FAM13A, fatty acid oxidation and ME

Lipid metabolism is dysregulated in ME patients (1, 2, 3, 4).

Family with sequence similarity 13 number A (FAM13A) regulates fatty acid oxidation (FAO) (5).

The gene FAM13A is hypomethylated (TSS200 and TSS1500) in peripheral blood mononuclear cells (PBMC) from ME patients (table S4 in ref 6).

The gene FAM13A is hypermethylated (1stExon/5'UTR), CPT1A is hypermethylated (5'UTR) and CPT1B is hypermethylated (TSS200/5'UTR) in PBMC from ME patients. This DNA methylation pattern is related to quality of life in the ME patients (table S7 in ref 7).

In whole blood from adolescent CFS patients FAM13A gene expression is elevated (table S3 in ref 8).

Use the link to see the figure:
Figure. FAM13A regulates the CPT1A–FAO pathway in chronic obstructive pulmonary disease (COPD).  (Ref. 5)

FAM13A regulates the CPT1A–FAO pathway in chronic obstructive pulmonary disease (COPD). FAM13 locus has been shown to be associated with increased expression of FAM13A and higher risk for COPD. FAM13 in association with SIRT1 induces the expression of CPT1A, a key enzyme that regulates fatty acid oxidation (FAO) in the mitochondria. Up-regulation of CPT1A promotes FAO and high production of ROS, leading to lung epithelial cell death; this is an important pathogenic mechanism that might lead to COPD. CAT, carnitine translocase; CoA, coenzyme A; CPT1A, carnitine palmitoyltransferase 1; CPT2, carnitine palmitoyltransferase 2; FACS, fatty acyl-CoA synthase; FADH2, flavin adenin dinucleotide; FAM13A, family with sequence similarity 13 member A; NADH, nicotinamide adenine dinucleotide reduced; ROS, reactive oxygen species; SIRT1, sirtuin 1; TCA, tricarboxylic acid cycle.  (Ref. 5)


Is FAM13A and CPT1A/CPT1B involved in the ME/CFS pathomechanism?

References

1. Naviaux et al: Metabolic features of CFS. www.pnas.org/cgi/doi/10.1073/pnas.1607571113
2. Germain et al: Metabolic profiling of a ME/CFS discovery cohort reveals disturbances in fatty acid and lipid metabolism. Mol. BioSyst. 2017, 13, 371.
3. Nagy-Szakal et al: Insights into ME/CFS phenotypes through comprehensive metabolomics. Nat Sci Rep 2018, 8.
4. Reuter and Evans: Long-chain acylcarnitine deficiency in patients with CFS. Potential involvement of altered carnitine palmitoyltransferase-I-activity. J. Int. Med. 2011, 270.
5. Hawkins et al. FAM13A, A Fatty Acid Oxidation Switch in Mitochondria. Friend or Foe in Chronic Obstructive Pulmonary Disease Pathogenesis?  Am J Respir Cell Mol Biol. 2017 Jun; 56(6): 689–691.doi: 10.1165/rcmb.2017-0080ED  https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5516298/
6. Trivedi et al: Identification of ME/CFS - associated DNA methylation patterns. Plos One 2018, 13, 7.
7. de Vega et al: Epigenetic modifications and glucocorticoid sensitivity in ME/CFS. BMC Medical Genomics, 2017, 10, 11.
8. Nguyen et al: Whole blood gene expression in adolescent CFS: an exploratory crosssectional study suggesting altered B cell differentiation and survival. J Transl Med. 2017,15,102

ACSL3, GPD2 og PDHX i ME

Forskere har anvendt en ny forbedret teknologi til at undersøge epigenetiske ændringer i blodceller fra ME patienter. Resultatet blev valideret på ME patienter fra både USA og Spanien (1).

De supplerende tabeller i artiklen rummer værdifulde informationer. Jeg vil nævne tre eksempler (ACSL3, GPD2 og PDHX) fra Tabel 7.

Tabel S7 er en liste over 144 hypomethylerede gen promotorer. Promotoren er det område på DNA'et, hvorfra transkriptionen af et gen starter. Ændret DNA methylering af promotoren har stor betydning for genekspressionen. Man kan formode, at de 144 hypomethylerede gener er "på overarbejde", og at dette er relateret til ME sygdomsmekanismen. Vi ved ikke hvordan. Måske er de på overarbejde for at kompensere for noget, der er gået galt i cellens metabolisme?

