Introduction to MCB 252 Topic 26 Microtubules in MitosisProf David Rivier MCB 252 Spring 2015 MCB 252 Topic 26 Motors in Mitosis Reading Lodish 18 6 Roles of MTs in Interphase and Mitosis Lodish 18 5 In Mitosis 1 Cease Interphase Roles Organelle transport and positioning 2 New Role Attach to Position and Move Chromosomes Roles of MTs in Interphase and Mitosis Lodish 18 5 What Happens During this Transition Events in Mitosis Interphase prophase Mitosis DNA Compaction Angel Hair Pasta in Lincoln Theater 2 x 3 x 109 bp x 34 nm bp 2 meters DNA nucleus Scale by 1 000 000 Nucleus 10 mm 10 meters DNA diameter 2 nm 2 millimeter DNA length 2 meters 2000 kilometers 1200 miles UIUC to Orlando Florida 1066 miles Events in Mitosis Events in Mitosis part 2 Roles of MTs in Interphase and Mitosis In Mitosis Lodish 18 5 1 Cease Interphase Roles Organelle transport and positioning 2 New Role Attach to Position and Move Chromosomes Regulation of Dynamic Instability in Mitosis Balance of MAPs v Catastrophe Factors Dynamics Increased Gamma TURC at centrosomes Three Classes of Mitotic Microtubules Kinetochore Polar and Astral Chromosome Segregation General Features Structure of Kinetochore by EM EM of MTs attached to the Kinetochore Spindle Assembly and Function 2 Opposite Types of Chromosome Movements 1 Prior to metaphase move chromosomes to middle a capture chromosomes b move chromosomes to middle 2 Anaphase Move chromosomes to poles Proper Attachment to 2 Spindles Proper attachment tension stabilize MT at Kinetochore stabilize capture Improper attachment no tension de stabilize MT de stabilize release Sensing Tension Proper attachment tension stabilize MT at Kinetochore stabilize capture Improper attachment no tension de stabilize MT de stabilize release Opposing Forces Act on Chromosomes During Congression Opposing Forces Act on Chromosomes During Congression Motors bind chromosomes at sites other than kinetochores Note how Similar this Process Looks to Centering of MTs in Interphase Spontaneous Spindle Assembly in the Absence of Centrosomes How do Do We Get More MTs in the Vicinity of the Chromosomes than in the Remainder of the Cell How Do MTs Find the Chromosomes How is the Nucleo cytoplasmic trafficking regulated Ran Ras related nuclear protein GEF GAP Small GTPase of 24 kDa Role of Ran in Spindle Assembly Ran GTP signals MAPs locally stabilizes MTs and Promotes Assembly of MTs Ran GEF bound to chromosomes high concentration of Ran GTP around chromosomes stabilize and promote assembly near chromosomes part of capture mechanism in vivo Ran GAP evenly distributed in cytoplasm therefore only high conc of Ran GTP is around the chromosomes RAN GTP Concentration Gradient Favors MT Assembly and Stabilization in the Vicinity of the Chromosomes RAN GTP Facilitates Search and Capture How do the MTs that are Attached to the Kinetochores Reel In the Chromosomes More MT Dynamics 1 In Prophase Prometaphase and Metaphase MTs Treadmill 2 In Anaphase MTs de polymerize from both ends Key point energy of de polymerization contributes to movement of chromosomes in Anaphase A Speckle Microscopy Kinetochore Microtubules Treadmill Treadmilling Prior to Anaphase Why is this puzzling Tubulin Treadmilling How do both treadmilling and anchoring occur simultaneously Anaphase A Motors and De polymerization How to Harness the Energy of De polymerization Ring Proteins Identified Genetically in Yeast Dam1 Rings on Yeast MTs What happens when MTs De polymerize Sliding of Rings During Disassembly of MTs Rings slide together Dam1 Ring Surfs Down De polymerizing MT Energy of De polymerization Drives Chromosome Movement in Anaphase A Dam1 Ring Surfs Down De polymerizing MT Ring Also Provides a Possible Answer to How the Kinetochore can Hold onto the End and also Allow Treadmilling Ring in between Plus end and Dyneins Motors and Movements in Anaphase B Sensing Tension PO4 by Aurora Kinase Less Stable No PO4 by Aurora Kinase More Stable Proper attachment tension stabilize MT at Kinetochore stabilize capture Improper attachment no tension de stabilize MT de stabilize release