Cell Cycle and Control Flashcards
DNA replication occurs in ___ phase
2 daughter cells are generated in ___ phase
S phase = replication
M phase = 2 daughter cells generated (mitosis + cytokinesis)
interphase = __ + ___ + ___
what is purpose of interphase
interphase = G1 + S + G2
time of delay to accumulate mass, monitor intra/extra-conditions, point of regulation
explain goals of restriction point of G1
“should I invest all this energy in replication?”
measures favorably of environment
if passes, cell is COMMITTED
(if environmental signals are unfavorable AFTER restriction point, oh well)
senescence
cells don’t pass the bar at G1 restriction point, hang out in G0 until conditions become more favorable
(after passing, cell is committed)
4 goals of cell cycle control
- turns on at specific times (clockwork)
- events are in correct order
- each event is only 1x time
- on/off switches that trigger events in complete and irreversible fashion
major enzymes of cell cycle control
cyclin-dependent kinases (Cdks) - activity of these increase or decrease during different phases
phosphorylation of target proteins initiatives or regulates cell cycle events
DEPENDENT ON CYCLINS (different cyclins for different phases)
4 classes of Cdks?
- G1 Cdk - promotes passage through restriction point (cyclin D)
- G1/S Cdk - commits cell to replication (cyclin E)
- S Cdk - initiates replication (cyclin A)
- M Cdk - promotes mitosis (cyclin B)
how does S-Cdk also prevent re-replication?
phosphorylation of Cdc6 by S-Cdk causes Cdc6 to dissociate from ORC (pre-RC disassembled)
dissociation/phosphorylation of Cdc6 causes its degradation
S-Cdk also phosphorylates Mcm (helicase) proteins to cause export from nucleus
describe steps of entry into S phase
- ORC binds origins and recruits regulatory proteins
- Cdc6 increases transiently in early G1, binds ORCs —> Mcm (helicases) recruited (ORC + Cdc6 + Mcm = pre-replicative complex)
- S-cyclin transcription activated in late G1, S cyclin-Cdk complex phosphorylates DNA pol (pre-RC activated)
- S-Cdk assembles DNA pol/replication machinery at origins, activates Mcm (helicases) —> replication is triggered
how does M-Cdk prevent replication occurring twice?
phosphorylates Cdc6 and Mcm (helicase) proteins
*keep in mind S-Cdk activity remains high during G2 and mitosis to keep Cdc6 phosphorylated/inactivated
[at end of mitosis, all Cdk activity is zeroed so Cdc6 and Mcm can be DEphosphorylated to allow for pre-Rc assembly again]
regulation of M-Cdk is controlled by 3 proteins:
- M cyclin transiently increases during G2 and M phases (transcribed)
- CAK (Cdk activating kinase)
- Wee1 (Cdk inhibitory kinase, inhibitory phosphorylation)
M-Cdk is “poised” for activation - what needs to be done to activate it?
Cdc25 (phosphatase) removes inhibitory phosphate (Wee1, Cdk inhibitory kinase)
***not all Cdc’s are phosphatase - they each have unique mechanisms and roles (don’t group them)
M-Cdk phosphorylates proteins responsible for: (3 things)
- assembly of spindle for chromosome segregation
- chromosome condensation
- breakdown of nuclear envelope
describe the feedback loops (2) to increase M-Cdk activity
- M-Cdk inhibits Wee1 (inhibitory kinase of M-Cdk)
- M-Cdk phosphorylates more Cdc25 (phosphatase which removes Wee1)
exit from mitosis requires inactivation of M-Cdk which occurs primarily through _____
ubiquination (degradation)
M-Cdk regulates its own degradation
during what phase of the cell cycle is Cdk activity ABSENT, and what mechanisms ensure this?
G1 - NO Cdk activity, via:
- ubiquination of Cdks
- Cyclin Kinase Inhibitor (CKI) accumulates
- decreased cyclin transcription via Rb (retinoblastoma) protein and E2F transcription factor (Rb INHIBITS E2F)
what protein and transcription factor are needed to decrease cyclin transcription during G1 (to ensure absence of Cdk activity)?
Rb (retinoblastoma) protein and E2F transcription factor
E2F function is controlled by Rb protein —> during G1, Rb binds/blocks E2F (G1/S and S cyclins NOT transcribed)
extracellular signal to divide —> G1-Cdk accumulates and phosphorylates/inactivates Rb (loses affinity for E2F) —> G1/S and S cyclins transcribed
how do CDK inhibitor proteins (CKIs) such as p27 work?
binds active cyclin-Cdk complex —> inactivated
why does loss of both copies of Rb gene cause a cancer, retinoblastoma?
Rb’s job is to bind/block E2F transcription factor for cyclin transcription
without it, cyclins are transcribed, which continue to activate Cdks, and cell cycle continues unregulated
describe the feedback loop for Rb protein (3 ways)
when E2F is released from Rb, it increases its own expression
E2F expression leads to production of G1/S-Cdk and S-Cdk —> G1-Cdk phosphorylates Rb (inactivates, MORE E2F released)
increased G1/S-Cdk and S-Cdk enhance phosphorylation of ubiquitin ligases and CKIs, leading to their destruction in proteosome — G1/S-Cdk and S-Cdk activated
what is the purpose of G2 checkpoint
prevents mitosis when there is DNA damage so it’s not passed on
damaged DNA sends signal that blocks Cdc25 activity (phosphates that removes Wee1 inhibitory kinase) —> M-Cdk remains phosphorylated and inactive
what is the purpose of the G1 checkpoint and how does it work?
G1 checkpoint prevents progression into S (replication) phase
inhibits activation of G1/S-Cdk and S-Cdk complexes - controlled by p53
p53: stimulates expression of several genes including CKI and p21
CKI then binds to G1/S-Cdk and S-Cdk to inactive them
how does p53 work to check cell cycle in G1 checkpoint?
during G1 DNA damage checkpoint, stimulates expression of several genes including CKI and p21
CKI binds to G1/S-Cdk and S-Cdk to inactive them
entry into S phase prevented
Fill in:
At the beginning of __ phase, ____ activates a ubiquitin-ligase that then results in the degradation of _____
At the beginning of M phase, M-Cdk activates a ubiquitin-ligase that then results in the degradation of M-Cdk
regulates itself
Which of these feedback mechanisms is NOT regulated by M-Cdk?
a. phosphorylation and further activation of Cdc25
b. activation of APC ubiquitin-ligase
c. Inhibition of Wee1
d. Activation of Wee1
M-Cdk DOES regulate these:
a. phosphorylation and further activation of Cdc25
b. activation of APC ubiquitin-ligase
c. Inhibition of Wee1
does NOT activate Wee1
Which of these is not an immediate effect of Rb deletion?
a. increased levels of E2F
b. increased levels of S-Cdk
c. re-entry into G1
d. decreased expression of Cdc25
these are immediate effects of Rb deletion:
a. increased E2F
b. increased S-Cdk
c. re-entry into G1
Rb deletion would not inherently cause decreased expression of Cdc25
