Cell growth + division lecture 2

Question 1

Tubulin

Answer 1

• Building block of spindle
• • and 0 end
• If meets chromosome, becomes stabilised

Question 2

Centrosome

Answer 2

• Formed around centrioles, 2 centrioles liked per centrosome
• Centrosome organises - ends of microtubule at opposite end of cell
• Overlapping microtubules captured by Eg5
• Dynein = at - pole

Question 3

Kinesin + dynein

Answer 3

• Force generating ATPases

- Move chromosomes

Question 4

Kinetochore

Answer 4

• Kinetochore proteins
• CENPA + CENPC form template on DNA, NDC80 + KNL1 sit on top
• When attached, microtubule pulls back, generates force
• TIRF microscopy, NDC80 + CENP-T track + end of microtubule

Question 5

Spindle attachment geometry

Answer 5

• Correct = amphitelic
• Incorrect = monotonic, systelic + merotelic
• Need way of sensing

Question 6

Aurora kinases

Answer 6

• Adopt similar active conformation to cyclin bound to Cdk?

- Phosph to stabilise AS and have nearly identical consensus motif

Question 7

Gradient of aurora

Answer 7

• Gradient of aurora A at spindle, B at centromere
• ↑ aurora A = kinetochore released from microtubule (at pole released + moves to middle
• Aurora B = near centromere, initial attachment = unstable due to aurora B
• Aurora B phosph NCD80 + KNL1, ↓ affinity for microtubule
• w/ tension, pull away, escape aurora B → dephosph

Question 8

Spindle checkpoint

Answer 8

• Check chromosome alignment
• Based on number of free kinetochores
• MAD/BUB regulate APC. Localise to kinetochores x attached to microtubules

Question 9

MPS1

Answer 9

• Senses + binds unattached kinetochores
• MPS1 phosph MELT in KLN1
• Aurora B phosph NDC80 → MPS1 binds phosph NDC80 → phosph MELT KLN1 → signals to SAC pathway
• Cdk activates MPS1 by phosph

Question 10

BUB3

Answer 10

• Reads phosph of KLN1

- Binds GLEBS motif in BUB1/BUBR1

Question 11

MAD2

Answer 11

• Open/closed conformation
• Close locks w/ Cdc20 + forms part of MCC
• MPS1 phosph. unattached kinetochore + recruits to checkpoint complex
• MPS1 phosph MAD1 which is in complex with Mad2c + recruits Mad2o
• Mad1-Mad2c-Mad2o-Bub1-Bub3-Cdc20 → MCC (Mad2c-BubR1-Bub3-Cdc20)

Question 12

APC/C regulation

Answer 12

• Cdc20 binds cyclin
• MCC inhibits APC/C by occupying substrate bs w/ BUBR1
• BUBR1 captured by 2nd CDC20 in MCC + MAD2
• At start of M, ACC phosph by cyclin B → active, binds Cdc20 when unattached kinetochore → inhibited by MCC
• MCC turnover by TRP13
• When kinetochores attached → MCC x made but is turned over so x inhibit ACC