Mitosis and the Cell Cycle 10/21/05

Lecture Outline Two goals of the Cell Cycle: Make one cell into two Must accurately replicate the genetic material Mitosis (replicate and distribute the chromosomes) Major phases Mechanics of chromosome segregation Cytokinesis (how does one cell become two?) Replication of the cytoplasm and organelles Control of the Cell Cycle Cyclins and CDKs The importance of checkpoints for quality control

Phases of the Cell Cycle The cell cycle consists of The mitotic phase (M) Interphase G1 S G2 INTERPHASE G1 S (DNA synthesis) G2 Cytokinesis Mitosis MITOTIC (M) PHASE Figure 12.5

Mitosis and the Cell Cycle Genetic information is copied exactly into each daughter cell See it in action

Each duplicated chromosome Has two sister chromatids, which separate during cell division 0.5 µm One chromosome, one DNA molecule Duplication Centromere One chromosome, two DNA molecules (Two attached chromatids) Sister chromatids Sister chromosomes separate during mitosis Figure 12.4 Centromeres Sister chromatids

Kinetochore microtubule Overview of Mitosis G2 OF INTERPHASE PROPHASE PROMETAPHASE Centrosomes (with centriole pairs) Chromatin (duplicated) Early mitotic spindle Aster Centromere Fragments of nuclear envelope Kinetochore Nucleolus Nuclear envelope Plasma membrane Chromosome, consisting of two sister chromatids Kinetochore microtubule Figure 12.6 Nonkinetochore microtubules Prometaphase: Nuclear envelope breaks down. Chromosomes attach to spindle DNA replication during Interphase Prophase: Chromosomes begin to condense. Spindle starts to form

TELOPHASE AND CYTOKINESIS Overview of Mitosis Centrosome at one spindle pole Daughter chromosomes METAPHASE ANAPHASE TELOPHASE AND CYTOKINESIS Spindle Metaphase plate Nucleolus forming Cleavage furrow Nuclear envelope forming Figure 12.6 Telophase: Complete set of chromosomes at each pole Metaphase: Chromosomes align in center of cell Anaphase: Sister chromatids separate

“tug of war” Balanced attachment of spindle fibers to both chromatids aligns chromosomes in metaphase “tug of war”

Both chromatids must be captured by spindle fibers Both chromatids must be captured by spindle fibers. If any kinetochores remain unattached, chromosomes will not separate Kinetochore microtubules attach to centromeres and direct the poleward movement of chromosomes Centrosome Aster Sister chromatids Metaphase Plate Kinetochores Overlapping nonkinetochore microtubules Kinetochores microtubules Chromosomes Microtubules 0.5 µm 1 µm Figure 12.7 Nonkinetechore microtubules from opposite poles overlap and push against each other, elongating the cell

Spindle fibers shorten at the kinetochore Mark

Kinetochore Chromosome movement Kinetochore Tubulin subunits Microtubule Motor protein Chromosome Kinetochore Tubulin subunits

Cytokinesis Animal cells divide by constriction Plant cells build a partition (cell plate) Cleavage furrow Contractile ring of microfilaments Daughter cells 100 µm (a) Cleavage of an animal cell (SEM) Figure 12.9 A Daughter cells 1 µm Vesicles forming cell plate Wall of patent cell Cell plate New cell wall (b) Cell plate formation in a plant cell (SEM) Figure 12.9 B

How do the cytoplasmic organelles divide? Mitochondria (and chloroplasts) are present in multiple copies, and randomly segregate into the two daughter cells. Membrane bound organelles (e.g. ER) fragment along with the nuclear membrane and are reconstructed in the daughter cells

Phases of the Cell Cycle The cell cycle consists of The mitotic phase (M) Interphase G1 S G2 INTERPHASE G1 S (DNA synthesis) G2 Cytokinesis Mitosis MITOTIC (M) PHASE Figure 12.5

The clock has specific checkpoints: the cell cycle stops until a go-ahead signal is received See cell-cycle game at: http://nobelprize.org/medicine/educational/2001/cellcycle.html G1 checkpoint G1 G0 (a) If a cell receives a go-ahead signal at the G1 checkpoint, the cell continues      on in the cell cycle. (b) If a cell does not receive a go-ahead signal at the G1checkpoint, the cell exits the cell cycle and goes into G0, a nondividing state. Figure 12.15 A, B

Cell Cycle Control System S-PHASE ENTRY (G1/S) Mitosis Complete? Growth/ Protein Synthesis adequate? No DNA Damage? G1 checkpoint MITOSIS EXIT: All chromosomes attached to spindles? Control system S G1 G2 M MITOSIS ENTRY (G2/M) Replication Complete? Growth/ Protein Synthesis adequate? No DNA Damage? Figure 12.14

The Cell Cycle Clock: Cyclins and Cyclin-dependent kinases G1 cyclin (cyclin D) S-phase cyclins (cyclins E and A) M-phase cyclins (cyclins B and A) Cyclin-dependent kinases (Cdks) G1 Cdk (Cdk4) S-phase Cdk ((Cdk2) M-phase Cdk (Cdk1) Cyclin levels in the cell rise and fall with the stages of the cell cycle. Cdk levels remain stable, but each must bind the appropriate cyclin (whose levels fluctuate) in order to be activated.

Phosphorylation of CDK Targets Changes Their Activity Now performs a cell cycle function

The Human Cell Cycle ~ 1 hour ~ 4 hours ~ 10 hours ~ 9 hours

How does the cell cycle cycle? Focus first on entry and exit from mitosis

Cyclin-CDK controls the cell cycle Cyclin B synthesized in S phase; Combines with cdk1 to make active MPF Cyclin component degraded in anaphase MPF triggers: assembly of the mitotic spindle breakdown of the nuclear envelope condensation of the chromosomes

The Cell Cycle According to Cyclin Abundance Cyiclin-CDK activity can also be controlled by inhibitors

Isolate mutants that divide too early or too late How are CDKs Regulated? Isolate mutants that divide too early or too late

CDKs are Regulated by Phosphorylation is a kinase is a phosphatase CAK Activating Kinase) is a kinase is a phosphatase

Conformational Changes Associated with CDK Phosphorylation Free CDK CDK + Cyclin T161 phosphorylation The T-loop blocks substrate access Binding of cyclin moves the T-loop Phosporylation moves the T-loop more

Cyclin Dependent Kinase Inhibitors (CKIs) CDK CDK p21 Cyclin p16 Cyclin CDK4 CDK4 p16

Cell Cycle Regulators and Cancer

Anaphase promoting complex Triggers: Chromosome separation Breakdown of cyclin to re-start the cycle Breakdown of geminin (to again allow replication)