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Vitamin B9

VitB9

Folate, Tetrahydrofolate(THF), 5,10-methylene THF, 10-formyl THF, 5-formimino THF, 5,10-methylenyl THF, 5-methyl THF

VitB9

Daily Requirement:

Modified DV:

RDA ?:

Adequate Intake ?:

400

true

DFE/d

DFE/d

Min Deficiency:

Max Toxicity:

Tolerable UL

Animal:Plant Conv:

1000

DFE/d

DFE/d

DFE/d

Date Discovered:

1941

Short Description:

One-carbon transfer reactions for nutrient metabolism and energy production; gene expression; purien and pyrimidine synthesis for DNA and RNA; hematopoiesis. Major Food Sources: Green vegetables = spinach, asparagus, and greens; legumes; fortified grain products. RDA: 400 mcg

Interpretation:

Main sources:


Animal Products: Liver, dairy products, egg yolks


Plant Products: Oranges, Dark leafy vegetables, peanuts & beans


Dietary methyl donors and DNA methylation

• What nutrients are considered methyl donors

Single carbon metabolation pathway, folate, zn, b6, b12, methionine, choline

• Where does DNA methylation normally occur

CpG islands, promoters of genes. folate deficiency has - dutch hunger study - starvation causes problems later in life, first trimester - igf2 dna methylation lack. folate deficiency uracil misincorporation. If folate deficiency, uracil can't turn into thymine. Causes strand breaks, two uracil residues = double strand breaks - bad.

• What is the significance of the agouti mouse experiments

Dietary exposure of your grandma can effect your genotype. Your egg was exposed to that that diet.

• Why are dietary methyl donors important in cancer, what are the mechanisms

If low methyl donors, aberrant methylation, chance for uracil -> cancer

History & Discovery:

1941



Digestion:

Jejunum is the main site of digestion and absorption.


Food folates with many glutamine (3 or 7) need to be broken down by proton-coupled folate transporter PCFT in the membrane of the enterocyte. Then the food folate with one Glu is brought into the enterocyte as either THF or 5,10-methylene-THF, which goes to 5-methyl-THF for transport to the Plasma through multidrug resistant-associated protein (MRP).




Stored in small quantities in liver, kidneys, and spleen for 3-4 months.


Uptake into tissues requires proton-coupled folate transporter (PCFT) or organic anion transporting polypeptides (OATP) B1 and B3.


5-methyl-THF is the main form in blood plasma.

Folate is found as free (33%) or bound to albumin and alpha2-macroglobin (66%)

Absorption and Storage:

AI of folate = 320 mcg/day.


Folic acid from supplements/fortified foods is more bioavailable. This is taken into account for Dietary Folate Equivalents (DFE).


1 DFE = 1 mcg of food folate = 0.5 mcg folic acid from supplement on empty stomach


1 DFE = 1 mcg of food folate = 0.6 mcg folic acid from fortified food (1.7 factor)


Example:


Half cup of avocodo = 60 mcg of folate

Slice of bread = 50 mcg of folic acid * 1.7 = 85 mcg


Folate is degraded by heat, oxidation and UV light.

Overall absorption is 10-90% or 50% on average due to food matrix and inhibitors.


Folate bioavailability of beef liver, lima beans, peas, spinach, mushrooms, collards, orange juice and wheat germ was estimated with a protocol of folate depletion-repletion using growth and liver, serum and erythrocyte folate of weanling male rats. Diets with 125, 250 and 375 micrograms folic acid/kg were standards. Individual foods were incorporated into a folate-free amino acid-based diet alone (250 micrograms folate/kg diet from food) or mixed with folic acid (125 micrograms folate from food + 125 micrograms folic acid) to evaluate folate bioavailability and effects of food matrix. Beef liver and orange juice folates were as available as folic acid, whereas those of wheat germ were less bioavailable. Folates of peas and spinach were also less available than folic acid using liver and serum folate concentrations and total liver folate as response criteria, but they were not lower when based on growth and erythrocyte folate concentrations. Lima bean, mushroom and collard folates were as available as folic acid using four of five response criteria. Folate bioavailability of all foods generally exceeded 70%. All response criteria gave approximately equivalent results, indicating that growth and tissue folate levels are appropriate criteria. No food matrix effects were observed for any food except lima beans. Foods rich in polyglutamyl folates were less bioavailable than those of foods rich in short-chain folates. https://pubmed.ncbi.nlm.nih.gov/2007897/


Folic acid, 5-formyl THF and 5-methyl THF absorbed intact.


