LT12 NRF2

Question 1

What is the function of NRF2?

Answer 1

Induction of detoxifying enzyme GENES thorugh antioxidant response elements

Question 2

What does NRF2 make a heterodimer with?

Answer 2

Small Maf proteins

Question 3

How does KEAP1 mediate NRF2?

Answer 3

Binds the amino-temrinal Neh2 domain on NRF2 = keeps it in cytoplasm

This represses nuclear activation of antioxidant responsive elements by NRF2

OxS weakens KEAP1 binding on NRF2 so it can move to nucleus and bind antioxidant responsive elements

Question 4

What is NRF2?

Answer 4

Inducible transcription factor

Question 5

Define cytoprotective

Answer 5

Providing protection to cells against harmful substances

Question 6

What degrades NRF2?

Answer 6

Ubiquitin-proteasome pathway

Primarily through the interaction with Keap1 and subsequent ubiquitination by the Cul3 E3 ubiquitin ligase.

Question 7

What is the half life of NRF2 and why?

Answer 7

It has a very short half

So it isn’t active all the time = only responds to stressors

Question 8

What is an inducer of NRF2?

Answer 8

Oxidants and electrophiles

They react with specific cysteine sensors of Keap1 [6], inhibiting its substrate adaptor function, and resulting in Nrf2 accumulation and transcriptional activation of its target genes encoding proteins with diverse cytoprotective functions.

Question 9

What is NQO1?

Answer 9

Antioxidant enzyme

Genes regulated by NRF2

Question 10

How is NRF2 activation monitored in live animals?

Answer 10

Reporter mice

Luciferase

Question 11

What areas does NRF2 regulate?

Answer 11

Redox homeostasis = glutathione & thioredoxin biosynthesis and utilization + NADPH regeneration & utilization

Intermediary Metbolism = carb, lipid, AA and nucleotide metabolism

Question 12

What does NRF2 negatively regulate?

Answer 12

Lipid metabolism

Pyruvate kinase

Glycerol phosphate dehydrogenase 2

Question 13

How is NRF2 regulated post-translationally?

Answer 13

Neh2 regulated by KEAP1

Neh6 = regulated by b-TrCP, which needs to be phosphorylated by GSK-3 first!!!

Both inhibit NRF2

Question 14

What TFs does NRF2 regulate and why?

Answer 14

Regulate TFs with antioxidant response element in their gene promotor

MafG, PPARa and PPARy, RXRa
C/EBPb and AhR

Question 15

What is the role of heterodmier RXRa and PPARa?

Answer 15

Stimuluation of uptake, utilization and catabolism of FA by the LIVER

Question 16

What is the role of heterodmier RXRa and PPARy?

Answer 16

Stimulation of:
FA storage
Glucose metabolism
Adipocyte differentiation

REduction in inflammation

Question 17

What is the role of AhR and ARNT?

Answer 17

Stimuluation of phase I drug metbaolism and differentiation

Inhibits adipogenesis

Question 18

What is the role of C/EBPbeta and alpha?

Answer 18

Contributes to early stages of adipocyte differentiation

Required for macrophage function

Question 19

What affect does NRF2 have on ATP levels?

Answer 19

When NRF2 KO, ATP levels decreased

When KEAP1 KD = NRF2 increased and ATP levels increased

Question 20

How does NRF2 affect ATP synthesis?

Answer 20

Affects relative contribution of oxidative phoshporylation and glycolysis towards ATP synthasis

NO NRF2 = ATP is produced primarily in glycolysis and ATP is used by ATP synthase, which functions in reverse as ATPase to maintain membrane potential

Basically instead of producing ATP = they use ATP to maintain mitochondrial membrane potential

Question 21

What is the function of IAA?

Answer 21

Inhibitor of glycolysis

Question 22

What happens to membrane potential without NRF2 and what else affects this?

Answer 22

No NRF2 means lower potential

This is especially prominent in the absence of glucose

Question 23

What affect does NRF2 have on mitochondrial FA oxidation?

Answer 23

NRF2 increased oxidgen consumption

Without NRF2 = less oxygen consumed

Much more long chain FA oxidized with NRF2

Question 24

How was mitochondrial fusion stimulated and why does this occur?

Answer 24

LPS stimulation

NRF2 is important in inflammatory stress

Mitochondrial fusion = enahnced by NRF2 activation
inhibitbed by NRF2 disruption

Mitochondrial fusion enhances energy production, mtDNA stability, mitochondrial quality control, and cellular resilience, ultimately improving metabolic flexibility and supporting cellular health and longevity.

Question 25

What does NRF2 depletion in A549 cells cause?

Answer 25

Decreases in respiration and ATP production Increases colocalization of mitochondria with lysosomes Increases number of mitolysosomes = suggesting abnormalility in mitoautophagy

Question 26

Is NRF2 up or downregulated in a fasted metabolic state?

Answer 26

NRF2 is activated because need to upregulate glycolysis = may increase energy expenditure NRF2 needs to downregulate FA storage

Question 27

What is the evidence that NRF2 activation may increase energy expenditure?

