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95 Cards in this Set
- Front
- Back
what would you see in FACS if cell couldn't divide? |
ALL cells would eventually enter G2 and all would be fluorescing w/ double the intensity (double the amount of DNA) |
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mutations of what regulatory proteins causes cell to stop dividing |
if Wee1 is hyperactive (inhib. phosphate on CDK) or if cdc25 not working (phosphatase doesn't remove inhib. phosphate) |
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what're cells called when they terminally differentiate? |
post- mitotic enter G0 and stay there neurons |
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FACS analysis: where are cells in g0 |
cells in G0 have same amount of DNA as cells in G1 if all cells were to stop going through cell cycle and enter g0: FACS would show all cells at same intensity as G1 phase |
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mitosis phases |
prophase metaphase anaphase telophase |
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aneuploidy |
chromosomes arent separated equally- one cell gets more chromosomes |
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prophase |
nuc. envelope breaks down spindle fibers and poles formed chromosomes condense |
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metaphase |
chromosomes line up in center |
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anaphase |
separation of sis. chromatids APC/C ACTIVATED --> degrades cyclin |
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why are yeast instead of mammal cells used to study cell cycle? |
mammal= can only see some details of only mitosis -can't see details of interphase: G1, S, G2 |
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S. cervisae (which yeast?) |
budding yeast |
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sci. name for budding yeast |
s. cervisae |
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s. pombe |
fission yeast (rod-like) |
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sci name for fission yeast |
s. pombe |
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properties of s. cervisiae cell cycle and growth |
as they undergo cell cycle: bud grows larger and larger *2 diff. sized cells produced LONG G1 PHASE most cells =found in G1 phase |
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cell cycle properties of s. pombe |
rod-like; symmetrical division septum is formed for cytokinesis LONG G2/M PHASES (most cells in these phases) |
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s. pombe vs. s. cervisae cell cycle diffs. |
pombe: rods, septum, G2/M cervisae: budding, asymmetric division, G1 |
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main regulatory factor of cell progression through cell cycle |
CDK: cyclin- dependent kinase --Kinase (phosphorylates) |
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3 protein families that control regulation of cell cycle |
kinases- enzyme phosphatases - enzyme cyclins |
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functional complementation |
procedure used to see what WT (normal) gene restored function of cell to grow (divide) in a mutant strand |
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what gene/ protein was first discovered w/ functional complementation?/ |
cdc28 CDK In s. cerevisiae-- ONLY ONE FOUND
*allowed for cell growth in temp.- sensitive s. cerevisiae |
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procedure for discovering genes responsible for cell growth |
1. mutant strand that didn't grow @normal temp. of 37--> nonpermissive temp. 2. using plasmid vectors, transform random genes of WT s. cerevisiae into mutant 3. try to grow in 37degrees IF CELLS GREW--> transcribed gene= regulates cell cycle |
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CDK in s. cerevisiae vs. s. pombe vs. humans |
s.cerev: cdc28 s. pombe: cdc2 HOMOLOGOUS TO EACHOTHER- same function -can use either (and even human CDK) to transform mutant strains and allow growth HIGHLY CONSERVED |
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what happens if you're missing CDK |
cells don't leave G2 phase and can't enter M phase --cells dont divide -in fission; you see cells getting longer and longer |
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which protein doesnt change in amount throughout entire cell cycle? |
CDK -amount =constant -activity= peaks @ late G2 |
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MPF |
mitosis promoting factor heterodimer CDK and mitotic cyclin CDK activity dependent on cyclin being bound and activation of CDK |
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mitotic cyclin levels throughout cell cycle |
gradually increases from G1--> G2 optimal/ maximal levels @ end of G2 after G2; amount of cyclin starts to decrease **gradual inc. of cyclin not associated w/ gradual increase of CDK (cdc2/cdc28) activity |
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classes of cyclins in cell |
G1 G1/S S M *ALL associate w/ one CDK and activate diff. components of the kinase |
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ubiquitin |
