summarise role of motor proteins in spindle organisation in prophase

A
  • KIF11 is involved in antiparallel microtubule-microtubule sliding
  • dyenins are involved in the microtubule pulling from the NE, and at the cell cortex
62
Q

Summarise role of motor proteins in spindle organisation in prometaphase

A
  • KIF11 is involved in antiparallel microtubule-microtubule sliding
  • dyenins are involved in the microtubule pulling from the NE, and at the cell cortex
  • self-organisation using KIF11, KIFC1 and KIF15, as well as spindle pole focussing by dyenin, KIFC1 and the centrosome
  • more antiparallel microtubule-microtubule sliding is achieved by KIF11, KIF15 and KIFC1, as well as sliding kinetochores on lattice by dynein and KIF10
  • kinetochore pulling and pushing with dynein, KF10, KIF2B, KIF2C, KIF15 and K18A, and the polar ejection force using KIF4 and KIF22, and the poleward microtubule flux, involving KIF2A and KIF2C.
63
Q

polar ejection force

A

the pushing away of a chromosome approaching a pole

64
Q

poleward microtubule flux

A

also termed treadmilling

Trinity Term 1 (38 decks)

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