Epigenetics, diet and the microbiome Flashcards
1
Q
What are the 5 major metabolites that act as co-factors and/or substrates for enzymes involved in epigenetic regulation of gene transcription?
A
- Acetyl-CoA
- NAD
- SAM
- FAD
- Alpha-keto-glutarate
2
Q
What is acetyl-CoA? (2)
A
- Substrate for histone acetyltransferases (HAT)
- End product of glycolysis (anaerobic resp)
3
Q
What are sirtuins? (2)
A
- Class 3 histone deacetylases (HDAC)
- NAD dependent
4
Q
What is NAD? (3)
A
- Derived from vitamin B3 (niacin)
- Essential co-factor for sirtuin function
- Oxidised (NAD+) or reduced (NADH) form
5
Q
What is indicated by high levels of NAD+?
A
Low metabolic activity
6
Q
What is indicated by high levels of acetyl-CoA?
A
High metabolic activity
7
Q
What is FAD? (3)
A
- Derived from vitamin B2 (riboflavin)
- Involved in electron transport in mitochondria
- Essential co-factor for lysine demethylases (KDM)
8
Q
What is SAM? (2)
A
- S-adenosyl-methionine
- Substrate for histone lysine(/arginine) methyltransferases (KMT) and DNA methyltransferases (DNMT)
9
Q
What is indicated by high levels of SAM?
A
High metabolic activity
10
Q
What is indicated by high levels of FAD?
A
Low metabolic activity
11
Q
What is alpha-keto-glutarate? (2)
A
- Intermediate in the TCA cycle
- Essential co-factor for TET demethylases and KDMs
12
Q
What is indicated by high levels of alpha-keto-glutarate?
A
High metabolic activity
13
Q
Where does acetyl-CoA come from? (2)
A
- Made in mitochondria from pyruvate or via fatty acid oxidation
- Acetyl-CoA enters the nucleus to be used by HATs
14
Q
Where does NAD come from? (3)
A
- NAD+ is made from tryptophan
- Converted to NADH in oxidative phosphorylation in the mitochondria
- NAD+ enters the nucleus to be used by sirtuins (HDACs) for gene resetting/repression
15
Q
Where does FAD come from?
A
Riboflavin (vitamin B2) converted to FAD which is recognised by lysine demethylases such as LSD1 (co-factor)
16
Q
What is LSD1? (2)
A
- Lysine specific demethylase 1 (KDM1)
- Specific for H3K4 methylation (trithorax mark, positive regulation)
17
Q
Where does SAM come from? (2)
A
- Made from folate (essential dietary component)
- Substrate for KMTs/DNMTs/PRMTs (arginine methyltransferases)
18
Q
Where does alpha-keto-glutarate come from? (2)
A
- Made from isocitrate in the TCA cycle by IDH1/2
- Co-factor for TET demethylases and KDMs
19
Q
What is IDH?
A
Isocitrate dehydrogenase
20
Q
Which KDMs require alpha-keto-glutarate as a co-factor? (2)
A
- KDMs specific for H3K36me3
- Important in cell proliferation
21
Q
What are agouti mice? (5)
A
- WT A/A agouti mice have a brown sandy coat because have black hairs with a yellow band
- Agouti (A) LoF (a) prevents yellow pigment production
- a/a mice have no yellow band so black coat
- Ay dominant agouti mutation causes agouti overexpression
- Ay/a mice have completely yellow coat
22
Q
What is the pseudo-agouti phenotype?