76 a.a.s expressed in all cells sequential addition--> polyubiquination= tag for degradation |
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degradation of mitotic cyclin |
ANAPHASE destruction box of mitotic cyclin is recognized by active APC/C -E3 + E2 (ligases) work together to add Ub ubiquinated protein's sent to 265 proteasome |
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proteasome responsible for degrading cyclins |
265 protease |
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ligases responsible for polyubiq. of cyclin |
E3--> ubiquitin ligase E2--> ubiquitin conjugating enzyme |
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wee1 function |
kinase- adds inhibitory phosphate to tyr15 (Y15) position - active from S--> end of G2 STOPS PREMATURE DIVISION STOPS CDK ACTIVITY |
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cdc25 |
phosphatase: removes inhib. phosphate added by wee1 active near end of G2 PROMOTES ACTIVATION OF CDK ALLOWS PROGRESSION THROUGH MITOSIS |
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deficit in Wee1 |
-CDK not inhibited w/ phosphate - premature cell division --> cell not bulked up enough yet: small cells |
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deficit in cdc25 |
inactivating phosphate not taken off CDK cells won't divide-- just get longer and longer |
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excess wee1 |
too much inhibition prevents cell division |
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excess cdc25 |
taking off inhibitory phosphate way too early premature cell division |
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why doesnt CDK activity gradually increase w/ gradual increase of cyclin |
CDK activity (and consequently MPF) is regulated by Wee1 and cdc25 stop CDK from becoming prematurely active UNTIL CYCLIN= AT MAXIMUM LEVELS |
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CAK |
kinase; adds activating phosphate to CDK @Thr161 (T161) position |
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position activating phosphate is added by and by what |
thr161 on CDK added by CAK |
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position inhib. phosphate is added by and by what |
Tyr15 (Y15) on CDK added by wee1 |
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steps for activation of MPF |
1. cyclin and CDK come together to form heterodimer (MPF)--> not active 2. Wee1 adds inhib. phosphate- late S and continually through G2 3. CAK adds excitatory phosphate 4. cdc25 removes inhib. P --late G2 |
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main role of MPF |
MPF = KINASE-- PHOSPHORYLATES **phosphorylation of diff. proteins controls processes needed for mitosis |
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processes controlled by activity of MPF |
chromosome condensation disassembly of nuclear membrane disassembly of interphase microtubules and formation of mitotic spindle turning off proteins for vesicular traffic |
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proteins regulated for chromosome condensation |
histones- ex. H1 phosphorylated by MPF to promote chromosome packing |
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G1 |
cell growth and metabolism --> prep for S LONGEST PHASE: ~11 HRS where most cells ARREST--> G0 **happens if not enough nutrients |
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S |
DNA synthesis and replication ~6-8 hrs |
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G2 |
prep. for chromosome segregation and M ~4hrs |
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M |
division of chromosomes and cell (MATC) ~1hr |
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when is DNA condensed |
beginning of M (prophase) **important for transport **done w/ MPF phosphorylation |
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when does dNA decondense |
right after M **relaxed chromosome important for replication |
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cdc28 |
CDK in s. cervsae (budding) |
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cdc2 |
CDK in s. pombe (fission) |
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APC/C |
anaphase promoting complex/ cyclosome |
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SPC/C |
synthesis promoting complex/ cyclosome --> regulates earlier phases of cell cycle and degradation of cyclins responsible for other phases |
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when is APC/C activated |
anaphase-- after cells have split |
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why is APC/C/ important |
TURNS OFF MPF QUICKLY!!! Lets cell quickly stop MPF activity in anaphase |
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when is wee1 activated |
end of S/early G2--> inhibiting P on MPF |
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when is CAK active |
as you progress through G2 adds inhib. p on CDK |
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when is cdc25 active |
end of G2 activates CDK |
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main cell cycle checkpoints |
DNA Damage checkpoint G1 checkpoint S phase checkpoint Metaphase checkpoint |
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S phase checkpoint |
