Lecture 14: Cell Division

Dynamic Nature of Cell Division

• Cell division is a highly dynamic process

• The dynamic movement of chromosomes during cell division is mediated by a microtubule-based spindle

Cell Cycle - Overview

• The cycle is composed of “interphase” and “mitosis”

• Interphase

  • The cell isn’t actively dividing; Rather, it is constantly preparing for division

  • DNA duplicates, centrosomes duplicate, and the cell increases in size

• Mitosis (M Phase): Divided into six stages

  • Prophase

  • Prometaphase

  • Metaphase

  • Anaphase

  • Telophase

  • Cytokinesis

Prophase

• Chromosomes condense (can be seen as individual threads)

• Centrosomes move apart

Prometaphase

• Nuclear envelope breaks down

• Spindle becomes fully formed with two poles

• Dynamic microtubules capture chromosomes

• Chromosomes move to the center of the spindle

  • Chromosome “Congression”

Metaphase

• Chromosomes are attached to the microtubules from opposite spindle poles

• Chromosomes align in the center of the cell

  • At the “metaphase plate”

Anaphase

• Duplicated chromosomes move to the opposite spindle poles

  • One set moves to each pole

Telophase

• Contracticle ring begins to form at the center of the cell

• Nuclear envelope begins to reform

Cytokinesis

• Contractile ring constricts, creating a cleavage furrow

• Cell pinches in two

• Chromosomes decondense

Essential Steps of Cell Division

• Chromosome duplication and condensation

• Spindle assembly

• Chromosome capture and alignment

• Chromosome segregation

• Cytokinesis

Essential Step 1: Chromosome Duplication and Condensation

• Mitotic chromosome organization

  • DNA duplicates prior to mitosis to form sister chromatids

    • In S “Synthesis” phase

    • Chromatid: One copy of the chromosome

    • Sister chromatids are linked together b/c of a protein complex

      • Cohesin Complex

        • Loads onto the duplicated DNA strands during S phase and forms a ring that links the two strands, holding sister chromatids together until anaphase

  • Chromosome is highly condensed

    • Contain centromeres, which represent “primary constriction”

    • Condensation occurs to fit a lot of DNA into small nucleus

• Chromosomes separate in mitosis

  • To do so, chromosomes must condense even further in mitosis so they can be easily segregated (makes them more distinguishable compared to interphase)

    • Condensins

      • A complex of proteins that form rings that function to condense the DNA in mitosis

      • Thought to organize the DNA into loops

  • The links holding sister chromatids are gone, causing them to come apart

Essential Step 2: Spindle Assembly

• Process

  • Centrosomes nucleate microtubules

  • Centrosomes duplicate in the S phase

  • Centrosomes move apart along the outside of the nuclear envelope

  • A bipolar football-shape spindle forms, with a centrosome at each pole nucleating microtubules

  • Nuclear envelope breaks down, allowing microtubules to capture chromosomes

• Microtubules are more dynamic in mitosis (compared to interphase)

  • Increased catastrophe frequency during mitosis

  • Increase in dynamics facilitates

    • Breakdown/rearrangement of the interphase microtubule array

    • Attachment of chromosomes to the spindle

• While microtubules are dynamic in mitosis, specific populations are stabilized

  • Astrial microtubules: Microtubules growing out of the centrosome that are highly dynamic and form asters

  • Interpolar microtubules

    • Astrial microtubules that become stabilized by cross-linking proteins

    • Overlap in the center of the spindle and help forms its football shape

      • Formed because microtubule minus ends are at the centrosomes (at two spindle poles), so the overlapping microtubules in the center are of opposite polarity

Kinesin-5

• Specialized kinesin that localizes to the “overlap zone” of interpolar microtubules

  • Specifically, in the center of the spindle to antiparallel interpolar MTs

• Tetrameric kinesin, with two heads on each side

• Both sets of heads can walk to MT plus ends

  • Two heads associate with the top MT, two heads associate with the bottom MT

• Since the MTs are antiparallel, this walking provides outward force

  • Allowing poles ot be pushed apart to promote centrosome separation during spindle assembly

Essential Step 3: Chromosome Capture and Alignment

• After the spindle forms, chromosomes start attaching to microtubules

  • Each sister chromatid has a centromere and a kinetochore

    • Centromere: DNA underlying kinetochore

    • Kinetochore: Protein structure that mediates attachment

  • As a result, microtubules specifically attach to these kinetochores on sister chromatids

    • Multiple microtubules can attach to each kinetochore

• Chromosome “Congression”

  • After the chromosomes attach to the spindle, they are moved to the middle, which is the metaphase plate

Kinetochore Organization

• Core kinetochore proteins

  • Form a platform on the centromere region

• Outer kinetochore proteins

  • Extended proteins that physically bind to the microtubules

Chromosome Capture and Alignment

• Microtubules coming out from centrosomes (“astrial microtubules”) grow and shrink (again, highly dynamic)

• When a mcirotubule hits a kinetochore in the process, it forms an attachment

  • The microtubule “captures” the kinetochore and becomes a “kinetochore microtubule”

• Eventually, chromosomes become “bioriented”

  • One sister is attached to one pole, and the other sister is attached to the other pole

Sister Chromatids Must Attach to Opposite Spindle Poles

• Chromomes must be bioriented to segregate properly

• Once a chromosome is bioriented, there is tension on the kinetochores

• Due to pulling from both sides by microtubules

• The cell can sense this tension, which gives it a way to monitor attachments

Chromosome Dynamics After Attachment

• Even after chromosomes become bioriented, the kinetochore MTs remain dynamic and chromosomes oscillate

• Outer kinetochore proteins form extended structures that bind to the sides of microtubules (not at the very end)

• This allows microtubules to add and lose subunits

  • MTs can remain dynamic without detaching from the kinetochore

Essential Step 4: Chromosome Segregation

• For sister chromatids to come apart, cohesin must be removed

• A protein called “Separase” cleaves the cohesin rings, allowing sisters to come apart

• After cohesin is removed, sister chromatids move to opposite spindle poles

• Two mechanisms for chromosome segregation (that happen at the same time)

  • Anaphase A

  • Anaphase B

Anaphase A

• Chromosomes move to spindle poles

• Movement driven by depolymerization of microtubule plus ends at the kinetochore

• Kinetochore microtubules shorten

• Chromosomes stay attached to shrinking microtubules and are pulled to the pole

Anaphase B

• Spindle poles elongate

• The spindle elongates, driving chromosomes even further apart

• Elongation is driven by:

  • Sliding force from the center of the spindle by kinesin-5

  • Pulling force from dynein on the cell cortex

    • Specifically, b/c dynein at the cell cortex walks towards the minus ends of these astrial microtubules, a pulling force is generated that reels in the astrial microtubule and pulls the centrosome closer to the plasma membrane

Chromosome Segregation Error in Anaphase

• Error in chromosome attachment

• If a single kinetochore becomes attached to both spindle poles, the chromosome will get stuck in the middle during anaphase

Essential Step 5: Cytokinesis

• A ring of actin and myosin forms at the center of the spindle, between the separating chromosomes

  • Contractile ring

• The ring contracts, eventually pinching the cell in two