A
Mice appear like WT agouti mice but genotype is Avy/a
22
Q
What is the agouti viable yellow (Avy) allele? (3)
A
- Mutation that causes variegated coat pigmentation in mice
- Avy/a heterozygotes show a range of different coat colours from yellow to agouti, many have a mixture of the 2 in a mottled pattern
- Decision is made early in development as to how much agouti protein is being expressed by Agouti viable yellow
23
Q
What causes the Avy mutation? (3)
A
- Insertion of an IAP element (transposon) into the regulatory region of the Agouti gene which causes strong activation of the gene
- IAP sensitive to DNA methylation which causes silencing of IAP and therefore weaker expression of Agouti
- Variable methylation of IAP in skin pigment cells so Avy allele shows variegating expression in different cells of genetically identical littermates meaning some hairs are yellow and some are brown (pseudo-agouti)
24
What is the impact of methyl donor (folate, vitamin B12, choline, betaine) supplementation of maternal diet during gestation of Avy/a offspring? (3)
- Shifts the coat colour distribution of Avy/a offspring towards the brown pseudo-agouti phenotype
- Increased methyl donors in the diet used by methylation machinery to methylate IAP and repress the activation of Agouti
- Demonstrates that methyl donors from the diet regulate DNA methylation and gene expression during development
25
What is the impact on seasonal variation in folate on DNA methylation in Gambia? (2)
- Dietary folate availability is high in the rainy season and low in the dry season
- Offspring conceived in the rainy season have higher DNA methylation of specific genes and overall than conceived in the dry season
26
How is PGC1alpha regulated? (2)
- Acetyl-CoA is used to acetylate PGC1alpha by GCN5/KAT2A which inactivates its DNA binding activity
- Acetylation removed by SIRT1 (NAD+ dependent deacetylase) to activate PGC1alpha when when low carbohydrate supply to use fatty acid oxidation for energy instead
27
What is the function of PGC1alpha? (3)
- Transcription factor that promotes mitochondrial biogenesis (for aerobic respiration) and fatty acid oxidation (when low carb supply) which are required in conditions of energy depletion
- Activated by nutrient deficiency
- Regulated by NAD+ and acetyl-CoA
28
How is PGC1alpha affected by fasting? (2)
- Fasted cells (energy depletion) contain PGC1alpha in a complex with SIRT1 so that PGC1alpha can be deacetylated and drive mitochondrial biogenesis/fatty acid oxidation
- Complex disappears when cells are refed and PGC1alpha is acetylated
29
What is the function of SIRT1? (2)
- Deacetylase which promotes PGC1alpha-mediated function of mitochondrial biogenesis and fatty acid oxidation
- Target gene expression is reduced by siRNA interference of SIRT1
30
What are the features of the gut microbiome? (4)
- Contains around 500-1000 species of bacteria
- Uses gut mucus as an attachment matrix which allows bacteria to form biofilms
- Different parts of the gut have different bacteria which carry out different steps of food breakdown and metabolite supply
- Human microbiome is acquired by embryo from the mother
31
How do you create germ-free mice? (3)
- Delivered via aseptic caesarean section
- Maintained in a germ-free environment
- Fed with germ-free food
32
What is seen in germ-free mice? (2)
- Wide range of abnormal physiological and behavioural phenotypes
- Microbiome is adaptive
33
What are examples of the developmental impacts of germ-free mice? (3)
- Reduced expression of immediate-early synaptic activity regulated gene NGF1-A in germ-free mice
- Blood capillary development impaired in germ-free mice
- Gut-associated-lymphoid-tissue (GALT) missing in germ-free mice
34
What is produced by the microbiome? (2)
- Short chain fatty acids: acetate, propionate, butyrate
- SAM precursors: folate, B vitamins
35
What is the function of acetate?
Precursor of acetyl-CoA
36
What is the function of butyrate? (5)
- Histone deacetylase (HDAC) inhibitor
- Actively imported into cells via monocarboxylate transporters and GPCRs
- Activates LSD1
- Supplies acetate for acetyl-CoA/ATP production
- Regulates PGC1alpha
37
What is GCN5/KAT2A?
Acetyl-CoA-dependent lysine acetyltransferase which catalyses acetylation of PGC1alpha
38
What is the methyl donor?
SAM
39
What is the acetyl donor?
Acetyl-CoA
40
How is the Krebs cycle linked to gene expression? (5)
- Input is acetyl-CoA made either from glucose or fatty acids
- Involves conversion of NAD+ to NADH and FAD to FADH2
- NAD+ is depleted in high metabolic activity, NADH rises which enters the electron transport chain for oxidative phosphorylation and ATP production
- NAD+ is high when energy is low which is the co-factor for SIRT1 which allows activation of PGC1alpha
- Lots of acetyl-CoA = lots of Krebs cycle = lots of ATP production
