CMB2001/L05 Transcriptional Responses to Stress & Infection I

Question 1

What 3 factors are required for adaptation to environmental challenge?

Answer 1

Sensors
Signalling pathway
Effector

Question 2

What is the common role of the NF-kB, p53 and HIF pathways?

Answer 2

Allow the cell/organism to respond to environmental threats

Question 3

Give 3 roles of the NF-kB family of transcription factors.

Answer 3

Regulating inflammation
DNA damage
Cell death
Cell adhesion
Proliferation

Question 4

What does the Rel Homology Domain of mammalian NF-kB encode?

Answer 4

DNA bin and dimerisation functions of NF-kB

Question 5

What precursor proteins are p50 and p52 proteolytically processed from?

Answer 5

p105 and p100 respectively

Question 6

What allows p100 and p105 to function as lkB-like inhibitors?

Answer 6

They contain ankyrin repeats in C-termini

Question 6

Name 2 non-conserved transcriptional activation domains in NF-kB.

Answer 6

TA1/TA2
TAD
SD1
SDII

Question 7

What form do NF-kB take to bind DNA?

Answer 7

Dimers

Question 8

What is the function of E3 Ubiquitin ligase?

Answer 8

Facilitates attachment of ubiquitin chains to target protein

Question 9

Describe the Ubiquitin-proteasome pathway of protein degradation. (6)

Answer 9

Ub conjugated to proteins for degradation by ATP-dependent process
5-Ub chain attached to substrate
Recognition by 26S proteasome
UB removed and protein linearised
Injected into proteasome core
Digested to peptides then amino acids by peptidases or used for antigen presentation

Question 10

What can induce NF-kB? (3)

Answer 10

Inflammatory cytokines
Bacterial products
Viral proteins & infection
DNA-damage
Cell stress

Question 11

Describe activation of NF-kB. (8)

Answer 11

Ligand binding
Activated IKK Kinase
IkB kinase complex
NF-kB in inactive cytoplasmic complex
IkB is phosphorylated
IkB is ubiquinated
IkB is proteolytically degraded
NF-kB translocates to nucleus

Question 12

Give the 3 core subunits of the IkB complex.

Answer 12

IKKa
IKKB
IKKy (NEMO)

Question 13

Give the 3 inhibitors of NF-kB.

Answer 13

IkBa
IkBB
Bcl-3

Question 14

How are ikB proteins similar to p105 and p100? (3)

Answer 14

Contain ankyrin repeat motifs (ANK) in C termini
PEST
Domain rich in proline, glutamate, serine and threonine

Question 15

Using NF-kB, give the sensor, signalling pathway and effector response of the body to an environmental challenge.

Answer 15

TNF receptor
IkB complex
NF-kB transcription factors

Question 16

What 2 pathways can lead to activation of NF-kB?

Answer 16

Classical/ canonical pathway
Non-canonical (alternative) pathway

Question 17

How are many NF-kB genes shared?

Answer 17

Between subunits

Question 18

Which part of NF-kB dimer faces in the 3’ and 5’ direction?

Answer 18

5’ - p50
3’ - RelA

Question 19

Describe the regulation of NF-kB function. (4)

Answer 19

Translocation to nucleus
Transcriptional repression, transcriptional activation, promoter targeting and selectivity

Question 20

What is Swi/Snf?

Answer 20

ATP-dependent chromatin remodeller (can move/remove nucleosomes)

Question 21

What is the role of the beta-interferon enhancer?

Answer 21

Regulates expression of the beta interferon gene, particularly in response to viral infections

Question 22

What complex is formed by the beta-interferon enhancer and what is its purpose?

Answer 22

Enhanceosome
High affinity platform for recruitment of transcriptional coactivators like p300/CBP

Question 23

Which complex works at the IFNB enhancer?

Answer 23

p50/RelA complex
Resulting from varying ability with other transcription factors and conformation adopted when binding IFNB kB element

Question 24

Why are coactivators often required? (3)

Answer 24

Interaction interface (higher affinity than normal activators) Weak interactions Chromatin remodelling Facilitating transcription initiation

Question 25

Explain how NF-kB combines multiple levels of regulation to give transcriptional specificity.

Answer 25

Phosphorylation and degradation of IkB a, B or e Translocation of NF-kB to nucleus Modification of NF-kB subunits DNA binding and gaining access to promoter/enhancer Transactivation - interaction with the basal transcription complex and coactivators Specific transcriptional response

Question 26

How can NF-kB directly induce transcription after chromatin remodelling?

Answer 26

Helping to recruit the basal transcription machinery (BTM)

Question 27

How many members of the NF-kB family form a range of homodimers and heterodimers?

Answer 27

5

Question 28

What targets proteins for degradation by the proteasome?

Answer 28

Ubiquitination

Question 29

Describe the inactive form of NF-kB complex.

Answer 29

Held in cytoplasm bound to inhibitory protein

Question 30

What does phosphorylation of the p100 subunit lead to in the alternative pathway?

Answer 30

Processing to p52

Question 31

Describe the first step of how NF-kB induces a transcriptional response.

Answer 31

Degradation of different IkB proteins, leading to activation of different NF-kB dimers

Question 32

Why is activation of gene transcription compared to a combination lock?

Answer 32

Each number of the combination lock is a different TF, or specifically modified TF or chromatin remodelling factor When all numbers are entered correctly, lock is opened (all TFs recruited to gene promoter/enhancer and transcription occurs)