ACSL3
Acyl-CoA synthetase long-chain family member 3 (ACSL3) aktiverer langkædede fedtsyrer til både syntese af cellulære lipider og til nedbrydning via beta-oxidation. ACSL3 er nødvendig for at bygge fedtsyrer ind i phosphatidylcholine.

ACSL3 interagerer med proteiner kodet af generne: CPT1A, CPT1B, PNPLA2 og MGLL. Disse proteiner har en central rolle i lipidmetabolismen. Et tidligere epigenetisk studie har vist, at disse gener er påvirkede hos ME patienter (2).

GPD2
Glycerol-3-phosphate dehydrogenase 2 (GPD2) er et mitokondrie enzym, som er komponent i den respiratoriske kæde og i glycerophosphate-shuttlen. Det er et nøgleenzym i skæringspunktet mellem glykolyse, oxidativ fosforylering og fedtsyrer metabolismen (3).

PDHX
Pyruvate dehydrogenase complex component X (PDHX) er en delkomponent i PDH-komplekset. Hvis der opstår fejl i dette gen, eller PDHX-proteinet udsættes for et autoimmunt angreb bliver man alvorlig syg. Læs om genet: https://www.ncbi.nlm.nih.gov/gene/8050

Spørgsmålet er hvorfor er dette gen epigenetisk ændret hos ME patienter? Et nedreguleret PDH-kompleks er allerede sat i centrum af ME sygdoms-mekanismen (4),

Referencer:

1. Trivedi et al: Identification of ME/CFS - associated DNA methylation patterns. Plos One 2018, 13, 7 http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0201066
2. de Vega et al: Epigenetic modifications and glucocorticoid sensitivity in ME/CFS. BMC Medical Genomics, 2017, 10, 11
3. Mracek et al: The function and the role of the mitochondrial glycerol-3-phosphate dehydrogenase in mammalian tissue. Biochimica et Biophysica Acta 1827, 2013, 401-410.
4. Fluge et al: Metabolic profiling indicates impaired pyruvate dehydrogenase function in myalgic encephalopathy / chronic fatigue syndrome. JCI Insight. 2016; 1(21):e89376. Doi 10.1172/jci.insight.89276

Functions of DNA methylation

Functions of DNA methylation
Abstract (ref 1):
DNA methylation is frequently described as a 'silencing' epigenetic mark, and indeed this function of 5-methylcytosine was originally proposed in the 1970s. Now, thanks to improved genome-scale mapping of methylation, we can evaluate DNA methylation in different genomic contexts: transcriptional start sites with or without CpG islands, in gene bodies, at regulatory elements and at repeat sequences. The emerging picture is that the function of DNA methylation seems to vary with context, and the relationship between DNA methylation and transcription is more nuanced than we realized at first. Improving our understanding of the functions of DNA methylation is necessary for interpreting changes in this mark that are observed in diseases such as cancer.

The effects of variations in CpG methylation on coding genes depend on the location

Quote (from ref 2, page 10):

The effects of variations in CpG methylation on coding genes depend on the location of the

differential methylation in relation to the gene of interest. There are two genic regions where the

relationship between CpG methylation and gene expression are generally well understood.

Increased CpG methylation in the TSS of genes tends to lead to a decrease in transcription either

due to the prevention of transcription factors from recognizing and binding to the promoter region for the initiation of transcription, or due to the recruitment of other methyl-binding

proteins and co-repressors [19, 56, 57]. On the other hand, increased CpG methylation in the

gene body is positively correlated with transcription [58]. Although the exact reasons are

unclear, gene body methylation is believed to be important for silencing repetitive elements,

retrotransposons, and regulating alternative splicing events [52]. (references are in ref 2)

References:

1) Peter A. Jones: Functions of DNA methylation: islands, start sites, gene bodies and beyond. Nat Revet Genet, 2012, 13, 7.

https://www.nature.com/articles/nrg3230

2) de Vega: 

DNA Methylation Modifications Associated with Glucocorticoid Sensitivity and Clinical Subtypes of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS)

https://tspace.library.utoronto.ca/handle/1807/89701

Mitokondrieproteiner i ME

Vi forstår endnu ikke mitokondriernes rolle i ME sygdoms-mekanismen. Mitokondrieproteiner dukker sporadisk op i ME forskningen. Lad os se om disse mitokondrie påvirkninger danner et samlet billede, som kan lede os på sporet af hvad der foregår i ME sygdomsmekanismen.

