Folate And Methylation - (Mar/31/2005 )

Kestutis,

I think what you are doing is a very noble effort in theoretical science and I commend you in doing it. Your views and ideas have certainly got me thinking "outside the square" and without such people, I think bench scientists will always think narrowly (this maybe an overgeneralisation).

I am a bit unclear with your diagram, I think you are trying to demonstrate all pathways that lead to DNA methylation? I should check up on all the biochemical pathways for this as from memory, it looks like what you are drawing.

On another note, which country are you from?

Nick

Hi Nick,
Thanks for support.
I remember the beginning of my research. Lots of numbers about the amount of B12 forms in a blood of stomach cancer sufferring and ulcer having. I was lucky to constructs the rule for differential dyagnosis, but I asked professor - why in lots of cases such a high amount of methylcobalamine when cancer. She smiled and gave me the heap of books and articles. I've tryied gave theorethical base for experimental data. Now, i'm finnishing with the general theory of malignant growth (everyone nows that B12 controls growth strongly...)-starting from initiation (part of this is real epigenetics) ending with ideas how and in which points to affect cancer cell. Some days ago I've asked myself -what does it mean to solve problem of cancer. Possible answer this, means - to be able controll this phenomena. But before control we need understand. So, lots of experiments needed - among them - folate system disturbances and Histones, we discuss. Experiments in vitro and vivo takes lots of time, so some brains started computer simulation, writting math equations for biochemical systems. This gives possibility to evaluate consequancies and dynamic starting from some beginning state of system...One example - folate system simulations made in North Carolina and Seattle, USA, published 2004 October, I've mentioned this some days before. Nice job, but problems in connection with reality and surrounding area.
I use more simple logic - this step of analysis is necessary before computer simulation. If i study methylation - I must evaluate all the inputs of carbon used for this (if one input in experimental system is not stable - different results then obtained). Very simplified model I'm drawing I hope will be usefull for better understanding of the problems (I think about byologists and oncologists first; biochemists could present much more complicated ...) - when we speak about this , we have some image of this in our imaginations.
Met, Ser...Gly, Chol...Bet, His, Trp and carriers of OCU - Folate, Met cycle is not too complicated to take into account at simplified level. The problem is - how to present this for better understanding . I have some ideas -trying to realise.
But yeasterday morning I draw one - our computer is on LINUX, not Windows - the new one for me, and this occuared in .sdx format but for publishing in protocol we need .gif. Life is life.I've found one more program - GIMP - studied drawing and yeasterday evening understood pencil and brush using finally (this like Adobe systems) - made some drawings - but they didn't satisfied me - not enough estethicall and usefull. Now I'm finnishing drawings.
Folate system is still under consideration - seems from PubMed. Understanding about folate antigonist MTX was found decades ago - to slow DNA nucleotide. This was impressive, but only with some effect on cancer. Why?
If folate goes down, Met cycle goes down too, not enough of SAM and produced DNA in cancer cell more and more hypomethylated globally (very nice ideas about stochastics of methylation I've found in Sue Clarc articles recently - and was happy becouse stressed this theoretically fourteen years ago making report at seminar - Kiev or St.Petersburg - thanks for experimentators). Then hypermethylation of some DNA region appears and changes in Histone methylation. To conclude - MTX slows nucleotides synthesys, but does bad job too - in DNA methylation-Histone direction maybe. So, MTX plus SAM, plus some Histone demethylating drug pluc acethylation controling drug mixture theoretically has the chance to be much more efective than MTX alone in some cancer treatment. This is, why such drawings we need. I've seen lots of drawings dealing with some part of methylation system (very nice about folate, about choline-Bet-Gly-Ser, about Ser-Gly, or Met cycle, but never more full image of OCU I'm going to finish today (for discussions of course too), now, because I can't found any even using google-image search system. Discussions become more effective and clear, I hope.

Kestutis Urba
P.S. I was born in Lithuania. This is middle, when someone drives from Germany to Finland. Mentioned in crusaders chronics middlages - 1009. The biggest country in Europe for some time. Very ancient and conservative language - some suppose not far from indoeuropean origin ( among the oldiest words in the world is milk, may be origin 20 000 -40 000 BC, we have some like this 'melzhti"... ) - Lithuania is known for basketball - bronze medals at olympic games, European champions and NBA star Sabonis. National heroes -Darius and Girenas crossed Atlantic in 1933 from USA to Baltic sea coast. Where are You from?

I am from Australia,

Lithuania is a great country, I would love to go visit someday.

Here is a summary of folate biosynthesis how we understand it (see attachment) from your comments Kestutis, serine and OCU transfer will enter the SAM/Adomet stage.

