The Mammalian Cell The Mammalian Cell CycleCycle

Rajat SinghaniaRajat Singhania

03/21/200603/21/2006

““Let the Truth be Told!!”Let the Truth be Told!!”

Overview of the Cell CycleOverview of the Cell Cycle

Source: www.els.net

Let’s Start with G1!!Let’s Start with G1!!

Fact: Most of the cells in our body are Fact: Most of the cells in our body are NOT replicating at any given point of time.NOT replicating at any given point of time.

Instead, most cells in our body remain in Instead, most cells in our body remain in the G0/G1 phase for their entire lifetime.the G0/G1 phase for their entire lifetime.

Most of the cells that go through the cell Most of the cells that go through the cell cycle on a regular basis are the adult stem cycle on a regular basis are the adult stem cells in the bone marrow, skin, and the gut cells in the bone marrow, skin, and the gut - continual cell division occurs to replenish - continual cell division occurs to replenish the constantly dying cells in those organs. the constantly dying cells in those organs.

CdK VideoCdK Video

G1 restriction pointG1 restriction point

pRb: Tumor Suppressor Protein retinoblastoma

E2F-DP: Transcribes genes whose products are used in DNA synthesis as well as for the genes for Cyclin E, Cyclin A, and Cdk2.

Various growth factors and hormones activate Cdk4/6 & Cdk2 which phosphorylate pRb…

Source: www.eurogene.org

DNA Damage CheckpointDNA Damage Checkpoint

Source: www.eurogene.org[p21 is a CDKI which binds Cdk2 and Cdk4/6]

Triggering the S phaseTriggering the S phase

The newly formed CycA-Cdk2 complexes The newly formed CycA-Cdk2 complexes help “trigger” pre-replication complexes help “trigger” pre-replication complexes assembled on the DNA by causing the assembled on the DNA by causing the assembly of DNA polymerase, thus assembly of DNA polymerase, thus initiating the S-phase.initiating the S-phase.

Source: www.med.upenn.org

More CycA-Cdk2 S-phase More CycA-Cdk2 S-phase RegulationRegulation

CycA-Cdk2 phosphorylates E2F to CycA-Cdk2 phosphorylates E2F to inactivate its function as a transcription inactivate its function as a transcription factor.factor.

CycA-Cdk2 also keeps the pRb protein CycA-Cdk2 also keeps the pRb protein inactive during the S phase.inactive during the S phase.

CycA-Cdk2 also phosphorylates Cdc6 CycA-Cdk2 also phosphorylates Cdc6 (part of the pre-replicative complex) and (part of the pre-replicative complex) and thus marks it for degradation – thus thus marks it for degradation – thus ensuring that re-replication does not occur.ensuring that re-replication does not occur.

G2/M transition (Cdc2=Cdk1)G2/M transition (Cdc2=Cdk1)

During the G2 phase of the cell cycle, inactive During the G2 phase of the cell cycle, inactive cyclin B1/Cdc2 complexes accumulate in cyclin B1/Cdc2 complexes accumulate in mammalian cells due to inhibitory mammalian cells due to inhibitory phosphorylation of Cdc2 on Thr-14 and Tyr-15 phosphorylation of Cdc2 on Thr-14 and Tyr-15 by the Wee1 and Myt1 kinases.by the Wee1 and Myt1 kinases.

CyclinB1/Cdc2 activation requires CyclinB1/Cdc2 activation requires dephosphorylation on Thr-14 and Tyr-15 by the dephosphorylation on Thr-14 and Tyr-15 by the phosphatase Cdc25C, and phosphorylation on phosphatase Cdc25C, and phosphorylation on Thr-161 by CAK3,4.Thr-161 by CAK3,4.

Activation of Cdc25CActivation of Cdc25C

The mammalian Plx1 homolog Plk1 The mammalian Plx1 homolog Plk1 phosphorylates Cdc25C.phosphorylates Cdc25C.

Active cyclin B1/Cdc2 activates Cdc25C by Active cyclin B1/Cdc2 activates Cdc25C by phosphorylating the N-terminal region of phosphorylating the N-terminal region of Cdc25C in an autocatalytic loop. Thus, it Cdc25C in an autocatalytic loop. Thus, it takes only a small amount of active cyclin takes only a small amount of active cyclin B1/Cdc2 for its concentration to rapidly B1/Cdc2 for its concentration to rapidly explode.explode.

G2 Checkpoint ActivationG2 Checkpoint Activation

Source: www.eurekah.com

In response to genotoxic stress such as Ionizing Radiation or Ultra Violet light, the ATM/ATR signaling pathway is activated. These two kinases are part of the phosphoinositide-3 kinase (PI-3K) family. ATM phosphorylates and activates Chk2; ATR, Chk1. Activated Chk1 and Chk2 phosphorylate Cdc25C on Ser-216, generating a consensus binding site for 14-3-3 proteins. Cdc25C is then exported out of the nucleus and is sequestered in the cytoplasm by the 14-3-3 proteins. This leads to G2 cell cycle arrest due to the inhibition of the ability of Cdc25C to activate cyclin B1/Cdc2. Cdc25C phosphorylation happens at a different site than when it is done by Plk1, as mentioned in the previous slide.

Mitosis Stage 1: ProphaseMitosis Stage 1: Prophase

The replicated chromosomes condense to become very thick and dense. Microtubules start sprouting from the centrosomes which had also replicated during the S-phase of the cell cycle. The centrosomes start to move apart.

Mitosis Stage 2: PrometaphaseMitosis Stage 2: Prometaphase

Source: www.sparknotes.com

The nuclear envelope abruptly breaks down.

Microtubules start attaching to the kinetochores of the chromosomes.

Centrosomes go towards opposite poles; chromosomes start moving towards center.

Mitosis Stage 3: MetaphaseMitosis Stage 3: Metaphase

The chromosomes align at the metaphase plate.

The metaphase checkpoint monitors the attachment of the mitotic spindle to kinetochores and the tension generated by mitotic spindle attachment. In the presence of even a single unattached kinetochore, the metaphase checkpoint halts the separation of sister chromatids and thereby provides additional time for spindle attachment.

Source: http://www.stanford.edu/group/fanglab/science/research_cycle.html

Mitosis Stage 4: AnaphaseMitosis Stage 4: Anaphase

The breakdown of the cohesin protein complex that holds the sister chromatids together is done by the Anaphase Promoting Complex (APC).

The APC works by degrading a protein that is binding a proteolytic enzyme that cleaves the cohesin when it is no longer held inactive by the now-degraded protein.

Mitosis Stage 5: TelophaseMitosis Stage 5: Telophase

Mitosis Stage 6: CytokinesisMitosis Stage 6: Cytokinesis

Cleavage occurs by the contraction of a thin ring of actin filaments that form the contractile ring.

If the ring is not positioned at the center of the cell, an asymmetrical division takes place.

Cell Cycle VideoCell Cycle Video

Importance of Cell SizeImportance of Cell Size

As the cell grows in size, more and more As the cell grows in size, more and more ribosomes are produced. So, more and ribosomes are produced. So, more and more cyclins are produced, and so more more cyclins are produced, and so more and more Cyclin-dependent Kinase and more Cyclin-dependent Kinase complexes form (e.g. CyclinA-Cdk2 going complexes form (e.g. CyclinA-Cdk2 going into S; CyclinB-Cdk1 going into M)…since into S; CyclinB-Cdk1 going into M)…since the nucleus size doesn’t increase, these the nucleus size doesn’t increase, these become more and more concentrated -> become more and more concentrated -> more active->carry out the transition into more active->carry out the transition into the next cell cycle phase.the next cell cycle phase.