Bio202 Lecture 14: Cell Division PDF

Lecture 14: Cell Division Today’s agenda: Quick overview of the stages of mitosis: Prophase, prometaphase, metaphase, anaphase, telophase, cytokinesis Focus on the essential events that happen during mitosis: 1) Chromosome duplication and condensation 2) Spindle assembly 3) Chromosome captu...

Lecture 14: Cell Division Today’s agenda: Quick overview of the stages of mitosis: Prophase, prometaphase, metaphase, anaphase, telophase, cytokinesis Focus on the essential events that happen during mitosis: 1) Chromosome duplication and condensation 2) Spindle assembly 3) Chromosome capture and alignment 4) Chromosome segregation 5) Cytokinesis  Walther Flemming 1843-1905 ▪ Coined the term mitosis in the early 1880s ▪ mitos = Greek for thread ▪ Drawings of mitosis, published in 1882! ▪ Mapped out the progressive stages of mitosis ▪ Remarkably accurate! 3  Cell division is a highly dynamic process Sand dollar embryo, fluorescent DNA The dynamic movement of chromosomes during cell division is mediated by a microtubule-based spindle Interphase Mitosis Microtubules in green  Cell division is a highly dynamic process Fluorescent tubulin  Cell division is a highly dynamic process Fluorescent tubulin and DNA  Cell division is a highly dynamic process Fluorescent tubulin, DNA, and plasma membrane  The cell cycle is composed of “interphase” and “mitosis” Figure 5-13 Interphase: ▪ DNA duplicates, centrosomes duplicate, the cell increases in size Mitosis (M phase): divided into six stages ▪ Prophase, prometaphase, metaphase, anaphase, telophase, cytokinesis  From your textbook, Panel 18-1 Stages of mitosis: Prophase ▪ Chromosomes condense (can be seen as individual threads) ▪ Centrosomes move apart  From your textbook, Panel 18-1 Stages of mitosis: 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”)  From your textbook, Panel 18-1 Stages of mitosis: Metaphase ▪ Chromosomes are attached to microtubules from opposite spindle poles ▪ Chromosomes align in the center of the cell (at the “metaphase plate”)  From your textbook, Panel 18-1 Stages of mitosis: Anaphase ▪ Duplicated chromosomes move to opposite spindle poles (one set moves to each pole)  From your textbook, Panel 18-1 Stages of mitosis: Telophase ▪ Contractile ring begins to form at the center of the cell ▪ Nuclear envelope begins to reform  From your textbook, Panel 18-1 Stages of mitosis: Cytokinesis ▪ Contractile ring constricts, creating a cleavage furrow ▪ Cell pinches in two ▪ Chromosomes decondense  Essential steps of cell division Figure 5-13 1) Chromosome duplication and condensation 2) Spindle assembly 3) Chromosome capture and alignment 4) Chromosome segregation 5) Cytokinesis Essential step 1: Chromosome duplication and condensation Mitotic chromosome organization: EM of metaphase chromosome: “Primary constriction” – site of centromere Chromatid = one copy of the chromosome Figure 5-15 DNA duplicates prior to mitosis, Chromosome is so has “sister chromatids” highly condensed  DNA replication duplicates chromosomes Chromosomes duplicate in “S phase” (“Synthesis”) and segregate in “M phase” (“Mitosis”) Replication starts at origins, proceeds bidirectionally (sister chromatids) Sisters linked together Sisters have come Figure 5-14 after replication apart (links are gone)  Sister chromatids are linked by the cohesin complex ▪ A complex of proteins called the “cohesin” complex loads onto the duplicated DNA strands during S phase ▪ The cohesin complex forms a ring that links the two duplicated DNA strands ▪ This “glue” holds the sister chromatids together until anaphase Figure 18-19 Chromosome condensation Humans: 2 meters of DNA in each cell ▪ Chromosomes are condensed in interphase to fit this length of DNA into a 10 μm diameter nucleus ▪ Chromosomes must condense further in mitosis so that they can be easily segregated ▪ Note: images on the left are shown at the same scale Final result: In mitosis, the DNA molecule is 10,000-fold shorter than its extended length! Figure 5-18  Chromosomes are more condensed in M phase Figure 5-1 ▪ Can visualize individual chromosomes in mitosis ▪ Harder to distinguish one chromosome from another in interphase – chromosomes less condensed  Condensins help compact chromosomes in M phase Figure 18-19 ▪ A complex of proteins (called the “condensin” complex) form rings that function to condense the DNA in mitosis ▪ Thought to organize the DNA into loops, which results in full condensation of mitotic chromosomes  Essential step 2: Spindle assembly ▪ Centrosomes nucleate microtubules ▪ Centrosomes duplicate (S phase) ▪ Centrosomes move apart along the outside of the nuclear envelope ▪ A bipolar football-shaped spindle forms, with a centrosome at each pole nucleating microtubules ▪ Nuclear envelope breaks down, allowing microtubules to capture chromosomes Figure 18-22  Reminder: Microtubule dynamic instability ▪ MTs undergo phases of growth and shrinkage by addition/loss of tubulin subunits ▪ This movie shows interphase; dynamics increase in mitosis  Microtubules are more dynamic in mitosis Average lifetime of an individual microtubule: Interphase = 5 minutes Mitosis = 15 seconds Interphase Mitosis Increased catastrophe frequency during mitosis Increase in dynamics facilitates: ▪ Breakdown/rearrangement of the interphase microtubule array ▪ Attachment of chromosomes to the spindle  Microtubule dynamics in the mitotic spindle ▪ While microtubules are dynamic in mitosis, specific populations are stabilized ▪ Microtubules growing out of the centrosome are highly dynamic and form asters – these are “astral microtubules” ▪ Some of these microtubules become stabilized by cross-linking proteins and become “interpolar microtubules” - these microtubules overlap in the center of the spindle and help form its football shape ▪ Microtubule minus ends are at the centrosomes (at the two spindle poles), so the overlapping microtubules in the center are of opposite polarity (forming Figure 18-24 the “overlap zone”) (5th edition) Kinesin-5 is a specialized kinesin that localizes to the “overlap zone” of interpolar microtubules Simplified diagram of overlap zone with two microtubules: Kinesin-5 ▪ Kinesin-5 is a 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  Kinesin-5 slides overlapping microtubules in the center of the spindle apart Spindle Kinesin-5 microtubule ▪ Kinesin-5 localizes in the center of the spindle to antiparallel “interpolar” MTs (in the “overlap zone”) ▪ Each set of heads walks to a MT plus end ▪ Since the MTs are antiparallel, this walking provides outward force, pushing poles apart to promote centrosome separation during spindle assembly  Essential step 3: Chromosome capture and alignment From your textbook, Panel 18-1 ▪ After the spindle forms, chromosomes start attaching to microtubules ▪ After the chromosomes attach to the spindle, they are moved to the middle (the metaphase plate) – this is called chromosome “congression”  Microtubules attach to kinetochores Figure 18-24 Kinetochore = Centromere = Protein structure DNA underlying that mediates kinetochore attachment Kinetochore Each sister chromatid Multiple has a centromere and microtubules a kinetochore attach to each kinetochore Kinetochore organization: Figure 18-24 ▪ “Core” kinetochore proteins in red – form a platform on the centromere region ▪ “Outer” kinetochore proteins in blue – extended proteins that physically bind to the microtubules  Chromosome capture and alignment “Search and capture” process: Bioriented chromosome: Dynamic astral microtubules capture ▪ Microtubules emanating from centrosomes (“astral microtubules”) grow and shrink ▪ When a microtubule hits a kinetochore, 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 Bioriented chromosome: Incorrect Incorrect Correct! ▪ Chromosomes must be bioriented to segregate properly ▪ Once a chromosome is bioriented, there is tension on the kinetochores (pulling from both sides by microtubules) ▪ The cell can sense this tension, which gives it a way to monitor attachments Figure from Alberts, Molecular Biology of the Cell Chromosome dynamics after attachment: ▪ Even after chromosomes become bioriented, the kinetochore MTs remain dynamic and chromosomes oscillate How do chromosomes stay attached to the spindle when the microtubules are growing and shrinking? Cultured rodent cell, microtubules, kinetochores and spindle poles Kinetochore organization : Figure 18-24 ▪ 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; microtubules can remain dynamic without detaching from the kinetochore  Essential step 4: Chromosome segregation ▪ For sister chromatids come apart, cohesin must be removed ▪ A protein called “separase” cleaves the cohesin rings, allowing sisters to come apart Figure 18-19, 18-28 After cohesin is removed, sister chromatids move to opposite spindle poles Figure 18-21 ▪ Two mechanisms for chromosome segregation: “Anaphase A” and “Anaphase B” ▪ These happen at the same time Anaphase chromosome movements: 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 Figure 18-29 Anaphase chromosome movements: Anaphase B: spindle poles separate The spindle elongates, driving chromosomes even further apart Elongation is driven by: Dynein Kinesin-5 1) Sliding force from the center is here is here of the spindle, by kinesin-5 2) Pulling force from dynein on the cell cortex Figure 18-29 How dynein contributes to Anaphase B spindle pole separation ▪ Astral microtubules extend towards the cell cortex ▪ Dynein at the cell cortex walks towards the minus ends of these microtubules ▪ This generates a pulling force that reels in the astral microtubule and pulls the centrosome closer to the plasma membrane What happens if there is an error in chromosome attachment? Incorrect Incorrect Correct!  Chromosome segregation error in anaphase: Cultured rodent cell, fluorescent tubulin, kinetochores, and spindle poles ▪ If a single kinetochore becomes attached to both spindle poles, the chromosome will get stuck in the middle during anaphase ▪ Note: You can see both Anaphase A and Anaphase B in this movie if you look at the chromosomes that are separating properly  Essential step 5: Cytokinesis Figure 18-20 ▪ A ring of actin and myosin forms at the center of the spindle, between the separating chromosomes – this is the ”contractile ring” ▪ The ring contracts, eventually pinching the cell in two Cytokinesis: Figure 17-39, 18-20, 18-33 Reminder from lecture 11: ▪ Myosin II motors can arrange in a bipolar fashion (heads pointed out) ▪ Actin filaments in the contractile ring are arranged in an antiparallel fashion ▪ Myosin heads on each side walk to actin plus ends ▪ Filaments slide relative to each other, which moves the filaments closer together and “contracts” the ring  Cytokinesis: the contractile ring Figure 18-33 Fluorescent myosin II, concentrated in the contractile ring  Wignall lab research Female reproductive cells Most cells (oocytes) Centrosome-mediated No centrosomes! Errors rare 10-30% error rate in humans!  Wignall lab research ▪ How are spindles formed without centrosomes? ▪ How are chromosomes aligned and segregated? Next lecture: Apoptosis (cell death)