Cell Cycle And Regulation

ALL cells would eventually enter G2 and all would be fluorescing w/ double the intensity (double the amount of DNA)

if Wee1 is hyperactive (inhib. phosphate on CDK)

or

if cdc25 not working (phosphatase doesn't remove inhib. phosphate)

post- mitotic

enter G0 and stay there

neurons

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

prophase

metaphase

anaphase

telophase

chromosomes arent separated equally- one cell gets more chromosomes

nuc. envelope breaks down

spindle fibers and poles formed

chromosomes condense

chromosomes line up in center

separation of sis. chromatids

APC/C ACTIVATED

--> degrades cyclin

mammal= can only see some details of only mitosis

-can't see details of interphase: G1, S, G2

budding yeast

s. cervisae

fission yeast (rod-like)

s. pombe

as they undergo cell cycle: bud grows larger and larger

*2 diff. sized cells produced

LONG G1 PHASE

most cells =found in G1 phase

rod-like; symmetrical division

septum is formed for cytokinesis

LONG G2/M PHASES (most cells in these phases)

pombe: rods, septum, G2/M

cervisae: budding, asymmetric division, G1

CDK: cyclin- dependent kinase

--Kinase (phosphorylates)

kinases- enzyme

phosphatases - enzyme

cyclins

procedure used to see what WT (normal) gene restored function of cell to grow (divide) in a mutant strand

cdc28

CDK In s. cerevisiae-- ONLY ONE FOUND

*allowed for cell growth in temp.- sensitive s. cerevisiae

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

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

cells don't leave G2 phase and can't enter M phase

--cells dont divide

-in fission; you see cells getting longer and longer

CDK

-amount =constant

-activity= peaks @ late G2

mitosis promoting factor

heterodimer

CDK and mitotic cyclin

CDK activity dependent on cyclin being bound and activation of CDK

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

G1

G1/S

S

M

*ALL associate w/ one CDK and activate diff. components of the kinase

76 a.a.s

expressed in all cells

sequential addition--> polyubiquination= tag for degradation

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

265 protease

E3--> ubiquitin ligase

E2--> ubiquitin conjugating enzyme

kinase- adds inhibitory phosphate to tyr15 (Y15) position

- active from S--> end of G2

STOPS PREMATURE DIVISION

STOPS CDK ACTIVITY

phosphatase: removes inhib. phosphate added by wee1

active near end of G2

PROMOTES ACTIVATION OF CDK

ALLOWS PROGRESSION THROUGH MITOSIS

-CDK not inhibited w/ phosphate

- premature cell division

--> cell not bulked up enough yet: small cells

inactivating phosphate not taken off CDK

cells won't divide-- just get longer and longer

too much inhibition

prevents cell division

taking off inhibitory phosphate way too early

premature cell division

CDK activity (and consequently MPF) is regulated by Wee1 and cdc25

stop CDK from becoming prematurely active UNTIL CYCLIN= AT MAXIMUM LEVELS

kinase; adds activating phosphate to CDK @Thr161 (T161) position

thr161 on CDK

added by CAK

Tyr15 (Y15) on CDK

added by wee1

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

MPF = KINASE-- PHOSPHORYLATES

**phosphorylation of diff. proteins controls processes needed for mitosis

chromosome condensation

disassembly of nuclear membrane

disassembly of interphase microtubules and formation of mitotic spindle

turning off proteins for vesicular traffic

histones- ex. H1

phosphorylated by MPF to promote chromosome packing

cell growth and metabolism

--> prep for S

LONGEST PHASE: ~11 HRS

where most cells ARREST--> G0

**happens if not enough nutrients

DNA synthesis and replication

~6-8 hrs

prep. for chromosome segregation and M

~4hrs

division of chromosomes and cell (MATC)

~1hr

beginning of M (prophase)

**important for transport

**done w/ MPF phosphorylation

right after M

**relaxed chromosome important for replication

CDK in s. cervsae (budding)

CDK in s. pombe (fission)

anaphase promoting complex/ cyclosome

synthesis promoting complex/ cyclosome

--> regulates earlier phases of cell cycle and degradation of cyclins responsible for other phases

anaphase-- after cells have split

TURNS OFF MPF QUICKLY!!!

Lets cell quickly stop MPF activity in anaphase

end of S/early G2--> inhibiting P on MPF

as you progress through G2

adds inhib. p on CDK

end of G2

activates CDK

DNA Damage checkpoint

G1 checkpoint

S phase checkpoint

Metaphase checkpoint

make sure all DNA is replicated equally and completely

before entering G2

make sure chromosomes lined up and connected to spindle properly before anaphase

spindle assembly checkpoint

at ALL phases!!

stops mutations from being passed on

mutation in gene that regulates cell cycle can cause cancer

--> can lead to uncontrolled growth

check to see if there are enough nutrients for cell to bulk

v. insufficient nutrients: cell--> G0

-- terminally differentiated

ataxia telangiectasia

kinases-- phosphorylates

proteins activated when theres DNA DAMAGE

-> activate p53

-> repair

-> inhibit cdc25

ATM/ ATR when DNA damage detected

phosphorylation of cdc25 inhibits it

--> INHIB. P ON CDK (cdc2/28) NOT TAKEN OFF

ATM/ ATR

transcription factors

-activates apoptosis

-activates p21

--CIK transcribed: physically inhibits CDK

stops cell division

NO ATM made

higher probability of cancer

Motor deficits -- locomotion and balance

dilation of blood vessels and cerebellum problems

-DNA damage not detected

--> p53 (tumor suppressor) not activated

- p53 not activated to stop cell division when damage

seckel syndrome

no ATR

small brains- neurons= dying off during development

primordial dwarfism

ATM/ATR detect DNA damage--> PHOSPHORYLATE p53 protein

stabilizes p53--> moved INTO NUCLEUS!!!

p53= activated when phosphorylated by ATM.ATR

--TRANSCRIPTION FACTOR :

--apoptosis genes

--p21 gene

p53= high turn over rate: ALWAYS PRESENT in all cells

stabilized when activated and MOVED INTO NUCLEUS TO work

inhibits ALL CYCLIN-CDK COMPLEXES

Stops cells from continuing any phase of cell cycle

50% of cancers have mutation in p53

tumor suppressor gene

if you inherit mutation; v. high chance of cancer (esp Fs)

wee1

cdc25

CIK

cyclin

things that produce extracellular signals to induce cell growth

unwanted

unwanted extracellular signal inducing growth

viral vectors bringing in ONCOGENES

GENETIC alterations: retrovirus

mitogens

transformations

damage

cell/cell contact

terminal differentiation --> G0

senescence

loss of mitogenic signal

genes that can induce cell growth

produced by p21 (activated by p53 phosphorylation)

active when DNA Damage

inhibit CDK activity

--> blocks activation/ ATP access

1. CDK

2. cyclins

3. CDK inhibitors

4. phosphatase

5. ub ligases

6. transcription factors

control DNA synthesis

transcription/synthesis of proteins needed to progress through cell cycle -> cyclins

cell cycle arrest

cyclin degradation

activation of CDKs by removing inhib. P

physically bind to stop activity

--> block activation or block substrate/ ATP access