Folate in food needs to be converted to tetrahydrofolate (monoglutamate form) by folate hydrolase, which requires Zn.


Jejunum is the main site of digestion and absorption.

Important Pathways:

Folate Appearance


Folate Forms


Folate = tetrahydrofolate (food form) + folic acid (oxidized version, found in supplement form)

Humans can synthesize each individual part but we cannot link pteridine to PABA

One glutamic acid on figure, but typically attached to 3 or 7 glutamic acids in the body and in food which is called a 'glu tail'. 75% of folate exists in this form.


Function of Folate:

One carbon transfer or methyl donor. "Dumping" a methyl on B12 (CH3), which can then be dumped onto homocysteine to be turned into S-adenosyl methionine = central methyl donor which is important for anabolic reactions.



DNA Synthesis

5,10-methyleneTHF reacts with dUMP to create dTMP which can be used for DNA synthesis. Or it can become 5-methylTHF to react with homocysteine to become methionine which can enter the SAM cycle to lead to DNA methylation.

Reactions between Forms of Folate

SAM Cycle

Homocysteine -> Cystathione -> Leads to succinyl CoA, glucose, glycine, Heme


Deficiency Diseases, Detection, Cures:

Red blood cells normally divide, yet this doesn't happen with folate or B12 deficiency.


Deficiency disease:

  • Common in early 1900's in young. female workers in former British colonies.

  • Tropical megaloblastic anemia - hyperpigmentation of fingers before treatment of anaemia (A)

  • Reduction 3 months after treatment (B)

  • Peripheral blood film shows hypersegmented neutrophils (green arrowheads), macrocytes (blue arrowheads), and a teardrop shaped red blood cell (red arrowhead). (C)


Erythropoiesis: maturation of red blood cells


Hematopoietic stem cell -> Proerythroblast -> Megaloblastic red blood cell due to folate deficiency -> Erythroblasts - two cytosols but no nuclei division (requires carbon donor folate) -> Large immature red blood cells -> Megaloblastic anemia - fewer red blood cells to take up iron -> heme iron deficiency



Recommendation: Take 400 mcg DFE/day 1-3 months before pregnancy and first month to help with embryonic neural tube closure to prevent anencephaly.



Symptoms of Deficiency:

  • cardiomegaly, tachycardia, shortness of breath

  • megaloblastic anemia

Genetic Diseases:

Dietary methyl donors and DNA methylation

• What nutrients are considered methyl donors?

• Where does DNA methylation normally occur?

• What is the significance of the agouti mouse experiments?

• Why are dietary methyl donors important in cancer, what are the mechanisms?


  • Higher eukaryotes use epigenetic modifications to reversibly suppress transcription of genes and repeat elements, often by stable silencing

  • Epigenetic marks are retained through mitosis, allowing maintenance of characteristic cell types

  • By their nature, epigenetic modifications are susceptible to environmental influence: the interposition of epigenetic modifications between genes and the environment provides a way in which the environment can exert heritable influences on phenotype



Methylation of DNA

  • Promoter regions of many genes are rich in CpG dinucleotides

  • Carbon 5 of cytosine in CpG dinucleotides is a prime target for methylation by DNA methyltransferases (DNMT)

  • Depending on cell type or tissue, 3–4% of all cytosines in vertebrate DNA and 70% of cytosines in CpG dinucleotides are 5-methylated

