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bio 2b03 > module 10 > Flashcards

module 10 Flashcards

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1
Q

question: how is phosphorylation mediated?

A
  • MPF
  • bc MPF = heterodimer of mitotic cyclin and CDK
  • active MPF = promotes phosphorylation
  • inactive MPF = reverses phosphorylation
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2
Q

question: what events in prophase are mediated by MPF phosphorylation (what gets phosphorylated for each one)? (5)

A
  • active MPF
  1. formation of mitotic spindle
    - phosphorylate microtubule assoc. prot.
    ⤷ bc they promote microtubule instability
  2. condensation of chromo.
    - phosphorylate condensins and histone prot.
  3. preparation for sister chromatid separation
    - phosphorylate cohesins
  4. break down of nuclear envelope
    - phosphorylate nuclear lamins
  5. fragmentation of golgi and ER
    - phosphorylate GM130
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3
Q

question: what events in telophase are mediated by MPF dephosphorylation? (4)

A
  • inactive MPF
  1. nuclear envelope reassembly
  2. chromo. decondensation
  3. mitotic spindle disassembly
  4. golgi and ER mem. reassemble
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4
Q

explain: histone prot. role in chromo. condensation

A
  • 5 types of histones
  • H1 and H3 = phosphorylated by Aurora B kinase during chromo. condensation
  • H3 forms prot. core for nucleosome
  • H1 links nucleosomes
  • get packed tighter during condensation
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5
Q

explain: cohesin prot. role in chromo. organization

A
  • form cohesin complex
    ⤷ holds sister chromatids together until anaphase
  • releasing them = 2 steps
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6
Q

question: how does the release of cohesins happen?

A
  1. release from chromo. arms
    - keep cohesins in middle (centromere)
    ⤷ protected from phosphorylation by phosphotases
    - makes the x shape
    - release by phosphorylation (cyclin B-CDK, Aurora B)
  2. release from centromere
    - cleaved by separase
    - in anaphase
    - allows chromatid separation
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7
Q

explain: condensin role in chromo. condensation

A
  • condensins get phosphorylated by cyclin B-CDK
  • allow assembly and chromo. condensation
  • MPF phosphorylation sites are on XCAP-D2 domain of condensins
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8
Q

question: how do chromo. decondense?

A
  • dephosphorylate condensins and histones
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9
Q

question: what happens of the nuclear envelope throughout mitosis (nuclear envelope disassembly at interphase, prometaphase, metaphase)

A
  • interphase = intact mem.
  • prometaphase = chromo. condense and envelope fragments
  • metaphase = no envelope
    ⤷ frag. into small vesicles and dist. throughout sytosol
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10
Q

explain: struc. of nuclear envelope

A
  • 2 lipid bilayers
  • outer = cont.d w/ rough ER
  • inner = assoc. w/ IF called nuclear lamina
  • nuclear pore complexes everywhere to allow transport
  • lamina = lamin A, B, C
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11
Q

explain: phosphorylation of lamin prot.

A
  • phosphorylated at serine by cyclin B-CDK
  • initiates envelope disassembly
  • only lamin B stays assoc. w/ nuclear mem.
    ⤷ A and C are dispersed
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12
Q

question: what happens when lamin A doesn’t get phosphorylated (explain w/ hamster exp.)?

A

HAMSTER + HUMAN LAMIN A
- interphase = intact envelope
- prophase = lamina breaking down
⤷ can see lamin A diffusing into cytosol
- metaphase = lamin no longer organized + chromo. condensed

HAMSTER + VARIANT OF HUMAN LAMIN A
- lamin A couldn’t be phosphorylated
⤷ serine became alanine
- interphase = same (intact lamina)
- prophase = lamin A not diffusing into cytosol
- metaphase = chromo. condensed but still has ring of lamin A

**lamin B and C could still be phosphorylated
⤷ so lamina intact in metaphase means lamin A needs to be phosphorylated to break down lamina

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13
Q

question: how does the nuclear envelope reassemble?

