Lecture 22- Autophagy II

Question 1

Describe the role of autophagy as a housekeeping process

Answer 1

Actively works at a low basal rate inside the cell to remove any damaged material in the cell.

Does this by selectively removing damaged proteins and organelles

Question 2

What accumulates in cells lacking autophagy?

Answer 2

Protein aggregates

Question 3

Why are neurons particularly sensitive to damage?

Answer 3

Because they are extremely long-lived cells, metabolically active and have long protrusions. This makes trafficking the proteins more difficult/less efficient

Question 4

What does neuronal-specific autophagy disruption is mice cause?

Answer 4

1. Accumulation of ubiquitinated aggregates

2. Increase apoptosis

Question 5

How can apoptosis be viewed/studied?

Answer 5

Using TUNEL staining

Question 6

What are proteinopathies?

Answer 6

Diseases where proteins are abnormally self-associating and aggregating due to confirmations changes

Question 7

What proteinopathy causes Huntington’s disease?

Answer 7

Huntingin protein aggregates

Question 8

What proteinopathy causes Alzheimer’s disease?

Answer 8

Amyloid-beta plaques aggregates

Question 9

What proteinopathy causes Parkinson’s disease?

Answer 9

alpha-synclein aggregates

Question 10

What is similar and different in the protein aggregates in Huntington’s, Alzheimer’s and Parkinson’s disease?

Answer 10

All share common phenotype that they have accumulation of ubiquitinated inclusions

However the proteins and areas in the brain where accumulation occurs is different so the symptoms can vary

Question 11

What causes Huntington’s disease?

Answer 11

Caused by polyQ expansion because it causes a mutation on the Huntingtin protein, making it missfold and aggregate

These proteins accumulate over time, causing the disease to worsen as you age

Question 12

Why does Huntington’s disease worsen with age?

Answer 12

Your lysosomal capacity and therefore your ability to remove the protein aggregates deceases with age.

As you get older, the accumulation of protein aggregates outpaces the ability to degrade the protein causing you to accumulate them

Question 13

What is the association of polyQ expansion and Huntington’s disease?

Answer 13

The more polyQ you have, the more susceptible to the disease you are and the earlier the onset

PolyQ less than 18 = healthy
PolyQ more than 35 = disease causing

Question 14

What are some mechanisms of toxicity?

Answer 14

1. Loss of normal protein function
2. Toxic oligomer
3. Aggresomes themselves may be toxic
4. Proteasomal damage
5. Adaptor sequestration

Question 15

What does the aggregation of alpha-synuclein cause in Parkinson’s disease?

Answer 15

Causes a loss of dopinergic neurons

Question 16

How is alpha-synuclein protein normally degraded?

Answer 16

1. By chaperone-mediated autophagy
2. The LAMP2 receptor on the lysosome surface is able to recognise specific amino acid sequences/proteins
3. These are then transported directly into the lysosome to be degraded

Question 17

How does alpha-synuclein mutations affect chaperone mediated autophagy?

Answer 17

1. The A53T mutated alpha-synuclein blocks the CMA pathway
2. The protein can still bind to the LAMP2 receptor but cannot be transported into the lysosome
3. Therefore, mutated alpha-synuclein cannot be degraded, forms aggregates and causes toxicity

Question 18

How does damaged mitochondria link with Parkinson’s disease?

Answer 18

1. Damaged mitochondria accumulate in Parkinson’s and are a big source of ROS
2. ROS damages cellular components
3. Therefore, Parkinson’s maybe caused by mitochondrial-derived oxidative damage

Question 19

What genes have been identified in familial Parkinson’s?

Answer 19

PINK1 and PARKIN genes

Question 20

How do PINK and PARKIN proteins normally function?

Answer 20

1. PINK and PARKIN collaborate to recruit ubiquitin to the surface of mitochondria and target them for mitophagy
2. If the mitochondria get depolarised, PINK1 is recruited which activates PARKIN

Question 21

What happens when PINK1 and/or PARKIN function is lost?

Answer 21

1. Loose the ability to target damaged mitochondria for mitophagy so it starts to accumulate
2. Accumulated damaged mitochondria work less efficiently so release more ROS and cause more cellular damage due to more protein misfolding
3. Feedback loop generated- as soon as you accumulate damaged mitochondria, start to damage even more

Question 22

Which steps of the autophagy pathway can be mutated and cause disruptions?

Answer 22

1. Impairing autophagosome formation
2. Disrupting cargo recognition
3. Cause autophagic cargo
4. Disrupting lysosomal formation
5. Inhibits autolysosome formation
6. Preventing secretion

Question 23

What causes cancer?

Answer 23

Accumulation of damaged DNA

Question 24

How is autophagy anti-oncogenic?

Answer 24

Before tumours have formed, autophagy is beneficial for cell homeostasis, damage removal, reducing ROS and inflammation

Question 25

How is autophagy pro-oncogenic

Answer 25

After tumour formation, autophagy can help tumour survival during starvation and chemotherapy and prevent apoptosis

Question 26

What are 3 autophagy therapy strategies?

Answer 26

1. Blocking survival to metabolic stress with autophagy inhibitors
2. Inhibit autophagy to increase apoptosis during chemotherapy
3. Elevate autophagy to remove damage and prevent cancer

Question 27

How does autophagy inhibit apoptosis?

Answer 27

1. Beclin 1 forms a complex with vps24 and UVRAG which makes a PIP lipid on the ER
2. This leads to the formation of the autophagosome
3. Apoptosis is driven by membrane permeabilisation caspase activation which is activated by Bcl2
4. Bcl2 and Beclin 1 directly interact with each other
5. When you activate autophagy you move Bcl2 away from the mitochondria and inhibit apoptosis
6. Therefore, autophagy promotes survival and inhibits apoptosis
7. However, if you inhibit autophagy, you promote apoptosis and inhibit survival

Question 28

What was observed and confirmed during experiments on Beclin 1 mice mutants?

Answer 28

Beclin 1 mice mutants accumulate tumours and don’t survive compared to wildtypes

Demonstrating Beclin 1 is a tumour suppressor