I studiet af DNA methyleringer på CD4⁺ - T celler fra ME patienter, er to gener, OXA1L og FTSJ2, henholdsvis hypo- og hypermethyleret. Generne koder proteiner, der anvendes af mitokondrierne (1).

En tommelfingerregel er, at hypomethylering af et gen er forbundet med øget transkription, og hypermethylering er en nedregulering af genet. Virkeligheden er mere kompliceret, idet methyleringens placering er afgørende for påvirkningen af genet. Som udgangspunkt må vi nøjes med at sige, at generne er epigenetisk ændrede hos ME patienter i forhold til raske kontrolpersoner.

OXA1L
I mitokondriernes membraner bygges de respiratoriske komplekser OXPHOS, der søger for, at cellen kan danne energimolekylerne ATP. Del-komponenter til OXPHOS komplekserne kodes af DNA fra cellekernen og fra mitokondriernes eget DNA. Når del-komponenterne er produceret, skal de bygges ind i mitokondriemembranerne og blive til OXPHOS komplekser. OXA1L har en vigtig rolle i OXPHOS byggeprocessen (2).

OXA1L interagerer også med mitokondrie ribosomerne, dvs proteinet er knyttet til translationen. Den tilknytning er relateret til den respiratoriske funktion.

Af genecards.org STRING interaction network kan man se de nærmeste proteiner, som OXA1L interagerer med. Nogle af disse OXA1L-samarbejdspartnere dukker også op i ME forskningen:

MRPL23, er mitochondrial ribosomal protein L23, og har øget genekspression hos ME patienter i tre studier (3, 4, 5).

ATP5G2, en ATP synthase knyttet til de respiratoriske komplekser. Genet er hypermethyleret (TSS1500) i et studie (6), og hypomethyleret (1stExon) i et andet studie af ME patienter (7).

ATP6V0C, en del komponent i v-ATPasen. Genet er hypermethyleret (body) og flere andre v-ATPase delkomponenter er fundet epigenetisk ændrede hos ME patienter (6).

FTSJ2 = MRM2
FTSJ2 hedder også mitochondrial rRNA methyltransferase 2, MRM2. Proteinet arbejder sammen med MRM3 om dannelse af en mitokondrie-ribosom del-komponent. Hvis man inaktiverer MRM2 og MRM3 i celler forminskes den respiratoriske kapacitet, som en konsekvens af formindsket mitokondrie-translation (8).

Defekt MRM2 vil resultere i MELAS- syndrom lignende symptomer. MELAS = mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (9).

MRM2 interagerer med proteinet thyroid adenoma associated (THADA). THADA regulerer balancen mellem energi opbevaring og varme produktion. THADA arbejder sammen med et protein, der hedder SERCA. SERCA regulerer Ca²⁺ ligevægten i cellen (10).

Genet THADA er hypermethyleret (body) hos ME patienter i et studie (6), og hypomethyleret (3'UTR) i et andet studie (7).

MRRF
I to af studierne hvor ME patienterne havde øget genekspression af MRPL23, så man også øget genekspression af mitochondrial ribosome recycling, MRRF (4, 5).

Mitokondrie-ribosomerne genbruges til kontinuerlig OXPHOS-produktion. I et forsøg med raske celler så man, at kunstig nedreulering af MRRF medførte:

• nedsat niveau af samlede OXPHOS-komplekser
• nedsat respiration

Det interessante var, at der var en forsinkelse på flere dage, førend den manglende proteinsyntese manifesterede sig (11).

Spørgsmål er om denne OXPHOS-levering-forsinkelse kan sættes i relation til ME patienters post-exertional-malaise (PEM), som bekendt er et ME-diagnosekriterie. Arbejder ME-celler på højtryk for at levere OXPHOS, og i givet fald - hvorfor?

GFM1

G elongation factor mitochondrial 1 (GFM1) er en mitokondrial translations faktor. Hvis GFM1 bliver nedreguleret resulterer det i OXPHOS mangel, og symptomerne er encephalopathy og multi-system sygdom (12).

Genet GFM1 er hypermethyleret (body) hos ME patienter, og dette er relateret til patienternes livskvalitet (6).

Jeg har ikke fundet alle relevante DNA methyleringer. Dette var blot et lille eksempel på hvad ME forskningen rummer af ikke udnyttet viden.

