Lecture 20: BICD 110 FINAL

Mitosis

What is Mitosis

The process of separating DNA equally between two daughter cells

Mitosis Overview: Major Steps

Prophase:

1. Chromosome condensation

2. Spindle assembly

Prometaphase/metaphase

3. Chromosome alignment

Anaphase

4. Chromosome separation

Telophase/Cytokinesis

2. Cellular separation

Aneuploidy

abnormal # of chromosomes (loss, gain, fragmentation, ploidy)

• chromosome misegregation = hallmark of cancer

Step 1: Chromosome Replication

• occurs in S phase when increase in cyclin A/CDK2 levels

• Cohesin joins sister chromatids at centromeres

Centromeric region

region where:

1. sister chromatids are paired

2. spindle MTs attach during mitosis

CENP-A (H3 variant) loaded at centromeric DNA

Sister chromatids

• identical copies of a chromosome (formed during DNA replication)

Step 2: Centrosome (spindle poles) duplication

Centrosomes (spindle poles) = MTOCS in proliferative cells

• composed of 2 centrioles

• duplicated (from 1 to 2) during S phase: each daughter inherits 1

• nucleates and anchors MTs for segregation

Prophase: Chromosome condensation

• Chromosomes condense BEFORE being aligned and partitioned

• Condensin: stabilizes DNA loops to condense chromo

  • Condensin II: activated by CDK1 phosphorylation

  • Condensin I: further condenses chromosomes in prometaphase

Prophase: Kinetochore Assembly

• large protein complex that binds MTs to centromere (marked by CENP-A)

• mediate MT attachment to capture the chromosomes

Prophase: Spindle Assembly

1. centriole pairs disengage from one another, promoting separation

2. during G2, each pair recruits PCM (making them STRONG MTOCs)

3. during M, matured centrosomes (now spindle poles) nucleate MTs

Prophase: Spindle Pole Separation

1. MT anchored at (-) ends at centrosomes

2. Kinesin 5 (+ end directed) binds to MTs and slides them apart

Prometaphase: NEBD

Nuclear envelope breakdown

• Lamins (IFs in nucleus) broken down by CDK1 to allow access to chromosomes

• Lamin A/C disperse freely in cytosol

• Lamin B are enclosed in vesicles

*w/o Lamin structural support, the nuclear membrane retracts into the ER, allowing full entry of dynamic MTs to capture kinetochores

Prometaphase: Search and Capture Model

1. Dynamic MTs “search” and “capture” kinetochore at their (+) ends

2. once captured, kinetochores congress towards spindle center via MT motors

Metaphase: Spindle Positioning

*ensure sister chromatids are attached via MTs at opposing ends

• Dynein-mediated pulling:

  • dynein interacts with astral MTs, exterting (-) end-directed pulling force

    to move the spindle

Astral MTs

• project from centrosome to cell cortex (cell membrane)

• spindle positioning and anaphase

Kinetochore MTs (K-fibers)

• connect centrosomes to chromosomes

• chromosome alignment (SAC)

Polar MTs

project to cell center w/ overlapping (+) ends

controls spindle size and anaphase

Metaphase: Bi-Orientation

• SAC: no progression to anaphase until all chromosomes are bi-oriented (amphitelic attachments)

  • 1 kinetochore attached to one MT, the other to the MT on an opposing pole

  • monotelic attachments are corrected by protein complex blocking APC/C activity

Anaphase

Goal: separate chromosomes, move to opposite poles

Anaphase: Cohesin Cleavage

SAC satisfied—> APC/C (ub protein ligase) activated

1. APC/C targets securin (which inhibits separase) for ub degradation

2. separase can now cleave cohesin, and chromosomes separate

Anaphase - Chromosome Separation

1. chromosomes move to opposite poles

2. spindle poles separate from each other

Kinesin - 13: depolymerizes MTs driving ‘poleward’ movement

Kinesin - 5: slides antiparralel polar MTs to move poles apart

Dynein (at cortex): pulls on astral MTs to move poles apart

Teophase and Cytokinesis: Contractile ring

GTPase RhoA: controls contractile ring and cleavage

1. RhoA-GTP (@ midzone) stimulates formin-mediated F-actin assembly

2. RhoA-GTP stimulates myosin II-mediated contraction, separating 2 daughter cells

Completing cell cycle steps

• APC/C degrades mitotic cyclins, resetting cell cycle back to G1