A
  • inactivation of cyclin B-CDK + phosphatase activity
  • dephosphorylation of lamins A, B, C
  • lamins reassemble and reform lamina
  • B still assoc. w/ vesicles from before
    ⤷ so it brings vesicles to lamina to form inner nuclear envelope
  • nuclear pore complexes dephosphorylated
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14
Q

explain: golgi fragmentation

A
  • before separation, golgi = fragmented to each pole of spindle
  • GM130 (golgi prot.) = phosphorylated by cyclin B-CDK
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15
Q

question: what is the purpose of golgi and mitochondria fragmentation?

A
  • to ensure organelles are distributed to both daughter cells
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16
Q

question: what are the cyclins that control each part of the cell cycle for vertebrates?

A

EARLY G1
- D type cyclins
- CDK4 or 6

S PHASE (trigger)
- cyclin E-CDK2

S PHASE (completion)
- cyclin A-CDK2
- cyclin A-CDK1

MITOSIS
- cyclin B CDK1

G0
- none

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17
Q

question: what is the restriction point?

A
  • time late in G1 when passage through cell cycle = indep. of presence of the mitogen
  • cell continues into S phase even w/out mitogen
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18
Q

question: what are mitogens?

A
  • sig. molecules that induce cell div.
  • cause expression of G1 cyclin-CDK
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19
Q

question: what is the diff. between early and delayed resp. genes in re-entry to the cell cycle?

Study These Flashcards
A
  • adding mitogen starts early gene expression
  • peaks at 1 hour and declines
  • delayed starts when early declines
20
Q

question: what regulates early resp. gene expression?

Study These Flashcards
A
  • transcription factors activated by MAP kinase
    ⤷ SRF
    ⤷ TCF
  • already in cell so only need to be phosphorylated
    ⤷ also means its not affected by inhibitors
21
Q

question: what do the early resp. genes code for?

Study These Flashcards
A
  • c-Fos
  • c-Jun
    (transcription factors)

^these activate delayed resp. genes

22
Q

question: what do the delayed resp. genes code for?

Study These Flashcards
A
  • cyclin D
  • cyclin E
  • CDK2
  • CDK4
  • CDK6
23
Q

question: how does c-Fos activate delayed gene resp.?

Study These Flashcards
A
  • c-Fos = early resp. gene
  • induces exp. of CDKs needed for cell division
24
Q

question: what happens if mitogens and inhibitors of prot. synthesis are added into cell?