Referencer:

1. Brenu et al: Methylation profile of CD4+ T cells in CFS/ME. J. Clin Cell Immunol 5, 228
2. Stoldt et al: The inner-mitochondrial distribution of OXA1 depends on the growth conditions and on the availability of substrates. MBOC, 2012, 23.
3. Kaushik et al: Gene expression in PBMC from patients with CFS. J Clin Pathol, 2005, 58
4. Kerr et al: Gene expression subtypes in patients with CFS/ME. JID, 2008, 197.
5. Frampton et al: Assessment of a 44 gene classifier for the evaluation of CFS from PBMC gene expression. Plos one, 2011, 6, 3.
6. de Vega et al: Epigenetic modifications and glucocorticoid sensitivity in ME/CFS. BMC Medical Genomics, 2017, 10, 11.
7. Trivedi et al: Identification of ME/CFS - associated DNA methylation patterns. Plos One 2018, 13, 7.
8. Rorbach et al: MRM2 and MRM3 are involved in biogenesis of the large subunit of the mitochondrial ribosome. Mol Biol Cell, 2014, 25, 17.
9. Garone et al: Defective MRM2 causes MELAS-like clinical syndrome. Hum Mol Genet, 2017, 26, 21.
10. Moraru et al: THADA regulates the organismal balance between energy storage and heat production. Dev Cell, 2017, 41, 1.
11. Rorbach et al: The human mitochondrial ribosome recycling factor is essential for cell viability. Nucleic Acids Research, 2008, 36, 18.
12. Simon et al: Activation of a cryptic slice site in the mitochondrial elongation factor GFM1 causes combined OXPHOS deficiency.  Mitochondrion 2017, 34.

SPI1 and ME

SPI1 (=PU.1) is a transcription factor involved in hematopoiesis. SPI1 regulates B-cell development, but is also important for maturation of macrophages, T cell progenitors and T helper 9 cells.

SPI1 can either activate or repress the transcription of genes. This is mediated by the ability of SPI1 to build different complexes with a number of different protein partners (1).

In leukemia SPI1 can be either an oncogene or a tumor suppressor. SPI1 overexpression can trigger cellular senescence, and may be a safeguard against leukemia (12).

EP300 is a transcriptional co-activator, which interacts with SPI1. Epstein-Barr Virus nuclear antigen leader protein (EBNALP) coactivates EP300 and hereby dysregulates SPI1. An EP300 inhibitor can abolish EBNALP coactivation and may have the potential to control EBV-associated diseases (3).

SPI1 is a putative upstream regulator in adolescent CFS patients (table S4 in ref 4).

The gene SPI1 is hypermethylated (body, 5'UTR, 1stExon) in Me patients in one study (5), and hypomethylated (TSS200) in another study (6).

SPI1 interacts with several proteins where the genes are epigenetic changed. Fx, FLT3, CREBBP, E2F3, ELANE, HOXA10, GFI1, TFDP1 and IRF4 (5)

SPI1 also interacts with KAT2B (= EP300 / CBP associated factor). The gene promoter of KAT2B is hypomethylated in ME patients (6).

EP300 and CREBBP interacts with CITED2 (= CBP / p300 - interacting transactivator). CITED2 geneekspression is upregulated in ME patients (7, 8).

References:

1. Riel and Rosenbauer: Epigenetic control of hematopoiesis. Biol Chem 2014, 395, 11.
2. Delestré et al: Senescence is a Spi1-induces anti-proliferative mechanism in primary hematopoietic cells. Haematologica 2017, 102, 11.
3. Wang et al: Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300. J Virol, 2018, 92, 9.
4. Nguyen et al: Whole blood gene expression in adolescent CFS: an exploratory crosssectional study suggesting altered B cell differentiation and survival. J Transl Med. 2017,15,102.
5. de Vega et al: Epigenetic modifications and glucocorticoid sensitivity in ME/CFS. BMC Medical Genomics, 2017, 10, 11.
6. Trivedi et al: Identification of ME/CFS - associated DNA methylation patterns. Plos One 2018, 13, 7.
7. Kerr et al: Gene expression subtypes in patients with CFS/ME. JID, 2008, 197.
8. Frampton et al: Assessment of a 44 gene classifier for the evaluation of CFS from PBMC gene expression. Plos one, 2011, 6, 3.