N

Thanks Nick,
This is OCU metabolism - the main features and pathes, before carbon atom appearance on DNA (in attachment).
The Ser-Gly path is essential in kidney, choline-Gly - in liver.... I'm going to improve this one- problems I'm still not skilfull drawer using GIMPS. Green for folate I've choosed - because the source - green vegetables; Histydine - not only in red wine; Trp -choclate (I'll change into brown)...Met - maybe fish, cheese - I'm trying to remember...The usage of corresponding colours in the drawing makes this one psychologically better acceptable and informative. Black colour from pencil tool seems like real carbon...
When I study Your drawing more deep -I'll discuss this one - of course such representing of folate input has advantages, but some more details may be must be considered too.
Kestutis Urba

Hi mtNick,

I would like to propose:
1.some general ideas about biochemical “maps” constructing.
2.some special remarks about “Folate deficiency and Histones methylation” experiment presentation by map. Then, about the question – how to obtain the biggest amount of information when studying this problem.
So, when constructing the OCU metabolism map, I presented systems of glycolysis, folate metabolism and Met cycle using displacement published in well known Roche- ExPASy “Biochemical pathways-Metabolic pathways map”: glycolysis above folate system, Chol-Bet-Gly path down and right side, because ....“In Roche we trust”...
When comparing pathways You study and my map, first, I need “a mirror” to fit. This small problem of information presentation acception made studying little problematic, because during years, may be for 1000... times, I've tryed to calculate scenaries generated by different events on OCU pathways something in the way Roche presented.
I propose in the future for all members of our “DNA methylation...” forum accept Roche compartamentalisation of metabolism as basic or the other one if there exist better map, and follow it's order . So, if joking, we need some agreemant and tritty Vilnius-Australia (Canberra...) in the beginning. I hope one day lots of the biochemists will decide to do the same (at some important meeting) with some exeptions, of course, in special situations ( there are some other global maps of biochemical pathways). Understanding of other ideas could be better – faster...What is Your opinion? About colors usage now. I've stressed – folate system let be green, because of natural origin. What to do with others? Met, His, Trp, Gly, Ser colours I'm going to choose according the food origin too. If possible, (computer is great tool in this situation) better to use.
2. Now, about Your map. I like it – simple, clear, easy understand the problem You study.
But when You write the article about obtained results, again, for better uderstanding by other experimentators, it is possible to stress, that Your map represents only the important part of total OCU system publishing some full OCU metabolism “map” (better than I proposed) and discussing some more interesting tasks ( for example, what could be expected if Chol input was higher to compensate Folate deficience an so on). We need such “politics”, because there was not yet done systematic analysis of OCU-methylation system, which seems to be more and more important and what we are trying to do in this forum part theoretically. Of course, in the begining, there must be studied single problems, but imagine the experiment –Liver cell, Folate deficiency, Chol input normal, Ser catabolism damaged but some SAM added without increment of dietary Met! If we change all situations at the input of the system point and study consequancies of it, we can try to plan adaptive control of OCU and methylation depending on the enzyme state. Powerfull coordinated lab concorcium is needed.
The PROBLEMS, I see. How to catch more, if not all the reasons of “methyl trap”? First, in some cases possible “methyl trap” depending on situation outside Your map, not depending only B12 (damaged some folate enzyme not presented....). It's interesting – DNA hypomethylated... because of this trap.
Second, about the regulation of methyl cycle. It was published, that the main regulator substrate of Met cycle is S-adenosyl-l-homocysteine (sHmc). It was found that sHMC facilitates the output from Met cycle activating cystathionine-beta-synthetase at all concentrations of Ser, Hmc (output from Met cycle), meanwhile repressing enzymes (input to Met cycle) betaine-homocisteine methyltransferase and 5-methyl THF-Hmc-methyl-transferase. This means – the more sHMC produced, the more intensive output from Met cycle appears but input appears under repression. Nice regulation. If folate is going down, amount of sHmc could be expected less, this intensifies input of OCU from folate...Hm..(this means, I think, about Hmc). Important circumstance, maybe, possible to add into the map, because that one seems to be exceptional important not only in this experiment. (To be continued.)

1.Kestutis Urba
P.S. Thanks for commend – my possitions sthrenghthened 20%.

Dear Kestutis,

I have a similar biochemical pathways map you cited from Roche and this is where I had adapted my map along with several references mainly from the Fenech group in Adelaide, Australia.

All chemical reactions are in equilibrium, this we know, however the reaction will favour either the reactants or the products so you have either more reactants of more products in the reaction at equilibrium, the enzymes are catalysts and push the reaction more to favour one or the other.

In this case it is all the transferases, most biochemical reactions are regulated by feedback, as you rightly point out accumulation of one particular substrate could increase the activity of the enzyme that metabolises it to try and compensate for the accumulation. As there are many steps involved in getting folic acid converted into a OCU that is then transferred to DNA/Histones and there are many pathways that feed into it, it is still unclear what is involved. Biology is a reductionist science and we like to simplify things as much as possible to test a hypothesis. I am now fully aware of the potential of other pathways feeding into this and will take all these into account.

I hope if other people are reading this forum and have some ideas they would like to present, to do so as well!

all the best.

