Cell Growth Control

Peter Takizawapeter.takizawa@yale.edu

What we’ll talk about…

• Cyclin and cyclin-dependent kinases and control of the cell cycle

• Start and regulation of cell division

• Signaling pathways that stimulate and inhibit cell division

• Checkpoints and regulation of the cell cycle

• Mitosis

Cell division requires cell growth, chromosome duplication and separation.

CellCycle

Cell cycle is divided into separate phases.

G1

S

G2

MC

G0

Cyclins and cyclin dependent kinases (CDK) drive cell cycle events.

G1

G2

M

S

CDK

Cyclin

Different cyclin CDK complexes initiate and control the phases of the cell cycle.

G1 S G2 M G1

Start G2/MMetaphase/Anaphase

CDK Off CDK On CDK Off

Cdk2

Cyclin E Cyclin A Cyclin B APC

A C

Waves of cyclin expression and degradation mediate ordered progression of cell cycle.

Cdk2

Cyclin E Cyclin A Cyclin B APC

Cyclin E Cyclin A Cyclin B

Cyclin protein level

G1 S G2 M G1A C

Positive feedback loops increase the amount of active cyclin-CDK.

Cyclin

CDK

Cdc25inactive

Cdc25active

Activating

Inactivating

Wee1

CAK

Active

Switch-like activation of CDKs ensures rapid and irreversible initiation of cell cycle events.

+Wee1 Cdc25

time

prot

ein

conc

entr

atio

n

CD

K a

ctiv

ity

CDK protein

Cyclin protein

CDK activity

+

time

prot

ein

conc

entr

atio

n

CD

K a

ctiv

ity

CDK protein

Cyclin protein

CDK activity

Positive Feedback

CyclinCDK

CDK Cyclin

Ubiquitylation and proteosome are required to digest cyclins and decrease CDK activity.

UbiquitinProteosome

CDK

Cyclin

Start and the Decision to Divide

Start marks the initiation of DNA replication and an irreversible commitment to cell division.

SStart

Anti-mitogensMitogens

G2 M CG1

Cyclin D

Cdk4

Cyclin E

Cdk2

Cyclin A

Cdk2

Mitogens activate signaling pathways that lead to expression of Cyclin D.

Signal transduction pathway

Mitogen

Cyclin D

CDK4

CDK2

Cyclin E

DNA replication

Cyclin A

CDK2

Receptor tyrosine kinases activate MAP kinase pathways to increase expression of cyclin D.

Receptor tyrosine kinases

Guanine nucleotide exchange factor

Ras

MAP kinase kinase kinase

MAP kinase kinase

MAP kinase

Phosphorylation

MycTranscription

Cyclin D

EGF

Anti-mitogens activate signaling pathways that inhibit formation of cyclin D-Cdk complexes.

Smad

Phosphorylation

Transcription

TGF-β

Smad4

Ink4Cdk4

Cyclin D

Receptor tyrosine kinases

E2F proteins regulate expression of cyclin E.

Cyclin EEnhancer Repressor

Cyclin EEnhancer Repressor

X

E2F1 or E2F2 or E2F3

E2F4 or E2F5

Cyclin E

pRB inhibits cell division by promoting binding of E2F to repressor and inhibiting binding to

XX

E2F1

pRb

E2F4

pRb

Cyclin EEnhancer Repressor

Cyclin D/CDK inactivate pRB to allow E2F to stimulate transcription of cyclin E.

Cyclin EEnhancer Repressor

X

CDK4Cyclin D

Cyclin E

E2F1

pRbE2F1

E2F1

pRb

E2F4

E2F4

pRb

Positive feedback loop keeps cyclin E - CDK active.

TranscriptionPhosphorylation

Phosphorylation

CDK4Cyclin D

Cyclin ECDK2

Positive Feedback Loop

E2F1pRb

E2F1pRb

E2F1

pRb

Mitogens and Increase in Cell Size

TOR complex integrates the nutritional status of the cell and regulates cell growth.

mTORC Active

Growth Factors[ATP] [Amino Acids]O2

Protein Synthesis Lipid Synthesis

Mitogen-activated signaling pathways can turn on TOR complexes to promote cell growth.

Receptor Tyrosine Kinase

Adaptor

PI3 KinasePDK

AKT

TSC1/2 Inactive

Mitogen

mTORC Activerheb-GDP rheb-GTP

Checkpoints in the Cell Cycle

Checkpoints ensure completion of one stage of cell cycle before starting the next.

G1/S-CDK S-CDK M-CDK APC

DNADamage

Unreplicated DNA

DNADamage

UnattachedChromosomes

G1 S G2 M G1A C

DNA damage activates p53 to arrest the cell cycle.Damaged DNA

Cyclin ECDK2

Mdm2

p53Mdm2

p53

p53

p53

p21p53

Enhancer

p21

Mdm2

Mdm2ATM/ATR kinasesChk1/Chk2 kinases

DNA damage also triggers degradation of Cdc25 to slow the cell cycle.

Damaged DNA

ATM/ATR kinasesChk1/Chk2 kinases

Cdc25

Proteosome

Ubiquitin Ligase

p53 is activated by oncogenes and slows cell division.

Oncogenic Transformation: Ras, Myc, E2F

p53

p21

p14-ARF Cyclin D

Cyclin E

CDK4

CDK2

p14-ARFMdm2

Inactive

P53 triggers apoptosis by inducing release of cytochrome c from mitochondria.

p53 Bax

Cytochrome c

Apoptosis

Mitosis

Mitosis proceeds in several defined stages that involve changes in microtubule organization.

CF

CS

Interphase MetaphaseProphase Prometaphase

Anaphase AAnaphase BTelophaseCytokinesis

Three types of microtubules comprise the mitotic spindle.

Chromosomes Astral microtubules

Interpolar microtubules

Kinetichore Microtubules

CentrosomeKinesin

Mitotic Checkpoint

Incomplete or incorrect attachment of microtubules arrests cells in metaphase.

Metaphase ArrestHigh cyclin B-CDK2 activity

Transition to AnaphaseDecrease in cyclin B-CDK2 activity

Cohesins tether sister chromatids to prevent premature separation.

SisterChromatids

Cohesins

SisterChromatids

Histones

Unattached chromosomes prevent activation of anaphase promoting complex.

Anaphase-promoting complex,inactive

Cdc20

Anaphase-promoting complex,

active

Metaphase Arrest

Anaphase-promoting complex ubiquitylates cyclin B and securin to trigger transition to anaphase.

Cyclin B

Cdk2

Securin

Separase

Ubiquitylation

Ubiquitylation

Proteosome

Anaphase-promoting complex

Cdc20

Separase Active

Separase digests cohesins which allows tension from the spindle to separate chromosomes.

Separase

Take home points

• Cyclins and CDKs initiate different stages of the cell cycle

• Positive feedback is critical for commitment steps in the cell cycle

• Mitogens and anti-mitogens work through signaling pathways to influence the decision to divide

• Oncogenes trigger cell division in the absence of mitogen

• Tumor suppressors slow cell division by inactivating cyclin-CDKs

• Checkpoints monitor the state of the cell and can arrest the cell cycle