  • Methylation of cytosines is a heritable modification of DNA. However, the following mechanisms are available to remove methylation marks in vivo:

  • enzyme-mediated removal of methyl groups

  • passive removal of methyl groups if methylation is not maintained at the time of replication

  • DNMT utilize S-adenosylmethionine (SAM) as a methyl donor

  • Folate plays a unique role in the generation of SAM, methylentetrahydrofolate reductase catalyzes the conversion of 5,10-methylenetetrahydrofolate to 5- methyltetrahydrofolate

  • The latter donates a methyl group to convert homocysteine to methionine

  • Methylenetetrahydrofolate reductase deficiency results in hypomethylation of DNA, consistent with a role for folate in DNA methylation

  • Nutrients other than folate also play roles in one-carbon metabolism: choline and methionine are the major dietary sources of methyl groups

  • whereas vitamins B-6 and B-12, riboflavin and zinc are coenzymes and cofactors, respectively, for various steps in one-carbon metabolism

  • Methylation of CpG islands is associated with transcriptional repression and provides a mean to control gene expression

  • DNA methylation plays a role in allele-specific gene expression (genomic imprinting), heritable transcriptional silencing of parasitic sequence elements and X-chromosome inactivation

  • Impaired DNA methylation is associated with perinatal death, decreased fertility, abnormal fetal development and tumorigenesis


Histone and DNA Methylation


Effect of Methyl Donors on the Epigenetic Process

  • Several observations point to the ability of several environmental factors, including diet, to be key regulators of epigenetic processes, and evidence exists for diet effects on both DNA methylation and histone posttranslational modulation

  • Diet can influence the degree of methylation by influencing the availability of methyl donors, including folate, choline, and methionine

  • Several dietary factors ranging from alcohol to zinc have been reported to influence the supply of methyl groups for the formation of SAM and thereby affect the methylation of CpGs

  • Dietary methyl deficiency (of folate, choline, and methionine) in a rat model is known to alter hepatic DNA methylation patterns and induce hepatocarcinogenesis in the absence of a carcinogen

  • Dietary components such as EGCG found in green tea can influence gene methylation

Gene Imprinting and Diet

  • Genomic imprinting is an epigenetic modification that inactivates one allele of a gene in a parent-of-origin-dependent manner. It occurs primarily by allelic-specific methylation of cytosines in CpG dinucleotides during gametogenesis

  • Although relatively few in number, imprinted genes are thought to be vital to human development, and their dysregulation appears to affect the risk of developing some diseases

  • One example by which early dietary exposures may affect genomic imprinting involves the imprinted gene insulin-like growth factor 2 (IGF2)

  • The maternally inherited allele of the imprinted gene encoding IGF-2 is normally epigenetically silenced, resulting in expression almost exclusively from the paternal allele

  • Interestingly, loss of imprinting at the IGF2 locus has been shown to induce biallelic expression of this growth factor in 10% of normal human adults and is implicated in several types of cancer

  • For the most part, the cause of the IGF2 loss of imprinting is unknown

  • Mice weaned on either a synthetic control or a synthetic methyl donorcofactor deficient diet compared with a standard diet displayed hypomethylation of IGF2 and consequent dysregulation of IGF2 allelic expression

  • The loss of imprinting caused by the deficient diet persisted during a subsequent 100-d recuperation period after the mice were switched to the control diet

  • Thus, early nutritional influences may stimulate changes in cytosine methylation to which imprinted genes, such as IGF2, may be susceptible

  • These findings in view of the relation between IGF-2, insulin, and metabolic syndrome suggest that early nutrition may influence susceptibility to adult obesity, diabetes, cardiovascular disease, and cancer

Dutch Hunger Winter Study - Heijmans B. T. et.al. PNAS 2008;105:17046-17049

Difference in IGF2 DMR methylation between individuals prenatally exposed to famine and their same-sex sibling. Periconceptional exposure: Difference in methylation according to the mother's last menstrual period (a common estimate of conception) before conception of the famine-exposed individual.