Study These Flashcards
A
  • translation inhibitors have no effect on early resp. gene exp bc already in cell (SGF, TCF)
  • but affects c-Fos and C-Jun -> affects delayed resp. gene
25
question: how do you turn off early resp. genes?
- expression and translation of transcriptional inhibitors encoded by early resp. genes - they turn themselves off
26
question: what happens when early genes turn off?
- early gene mRNA lvls stay high - no translation for early gene - delayed genes = never transcribed bc dep. on early genes
27
question: what happens when mitogen is added and removed before and after RP?
- adding growth factor/mitogen induces exit from G0 and entry into G1 - removing mitogen before RP passes -> decrease in cyclin D/CDK4-6 -> failure to proceed to S phase ⤷ retreat back to G0 - removing mitogen after RP passes -> decrease in cyclin D/CDK4-6 has no effect -> progresses to S phase ⤷ no effect bc cyclin E/CDJ2 is high enough
28
question: why can cell progress to S phase if mitogen removed after RP?
- E2F = regulates exp. of genes needed in S-phase - initially, E2f = inactive bc inhibitory prot. Rb present - cyclin D-CDK can phosphorylate Rb - if mitogen removed before RP, cyclin D lvls aren't high enough - but E2f also induces cyclin E/CDK which can also target Rb - if enough E2F is activated, induces exp. of E2f ⤷ +ive feedback loop - even if cyclin D decreases, cyclin E is self-sustaining and cell proceeds to S-phase
29
explain: functions of checkpoints
- recog. error and delay progression until its fixed - look for: ⤷ damaged DNA ⤷ unreplicated DNA ⤷ assembly of spindle ⤷ chromo. attachment during metaphase ⤷ completion of anaphase - if can't fix, cell does apoptosis
30
question: what are the types of damage found at damage checkpoints? (2)
1. ionizing radiation - creates double-stranded DNA breaks 2. ultraviolet radiation - creates thymine nucleotide dimers
31
question: what prot. recog. which types of damage? (2)
- ATR recog. UV damage (thymine dimers) - ATM recog. double strand breaks
32
question: how does ATR recog. + fix damage?
- targets Chk1 - Chk1 targets cdc25 **cdc25 regulates MPF between G2 to M-phase (reverses inhibitory phosphorylation) - Chk1 phosphorylates cdc25 - inactive cdc25 -> no MPF - arrests cell at G2 ⤷ gives time for cell to fix problem - when repaired, ATR dissociates ⤷ reverses everything ^^works for thymine dimers but also DNA replication checkpoints
33
question: how does ATM recog. + fix damage?
- targets Chk2 - Chk2 targets p53 prot. - p53 usually unstable but if phosphorylated, becomes stable - p53 activates p21 (CIP) ⤷ cyclin inhibitor - p21 binds to G1 cyclin-CDK to inhibit activity - cell stalls in G1 - when repaired, ATM dissociates ⤷ reverses everything
34
question: what happens if p53 can't fix the damage?
- constantly activated - induces pro-apoptotic genes to kill cell
35
question: what does spindle assembly checkpoint do?
- prevents entry into anaphase if chromo. aren't assoc. properly to spindle - delays cell in metaphase
36
question: how does cell fix error at spindle assembly checkpoint?
- wildtype cell reassembles the spindle when cell is arrested
37
question: what happens if SAC not functioning
- cell proceeds to anaphase even if chromo. aren't attached to spindle - chromatids don't get segregated to separate poles - nondisjunction and aneuploidy - one daughter cell has an extra chromo. - other cell missing a homologue
38
question: what regulates SAC?
- Mad2 - inactivates APC-Cdc20 ⤷ securins can stay ⤷ prevents separase from working - chromatids can't separate - loss of func. mut. in Mad2 -> cells proceed to anaphase early
39
question: what is Mad2 (what is it regulated by?)?
- can be open or close conformation - open Mad2 assoc. w/ kinetochores of chromo. that aren't on microtubules - Mad1 binding to Mad2 makes it close - closed Mad2 leaves chromo. and interacts w/ cdc20 ⤷ blocks cdc20 from associating with APC - keeps cell in metaphase
40
question: what happens when SAC error is fixed?
- Mad1 and Mad2 release - Mad2 back to open ⤷ releases the cdc20 - APC binds to cdc20 - APC-cdc20 reactivates ⤷ targets securin ⤷ separase can work - allows anaphase
41
question: what does mitotic exit network do?
- monitor completion of anaphase - activates cdc14 ⤷ to dephosphorylate cdh1 so it can bind to APC - activate APC-cdh1 ⤷ to inactivate MPF
42
question: where is cdc14?
- in nucleolus - can't reach it unless sister chromatids separate ⤷ means anaphase needs to happen
43
question: what happens in metaphase vs anaphase of budding yeast (tem1, cdc14)?
METAPHASE - Tem1 = GTPase - Tem1-GAP = GAP ⤷ keeps Tem1 in GDP bound form (inactive) - Tem1-GEF = on mem. (far) - when Tem1-GDP is inactive, cdc14 is hidden in nucleolus ANAPHASE - spindle gets longer - Tem1 gets closer to Tem1-GEF ⤷ exchanges GDP for GTP in Tem1 - allows passage through MEN checkpoint - allows release and activation od cdc14
44
question: what happens when cdc14 is released at anaphase?
- cdc14 dephosphorylates cdh1 - cdh1 activates APC - APC-cdh1 inhibits MPF - exit from mitosis can occur
45
question: what happens if chromo. segregation and anaphase don't happen?
- can't pass MEN - Tem1 stays inactive - cdc14 never gets released - MPF stays active - cell never exits mitosis
46
question: what does cdc14 do to inhibit S-phase?
- targets cdh1 (dephosphorylates) - inactivates MPF (degrades cyclin B) - keeps Sic1 unphosphorylated - Sic1 binds to S-phase cyclin-CDK - inhibits progression into S-phase **opposite can allow progression to S-phase if cdc14 is phosphorylated by G1 cyclin-CDK

bio 2b03 (8 decks)

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