make sure all DNA is replicated equally and completely before entering G2 |
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metaphase checkpoint |
make sure chromosomes lined up and connected to spindle properly before anaphase spindle assembly checkpoint |
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dna damage checkpoints |
at ALL phases!! stops mutations from being passed on |
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what type of mutation causes cancer |
mutation in gene that regulates cell cycle can cause cancer --> can lead to uncontrolled growth |
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restriction point in G1 |
check to see if there are enough nutrients for cell to bulk v. insufficient nutrients: cell--> G0 -- terminally differentiated |
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mutation/ deficit in ATM causes? |
ataxia telangiectasia |
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what is ATM/ ATR and main functions |
kinases-- phosphorylates proteins activated when theres DNA DAMAGE -> activate p53 -> repair -> inhibit cdc25 |
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what inhibits cdc25 |
ATM/ ATR when DNA damage detected phosphorylation of cdc25 inhibits it --> INHIB. P ON CDK (cdc2/28) NOT TAKEN OFF |
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what activates p53 |
ATM/ ATR |
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main roles of p53 |
transcription factors -activates apoptosis -activates p21 |
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what does p21 do |
--CIK transcribed: physically inhibits CDK stops cell division |
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ataxia telangiectasia |
NO ATM made higher probability of cancer Motor deficits -- locomotion and balance dilation of blood vessels and cerebellum problems |
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why does ATM/ATG deficit cause higher risk of cancer? |
-DNA damage not detected --> p53 (tumor suppressor) not activated - p53 not activated to stop cell division when damage |
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what happens when no ATR |
seckel syndrome |
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what causes seckel syndrome |
no ATR |
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seckel syndrome |
small brains- neurons= dying off during development primordial dwarfism |
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how is p53 activated |
ATM/ATR detect DNA damage--> PHOSPHORYLATE p53 protein stabilizes p53--> moved INTO NUCLEUS!!! |
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what happens when p53 is activated |
p53= activated when phosphorylated by ATM.ATR --TRANSCRIPTION FACTOR : --apoptosis genes --p21 gene |
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Regulation of p53 |
p53= high turn over rate: ALWAYS PRESENT in all cells stabilized when activated and MOVED INTO NUCLEUS TO work |
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role of p21 |
inhibits ALL CYCLIN-CDK COMPLEXES Stops cells from continuing any phase of cell cycle |
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p53 mutation significance |
50% of cancers have mutation in p53 tumor suppressor gene if you inherit mutation; v. high chance of cancer (esp Fs) |
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what regulates CDK activity |
wee1 cdc25 CIK cyclin |
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mitogens |
things that produce extracellular signals to induce cell growth unwanted |
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transformation |
unwanted extracellular signal inducing growth viral vectors bringing in ONCOGENES GENETIC alterations: retrovirus |
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growth promoting signals? |
mitogens transformations |
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growth inhibiting signals |
damage cell/cell contact terminal differentiation --> G0 senescence loss of mitogenic signal |
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oncogenes |
genes that can induce cell growth |
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CID and how does it work |
produced by p21 (activated by p53 phosphorylation) active when DNA Damage inhibit CDK activity --> blocks activation/ ATP access |
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6 major classes of cell-cycle regulatory proteins |
1. CDK 2. cyclins 3. CDK inhibitors 4. phosphatase 5. ub ligases 6. transcription factors |
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role of transcription factors for cell cycle regulation |
control DNA synthesis transcription/synthesis of proteins needed to progress through cell cycle -> cyclins cell cycle arrest |
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role of ub. ligases in cell cycle regulation |
cyclin degradation |
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role of phosphatases in cell cycle regulation |
activation of CDKs by removing inhib. P |
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how do CDK inhibitor proteins work |
physically bind to stop activity --> block activation or block substrate/ ATP access |