CellSig11 - 31

Question 1

How was notch identified?

Answer 1

Forward genetic screens in Drosophila - notch wing phenotype - by Thomas Hunt Morgan - 1866-1945

Question 2

How many notch receptors does Drosophila have?

Answer 2

1

Question 3

How many delta ligands do Drosophila have?

Answer 3

2 - delta and serrate

Question 4

How many delta ligands do vertebrates have?

Answer 4

At least 9 members of this family

Question 5

How many notch receptors do humans have?

Answer 5

4 (Notch 1-4)

Question 6

What are delta ligands collectively known as?

Answer 6

DSL family

Question 7

Characterise delta/notch structures and function

Answer 7

1TM domains, with many EC EGF repeats, juxtacrine in function, notch also has many IC ankyrin repeats

Question 8

What is important about EC EGF repeats?

Answer 8

Made up of repeated cysteines in regular intervals that form sulphide bridges, adding structure

Question 9

What regulates EGF cysteine pairing?

Answer 9

Protein disulphide isomerase in the ER

Question 10

Outline the delta-notch pathway

Answer 10

Notch activation > Notch-intra translocates to the nucleus > activation of CSL > transcription

Question 11

What constitutes CSL?

Answer 11

CBF1/SuH/LAG1

Question 12

What is furin’s role in notch activation?

Answer 12

Cuts the receptor to produce a heterodimer, S1, which then implants into the PM

Question 13

What is furin?

Answer 13

Serine protease - controls first cleavage of notch

Question 14

How is notch cleaved for a second time?

Answer 14

Ligand dependent, TACE cuts off EC domain, but notch remains in membrane because TM domain still attached

Question 15

What is TACE?

Answer 15

Metalloprotease

Question 16

What is presenilin?

Answer 16

Gamma-secretase - controls third cleavage, and release of notch from the PM

Question 17

What releases notch from the PM?

Answer 17

Presenilin

Question 18

Outline notch degradation

Answer 18

E1 - ubiquitin activating enzyme > E2 - ubiquitin conjugating enzyme > E3 ubiquitin ligase (notch specific) > proteasome

Question 19

Characterise Su(dx)/Itch

Answer 19

3 major domains - WW binds notch, HECT mediates ubiquitination, C2 localises to the PM

Question 20

Characterise Sel-10

Answer 20

Recognises Notch-intra, but lacks ubiquitin ligase activity, so recruits one via its F box

Question 21

Characterise Neur/Mib

Answer 21

Binds delta, RING domain ligases, required to activate endocytic pathway

Question 22

What domains are present in notch-intra?

Answer 22

NLS - nuclear localisation sequence, RAM and ANK domains - protein/protein interaction domains, activation domain - transcriptional activator

Question 23

Outline transcriptional activity without Notch present

Answer 23

CBF1 anchors HDAC via SMRT and SKIP near the promoter, reducing DNA acetylation

Question 24

Outline transcriptional activity with Notch present

Answer 24

Notch-intra displaces SMRT and HDAC and acts as a transcriptional activator

Question 25

How is Alzheimers related to Notch activation?

Answer 25

Similar mechanism - amyloid precursor protein (APP) cleaved by alpha or beta-secretase, then by presenilin in one of two places (40 or 42 a/a). The 42 a/a version is a major component of the amyloid plaques

Question 26

How was the first human notch cloned?

Answer 26

From T-cell acute lymphoblastic leukemia patients - Truncated Allele of Notch (TAN1)

Question 27

What is TAN1?

Answer 27

Translocation of TCR-beta gene fused to Notch-intra

Question 28

How does TAN1 cause leukemia?

Answer 28

TCR-beta strongly expressed in immature T-cells, therefore Notch is made consituently active

Question 29

What does CADSIL stand for?

Answer 29

Cerebral AD ateriopathy with subcortical infarcts and leukoencephalopathy

Question 30

What is CADSIL?

Answer 30

Recurrent strokes leading to dementia, due to build of granular material in smooth muscle of blood vessels

Question 31

What causes CADSIL?

Answer 31

Mis-sense mutations in Notch3 - usually the first 5 EGF repeats