Biocore 587 Unit 2

Spindle

structure based on microtubules and DNA that assembles in the M-Phase

Cytokinetic Apparatus

actin and myosin that cause the cell to contract during separation

Bipolar

two different ends, microtubules have plus and minus ends, Plus ends line up at the chromatin and Minus ends line up at the centrosome

Kinetichore (KT)

protein-based structures that bind MTs and link sister chromatids to the spindle. In metaphase, they are bound to the plus ends of MT Spindle Assembly

Spindle Composition

MTs are made of tubulin dimers, 1 alpha and 1 beta-tubulin subunit, each subunit binds to GTP, only the beta subunit can hydrolyze GTP

Centrosomes

MT organizing center based on centrioles, and they recruit Gamma-Turcs from the cytoplasm and concentrate it, which promotes MT nucleation

Chromatin

DNA, histones, and proteins that bind DNA, organized as condensed sister chromatids

Centromere

 a stretch of DNA where the kinetochore assembles

Self-assembly

the formation of structures that don't require continuous energy input

Catastrophe

transition from polymerization (growing) to depolymerization (shrinking)

Rescue

 transition from depolymerization (shrinking) to polymerization (growing)

Self-Organization

production of structures that require continuous energy investment, underlain by iterative cycles, structures arise gradually and produce

EB1

end binding protein, binds to growing plus ends of MTS

ASP

end binding protein, binds to minus ends and stabilizes them

Gamma-Turc

gamma tubulin ring complex that promotes MT polymerization, is concentrated at the centrosomes and other MT organizing centers (MTOCs)

Side-Binding proteins

bind MT sides and reduce the frequency of catastrophe (Example: Ensonsin)

Severing Proteins

bind to MT sides and extract a dimer from the MT lattice, causing the MTs to break

Motor Proteins

use ATP to move things with respect to the MTs

Cargo

what moves along MTs by the motor proteins, they can be many different things

Kinesins

transport cargo to the plus ends, can power slide the MTs within the spindles, and can also shorten MTs by removing dimers from the ends

Dyneins

move cargo toward the minus ends of MTs, help organize spindles, and power flagella rotation

CDK1

master regulator of the M-Phase, phosphorylation activates, and this phosphorylates hundreds of different proteins, all of which collectively stimulate entry into the M-Phase

CDK2

when active, phosphorylates proteins that promote the S-Phase events

Search and Capture Model Evidence

1. Spindle fits the known geometry of centrosomes and condenses chromosomes

2. Electron microscopy reveals that individual MTs can extend from poles to kinetochores

3. Most MTs grow out from the centrosomes

Chromatin/Chromatid Nucleation

states that spindle assembly results from MTs forming near or around chromatin

Chromatin/Chromatid Nucleation Evidence

1. Many spindles do not have centrosomes

Redundant Mechanisms

important to ensure the process is complete, very important for critical processes (Example: Search and Capture as well as Chromatin Nucleation)

Ran-GTPase

small GTPase, during interphase it regulates nuclear importin-cargo, during M-phase it regulates spindle assembly

Spindle Assembly Factors (SAFs)

proteins that, when activated, promote MT nucleation and stabilize MTs

RCC1

Ran’s GEF, located on the chromatids

Ran-GAP

soluble in M-phase and is located all around the cell

Inner KT Plate

close to chromatin, contains CenpA, CenpC, and the Mis-12 Complex, and remains as compact spots

Translucent Plate

middle layer, contains the Ndc80 Complex and the Knl1 Complex

Outer KT Plate and Fibrous Corona

contains the CenpF, CenpE, Mad2, and Ndc80 complex, very dynamic during prometaphase

CenpA

a variant of Histone H3, serves as a binding site

CenpC

binds to CenpA and serves as a binding site for the rest of the KT assembly

Knl1 Complex

scaffold for spindle assembly checkpoint proteins, orchestrates the Spindle Assembly Checkpoint

Mis12 Complex

part of the Inner KT Plate, physical link, connecting centromeric chromatin (specifically the protein CENP-A) to the outer kinetochore components

Ndc80 Complex

a large, linear complex that binds to the MTs, contains Hec1 and Nuf2

CenpE and CenpF

in the outer KT plate and fibrous corona, they both bind to MTs, dynamic

Spindle Assembly Checkpoint (SAC)

a mechanism that prevents anaphase entry until all KTs are attached to the KT MTs, mitotic checkpoint

Mitotic Checkpoint Complex (MCC)

 KTs recruit Bub1, BubR1, + Bub3. The Bubs recruit Mad 1+Mad 2, which leads to the formation, inherently unstable and has to be produced continuously to prevent transition

Anaphase

stage of M-phase following metaphase, the point where the biochemistry of the cytoplasm changes radically, and the process by which chromosomes are separated

Anaphase A

movement of Chromosomes to the poles

Anaphase B

movement of poles away from one anothe

Cdc20

 another protein that can associate with MCC, which prevents Cdc20 from activating the Anaphase Promoting Complex (APC)

Anaphase Promoting Complex (APC)

ubiquitin ligase attaches a small protein called ubiquitin to other proteins as targeted degraders, Two APC targets: Securin and Cyclin B, The attached KTs result in low MCC Content because of degradation. Cdc20 activates APC and the degradation of APC targets

Cyclin B

degradation leads to inactivation of Cdk1, and MT catastrophe frequency starts to drop

Securin

 protein that links sister chromatids together. High APC content leads to its degradation, which means the complex that holds chromatids together is disassembled

Caged Fluorescein

a caging group that renders the molecule non-fluorescent and a fluorescent molecule inside, the bonds that link them are UV liable

Poleward Flux Severing Proteins

Spastin and Fidgetin

Pacman Flux Severing Proteins

Katanin

Taxol (Paclitaxel)

MT stabilizer and cancer drug that prevents anaphase, Concentration in tumors was shown to be the same amount needed to produce a multipolar spindle, but a much lower amount than what was needed for mitotic arrest

Kinesins

move toward the plus ends

Dyneins

move toward the minus ends