LT12 NRF2 Flashcards
What is the function of NRF2?
Induction of detoxifying enzyme GENES thorugh antioxidant response elements
What does NRF2 make a heterodimer with?
Small Maf proteins
How does KEAP1 mediate NRF2?
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
What is NRF2?
Inducible transcription factor
Define cytoprotective
Providing protection to cells against harmful substances
What degrades NRF2?
Ubiquitin-proteasome pathway
Primarily through the interaction with Keap1 and subsequent ubiquitination by the Cul3 E3 ubiquitin ligase.
What is the half life of NRF2 and why?
It has a very short half
So it isn’t active all the time = only responds to stressors
What is an inducer of NRF2?
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.
What is NQO1?
Antioxidant enzyme
Genes regulated by NRF2
How is NRF2 activation monitored in live animals?
Reporter mice
Luciferase
What areas does NRF2 regulate?
Redox homeostasis = glutathione & thioredoxin biosynthesis and utilization + NADPH regeneration & utilization
Intermediary Metbolism = carb, lipid, AA and nucleotide metabolism
What does NRF2 negatively regulate?
Lipid metabolism
Pyruvate kinase
Glycerol phosphate dehydrogenase 2
How is NRF2 regulated post-translationally?
Neh2 regulated by KEAP1
Neh6 = regulated by b-TrCP, which needs to be phosphorylated by GSK-3 first!!!
Both inhibit NRF2
What TFs does NRF2 regulate and why?
Regulate TFs with antioxidant response element in their gene promotor
MafG, PPARa and PPARy, RXRa
C/EBPb and AhR
What is the role of heterodmier RXRa and PPARa?
Stimuluation of uptake, utilization and catabolism of FA by the LIVER
What is the role of heterodmier RXRa and PPARy?
Stimulation of:
FA storage
Glucose metabolism
Adipocyte differentiation
REduction in inflammation
What is the role of AhR and ARNT?
Stimuluation of phase I drug metbaolism and differentiation
Inhibits adipogenesis
What is the role of C/EBPbeta and alpha?
Contributes to early stages of adipocyte differentiation
Required for macrophage function
What affect does NRF2 have on ATP levels?
When NRF2 KO, ATP levels decreased
When KEAP1 KD = NRF2 increased and ATP levels increased
How does NRF2 affect ATP synthesis?
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
What is the function of IAA?
Inhibitor of glycolysis
What happens to membrane potential without NRF2 and what else affects this?
No NRF2 means lower potential
This is especially prominent in the absence of glucose
What affect does NRF2 have on mitochondrial FA oxidation?
NRF2 increased oxidgen consumption
Without NRF2 = less oxygen consumed
Much more long chain FA oxidized with NRF2
How was mitochondrial fusion stimulated and why does this occur?
LPS stimulation
NRF2 is important in inflammatory stress
Mitochondrial fusion = enahnced by NRF2 activation
inhibitbed by NRF2 disruption
What does NRF2 depletion in A549 cells cause?
Decreases in respiration and ATP production
Increases colocalization of mitochondria with lysosomes
Increases number of mitolysosomes = suggesting abnormalility in mitoautophagy
Is NRF2 up or downregulated in a fasted metabolic state?
NRF2 is activated because need to upregulate glycolysis = may increase energy expenditure
NRF2 needs to downregulate FA storage
What is the evidence that NRF2 activation may increase energy expenditure?
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)
How does KEAP1-KD cause mice on high-fat diet to gain LESS weight?
Repress haptic expression of gluconeogenic and lipogenic enzymes
What is p62?
Autophagy-asaptor protein
What is the role of p62 in autophagy?
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
How are KEPA1/NRF2 and autophagy linked?
p62
How does NRF2 affect cellular glutathione levels?
NRF2 activation = increases gluathione levels (antioxidant)
What is the relation between NRF2 and LC3B lipidation?
When NRF2 decreases so does LC2B lipidation
What does lipidation of LC3B indicate?
Autophagy is measured by lipidation of LC3B
What part of autophgay is not affected by NRF2 depletion?
Formation of new autophagosomes
How is nutrient sensing linked to redox stress signalling?
Glycosylation of KEAP1 links the two
How does glycosylation of KEAP1 link nutrient sensing and redox stress signalling?
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
What are Sestrins?
Sestrin1 and 2 are produts form two p53 target genes
Implicated in defense against ROS
What is the affect of Sestrins on NRF2?
Sestrins activate NRF2 by promoting p62-dependent autophagic degradation of KEAP1
What is Sestrins mechanism of action in activating NRF2?
Sestring promote autophagic degradation of KEAP1
KEAP1 degradation is dependent on p62
What is the protective role of Sesn2 in moues liver?
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
Why did only refeeding and not fasting promote KEAP1 degradation and NRF2 activation?
Because only refeeding induced p62 expression
What is Sulforaphane?
NRF2 activator
How is sulforaphane implicated in T2D?
May be a compound that could reverse the disease signature of T2D
What is sulforaphanes affect on T2D?
Suppressed glucose production and decreased expression of key enzymes in gluconeogenesis
Blood glucose and HbA1c levels reduced with NRF2 activation by sulforaphane
What are the most potent NRF2 activators?
Cyclic cyanoenones
Give an example of a potent NRF2 activator?
Tricyclic biscyanoenone
TBE-31
What is TBE-31 mechanism of action?
Acitvates NRF2 by reacting with Cys on KEAP1
What diet causes MASLD?
High fat and fructose diet
What affect does TBE-31 have on MASLD?
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
What contributes to an increased sensitivity to MASLD when NRF2-KO?
Without NRF2 = no protection against oxidative stress
Meaning inflammation occurs
What are the potential mechanisms TGF-beta signalling downregulates NRF2?***
What are the stategies to reverse MASLD using NRF2?
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
What affect does lithium have on Drosophila?
Lithium is a GSK3 inhibitor
This alters lipid metabolism and promotes survival under a high-sugar diet
Promotes longevity in Drosophila through NRF2 activation
