3/10/15 1 Mitosis Reading: ECB3 625-636 ECB4 616-632 Learning Objectives: • Understand the events that occur prior to M-phase and necessary for mitosis • Understand the organization of microtubules in the mitotic spindle and which associate with centrosomes and kinetochores • Understand the events that occur in each phase of mitosis • Understand how chromosomes become attached to appropriate microtubules: ‘bi-orientation’ • Understand how the checkpoint in mitosis is satisfied • Understand the movements and mechanisms of anaphase A and B • Understand the mechanisms of cytokinesis in animal and plant cells Last Lecture…….3/10/15 2 Figure 17-25 Molecular Biology of the Cell P • The Wee1 kinase puts on an inhibitory phosphate to keep M-Cdk inactive until the G2 checkpoint. Figure 17-25 Molecular Biology of the Cell P • The Wee1 kinase puts on an inhibitory phosphate to keep M-Cdk inactive until the G2 checkpoint. • When the G2 checkpoint is satisfied, then the phosphatase Cdc25 counteracts the inhibition by removing the inhibitory phosphate In essence, the Wee1 kinase enforces the G2 checkpoint until relieved by activation of the Cdc25 phosphatase3/10/15 3 Mitosis The events in the cell cycle are ordered by checkpoints and driven by activation of cyclin dependent kinases (Cdks) Figure 18-1, 18-3 combined Essential Cell Biology Cdks- checkpoint is: - 'activated' when it pauses the cell cycle to allow completion of the events of the previous phase - 'satisfied' when all the events of the previous phase are completebefore mitosis can occur, DNA in chromosomes has to be duplicated (S phase) Duplicated chromosomes are held together by cohesion proteins. Duplicated chromosomes consists of 2 chromatids, tightly joined at the centromere.3/10/15 4 Today’s topics 1. Events in S-phase. 2. Overview of events in M-phase 3. The mitotic spindle 4. Chromosome attachment in prometaphase and the checkpoint in mitosis 5. Metaphase to anaphase: separation of chromosomes 6. Cytokinesis and other events in M-phase Events in S Phase • Chromosomal DNA is duplicated precisely; mechanisms exist to ensure that it is not over-duplicated • Cohesins hold the duplicated chromatids together Duplicated Chromosome Centromere Chromatid Chromatid Figure 18-3, 18-15 Essential Cell Biology During S and G2 phases of the cell cycle, the interphase centrosome duplicates3/10/15 5 Additional events in S/G2 Phases and the G2 checkpoint • The single centrosome is duplicated in preparation for M phase Figure 18-1, 18-22 Essential Cell Biology Start of Mitosis Figure 17-25 Molecular Biology of the Cell The G2/M transition: activation of M-Cdk • Everything OK? Is the G2 checkpoint satisfied? • Let’s go into M phase! at end of G2, the G2 checkpoint assesses whether DNA is fully duplicated and has no damage - if there is damaged or unreplicated DNA then the cell cycle is activated while these events are finished or corrected - once everything is fine, then the cell moves into M phaseM phase is driven by rapid activation of M-Cdk3/10/15 6 Today’s topics 1. Events in S-phase. 2. Overview of events in M-phase 3. The mitotic spindle 4. Chromosome attachment in prometaphase and the checkpoint in mitosis 5. Metaphase to anaphase: separation of chromosomes 6. Cytokinesis and other events in M-phase M phase M-phase consists of two distinct, but overlapping, parts: • The precise and equal segregation of the duplicated chromosomes by the microtubule-based mitotic spindle – MITOSIS • The division of the cytoplasm in two halves to generate two cells by contraction of the microfilament-based contractile ring* - CYTOKINESIS * It is different in plants 2 parts of M phase: 1) mitosis: precise and equal segregation of the duplicated chromosomes by the microtubule based mitotic spindle 2) cytokinesis: division of the cytoplasm by microfilament- based contractile ringMitosis is in 5 stages3/10/15 7 Stages of Mitosis Prophase Prometaphase Metaphase Anaphase Telophase The mitotic spindle is a machine to distribute the chromosomes equally with a built in quality control system (the checkpoint). Cytokinesis Interphase Interphase M-Phase Today’s topics 1. Events in S-phase. 2. Overview of events in M-phase 3. The mitotic spindle 4. Chromosome attachment in prometaphase and the checkpoint in mitosis 5. Metaphase to anaphase: separation of chromosomes 6. Cytokinesis and other events in M-phase3/10/15 8 Microtubules: Review + + + + + + + - - - - + + + + + + + + + + + + + + + + + - - - - - - Favored for growth Associated with centrosome/spindle pole 13 protofilaments Figure 17-8, 17-9 Essential Cell Biology The Mitotic Spindle Parts List The spindle contains three classes of microtubules (shown at metaphase) The chromosomes (shown at metaphase) + + + + + + + + + + + + + + + + + Figure 18-24, 18-25 Essential Cell Biology Cohesins holding chromatids together 3 classes of microtubules are involved in mitotic spindle: 1) Aster: extends out to cell periphery 2) Kinetochore: extend from centrosome with plus end associated with kinetochore 3) Interpolar: extend from each pole and inter-digitate in the middle region of the spindle *all classes have their minus ends associated with the spindle poles*3/10/15 9 The Mitotic Spindle Parts List + + + + + + + + + + Figure 18-34 Molecular Cell Biology, Sixth Edition Kinetochore assembly Bioriented chromosome chromatid: - each one has a kinetochore: specific region associated with the centromeres to which the plus end of kinetochore microtubules can attachMetaphase: all chromosomes are bi-oriented and aligned at the metaphase plate3/10/15 10 Today’s topics 1. Events in S-phase. 2. Overview of events in M-phase 3. The mitotic spindle 4. Chromosome attachment in prometaphase and the checkpoint in mitosis 5. Metaphase to anaphase: separation of chromosomes 6. Cytokinesis and other events in M-phase Modified from Figure 17-38 Molecular Biology of the Cell Microtubule capture by kinetochoresres3/10/15 11 Figure 17-39 Molecular Biology of the Cell aaaaaaaaaaaaaaaaaaaaaaaaaaa Microtubules can capture chromosomes in many ways. How is the correct attachment selected? Figure 17-39 Molecular Biology of the Cell Tension creates a stable bipolar attachment3/10/15 12 Aurora B Kinase Figure 18-31 Molecular Cell Biology