Nick

Dear Nick,

I would like, when continuing discussion, to stress the importance of s-HMC consideration and presentation in such experiments and I would like to explain why? Simplifcation of complex problem sometimes leads to the simplification of truth (If some very important circumstance was lost)! Sorry, I'm afraid that there is such a situation and I'll try to describe some hipothetical model - how deficience of folate works.
It is accepted point of view that
Dietary folate deficiency or genetic polymorphisms in folate-metabolizing enzymes are associated with megaloblastic anemia, developmental abnormalities including neural tube defects and Down's syndrome, and various types of cancer, especially those of the gastrointestinal tract and leukemias.
Normal daily fluctuations of folate intake do not affect cellular folate levels very much, since, on the average, only 0.8% of total folate is consumed and excreted each day (see above). Long term folate deficiency that reduces total folate by one-half reduces the velocities of most of the reactions by approximately one-half. Thus, the basic folate model predicts that folate function should proportionally decrease with total folate, and therefore one should expect to see functional consequences such as hyperhomocysteinemia and megaloblastic anemia within months. However, long term folate deprivation studies in humans showed a rapid decline in serum folate followed by "weeks or months when the serum folate concentrations is low but there is no other evidence of folate deficiency" (65).
Institute of Medicine (1998) Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin, and Choline, p. 203, National Academy Press, Washington, D. C. ]
So, everyone would like to know WHY THE SERUM CONCENTRATION IS LOW BUT THERE IS NO OTHER EVIDENCE OF FOLATE DEFICIENCY WITHIN MONTHS?
Hypothesis. There exists some strong compensatory mechanisms in the cell obtained in evolution - only such humans ( when eating "the green food") survived. One of compensatory possibilities is based exactly on s-HMC regulatory mechanism which controls input into Met cycle corresponding to the output. Possibly, when defficience in folate appears this mechanism supports resynthesis of Met but not Hmc leackage from Met cycle, till some critical level of folate was reached. If the human has such system not working efficient enough for longer time - SAM concentration falls down and DNA hypomethylation... appears.
The other compensation is folate storage when binded to some protein.
Well, when someone plans and makes experiment in vivo he could obtain randomly some different result depending on effectivness of this sHmc compensatory mechanism - I think , in the most cases this works, but in some (very interesting)- not efficient enough. So, sHMC presentation seems to be usefull too when studying such systems.
Perhaps someone will study Folate, sHMC state and Histone methylation in the future. It is very interesting too.
Pregnancy and neural tube deffect are more special situations. Folate defficiency compensatory system appears at more complicated situation ....I must study this more deep.

Kestutis Urba
P.S. Of course, I explain some details in forum posts to make discussion better understandable! Some biochemists are more biologists, some - more chemists. You are strong biologist - I see reading the other coments and strong chemist trying to explain and solve the most interesting questions. I hope, the other scientists accept this, that we realy need to describe glycolysis above folate system, Ser-Gly - ...right side...following Roche. Otherwise we are in Alice among mirrors and scissors situation!

Hi Nick,

One more problem. Met cycle is controled not only by B12 but by B6 too. I suppose, 70% of readers -knows this very well - better than me, but 30%(students etc.) doesn't. So, I'm going to put B6 into my OCU metabolism map, too. What about Yours? For this forum, I think, better to stress this detail too. Oh, these simplifications.. !;)
PUBMED search results: cobalamin(B12), methylation[i] - 345 articles and B6(pyridoxal phosphate), methylation - 62 articles.

Kestutis Urba

In my case, of generalisation,

I know that B-vitamins together are required within the folate cycle, and of course I would expect there would be other micronutrients that are required as co-factors to enzymes within the OCU metabolism cycle. This is known. However, I think it is folate that is not synthesised or can be derived from the body and needs to be absorbed through our foods.

Nick

Hi Nick,
The last letters were more about scientific psychology - how to present information, but less about science. Generalisation has it's rules too. I'm going to study this using formal system research, system identification science.
But, first I have some important questions about methylation and neural tube deffects (NTD), because I' have some understanding about cancer cell and unfortunately very little about NTD. NTD is very rare when comparing with a cancer, but I see importance of this problem: if we obtain understanding about embryo cell and total embryo development program we obtain understanding - what switches on and out processes of growth. This seems to be of great importance in cancer research.
I've read - molecular mechanism of NTD reason is still unknown.
I'm trying to remember - totaly hypomethylated human embryo cell appears immediately and some methylation pattern appears aproximately on the 8 day of its existance. NTD features appears aproximately on 28th day and folate defficiency in serum supposed to be the reason of NTD. Folate deficiency (usually appears when not enogh in food) and DNR hypomethylation relation is complicated enough, but even theoretically is possible to make some conclusions. Usually folate system is described as nucleotides synthesys controling system. If nucleotides synthesys is slower much more, than normal, then...? Could it be the reason and the only reason of NTD? I've seen some ideas about folate dependent growth factor role, but what is the mechanism of dependence? The other possibility is: If something bad with methylation pattern creation (not enough of SAM) on the 8th day, some genes (and histones) remains hypomethylated too (this depends on the chance too), they are not swiched out - the normal program of development breaks...Is it possible? Are these questions real questions or they have no meaning for NTD research?

Kestutis Urba
in Vilnius warmer....