Answer 27

KEAP1-KD mice gain less weight than WT mice when placed on high-fat diet Indirect calorimetry in KEAP1-KD mice shows a tendency for increased energy expenditure NRF2 activator leads to weight reduction in humans Activator = CDDO (BARD)

Question 28

How does KEAP1-KD cause mice on high-fat diet to gain LESS weight?

Answer 28

Repress haptic expression of gluconeogenic and lipogenic enzymes

Question 29

What is p62?

Answer 29

Autophagy-asaptor protein

Question 30

What is the role of p62 in autophagy?

Answer 30

Phosphorylation of p62 increases its binding affinity for KEAP1 p62 is on Ub cargo for autophagy When KEAP1 is bound to p62 then NRF2 is free to enter nucleus and activate its gene targets

Question 31

How are KEPA1/NRF2 and autophagy linked?

Answer 31

p62

Question 32

How does NRF2 affect cellular glutathione levels?

Answer 32

NRF2 activation = increases gluathione levels (antioxidant)

Question 33

What is the relation between NRF2 and LC3B lipidation?

Answer 33

When NRF2 decreases so does LC2B lipidation

Question 34

What does lipidation of LC3B indicate?

Answer 34

Autophagy is measured by lipidation of LC3B

Question 35

What part of autophgay is not affected by NRF2 depletion?

Answer 35

Formation of new autophagosomes

Question 36

How is nutrient sensing linked to redox stress signalling?

Answer 36

Glycosylation of KEAP1 links the two

Question 37

How does glycosylation of KEAP1 link nutrient sensing and redox stress signalling?

Answer 37

O-GlcNAc transferase (OGT) inhibition activates NRF2 O-GlcNAcylation of KEAP1 is required for NRF2 Ub and degrdation Ser104 O-GlcNAcylation of KEAP1 promotes its productive interaction with CUL3 Ub ligase Glucose deprivation reduces KEAP1 O-GlcNAcylation and activates NRF2

Question 38

What are Sestrins?

Answer 38

Sestrin1 and 2 are produts form two p53 target genes Implicated in defense against ROS

Question 39

What is the affect of Sestrins on NRF2?

Answer 39

Sestrins activate NRF2 by promoting p62-dependent autophagic degradation of KEAP1

Question 40

What is Sestrins mechanism of action in activating NRF2?

Answer 40

Sestring promote autophagic degradation of KEAP1 KEAP1 degradation is dependent on p62

Question 41

What is the protective role of Sesn2 in moues liver?

Answer 41

Sesn2 was upregualted in the liver of mice subjected to 1. Fasting 2. Refeeding with high-carb, no fat diet Without Sesn2 = increased susceptibility of the liver to oxidative damage

Question 42

Why did only refeeding and not fasting promote KEAP1 degradation and NRF2 activation?

Answer 42

Because only refeeding induced p62 expression

Question 43

What is Sulforaphane?

Answer 43

NRF2 activator

Question 44

How is sulforaphane implicated in T2D?

Answer 44

May be a compound that could reverse the disease signature of T2D

Question 45

What is sulforaphanes affect on T2D?

Answer 45

Suppressed glucose production and decreased expression of key enzymes in gluconeogenesis Blood glucose and HbA1c levels reduced with NRF2 activation by sulforaphane

Question 46

What are the most potent NRF2 activators?

Answer 46

Cyclic cyanoenones

Question 47

Give an example of a potent NRF2 activator?

Answer 47

Tricyclic biscyanoenone TBE-31

Question 48

What is TBE-31 mechanism of action?

Answer 48

Acitvates NRF2 by reacting with Cys on KEAP1

Question 49

What diet causes MASLD?

Answer 49

High fat and fructose diet

Question 50

What affect does TBE-31 have on MASLD?

Answer 50

MASLD is associated with oxidative stress so TBE-31 improves MASLD by activating NRF2 Improves insulin sensitivity in HFFr-red mice decreased liver steatosis and expression of lipid synthesis genes, while increasing hepatic expression of fatty acid oxidation and lipoprotein assembly genes. Also, TBE-31 treatment decreased ER stress, expression of inflammation genes, and markers of apoptosis, fibrosis, and oxidative stress in the livers of HFFr-fed wild-type mice

Question 51

What contributes to an increased sensitivity to MASLD when NRF2-KO?

Answer 51

Without NRF2 = no protection against oxidative stress Meaning inflammation occurs

Question 52

What are the potential mechanisms TGF-beta signalling downregulates NRF2?***

Question 53

What are the stategies to reverse MASLD using NRF2?

Answer 53

Inhibiting KEAP1 in early stages of MASLD Inhibiting KEAP1 and beta-TrCP in mid stages of MASLD Inhibiting KEAP1, beta-TrCP and TGF-beta signalling in late stages of MASLD

Question 54

What affect does lithium have on Drosophila?

Answer 54

Lithium is a GSK3 inhibitor This alters lipid metabolism and promotes survival under a high-sugar diet Promotes longevity in Drosophila through NRF2 activation