metabolomics analysis reveals differential t-cell serine...
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![Page 1: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/1.jpg)
Metabolomics Analysis Reveals Differential T-Cell Serine Metabolism as a Target in Autoimmunity Gabriela Andrejeva Jeffrey Rathmell lab Vanderbilt University Medical Center [email protected] 2018 UK Metabolism Symposium Monday, 30th July 2018
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Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
2
AGXT2L-Cystine
DLD
L-Arginine
CoA
ADC
5,10-Methylenetetrahydrofolate"
CDO1
GOT2
4-Aminobutanoate
MPST
GOT1
2-OxoglutarateMercaptopyruvate
Pyruvate
L-Cysteate
Cyanide
3-Sulfino-L-alanine
GPT
Taurine
GAD1
beta-Alanine
GAD2
GPT2
CSAD
Hypotaurine
SDS
5-FormiminotetrahydrofolateThiocysteine
2-Oxobutanoate
2-Aminoacrylate
L-Glutamate
L-Cysteine
L-Alanine
Oxaloacetate
AGXTCTH
L-Cystathionine
4-Guanidinobutanoate
Succinyl-CoA
GAMT
Guanidinoacetate
Creatine
GSS
DMGDH
S-Adenosyl-L-homocysteine
Betaine aldehyde
CO2
S-Aminomethyldihydrolipoylprotein
L-Pipecolate
PDPR
CHDH
Choline
Sarcosine
GLDC
SARDH
ALAS1
Glutathione
ALAS2
PIPOX
GNMT
GCAT
L-2-Amino-3-oxobutanoic acid
5-Aminolevulinate
MTRBHMT2
"N,N-Dimethylglycine"
BHMT
Selenohomocysteine
AHCYL1
L-Homocysteine
BetaineCob(II)alamin
AHCY
PPCS
ANPEPHydroxypyruvate
CP
Acetyl-CoA
GATM
Se-Adenosylselenomethionine
L-Methionine
MAT1AMAT2B
S-Adenosyl-L-methionine
MAT2A
CBS
PPIGGlycine
IL4I1Cystathionine
ALDH1L2
ALDH1L1
Tetrahydrofolyl-[Glu](n)
Poly-L-glutamate
10-Formyltetrahydrofolylpolyglutamate 10-Formyltetrahydrofolyl L-glutamate
GGH
ATIC
MTHFD2
DHFR
MTHFD1
10-Formyltetrahydrofolate
Tetrahydrofolate
MTHFD1L
"5,10-Methylenetetrahydrofolate"
FPGS
"5,10-Methenyltetrahydrofolate"
Formate
O-Phospho-L-serine
5-MethyltetrahydrofolateMTHFR
MTHFS
FTCD
MTFMT
N-Formimino-L-glutamatePSPH
GART
PHGDH
3-Phosphonooxypyruvate
2-Oxo-3-hydroxy-4-phosphobutanoate
3-Phospho-D-glycerate
O-Phospho-4-hydroxy-L-threonine PSAT1
NAGS
SHMT1
NH3 L-Serine
AMT5-Formyltetrahydrofolate
SHMT2
dUMP dTMP
Folate
DihydropteroateDihydrofolate
TYMS
Glucose
Serine
Methylene-THF
Serine
ExtracelularSerine
Methylene-THF
FormateFormate
MTHFD2
NADPH NADPH/NADHPURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Treg
Th17
Th1
![Page 3: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/3.jpg)
Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
3
![Page 4: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/4.jpg)
CD4 T Cells and Metabolism 4
Adapted from Swain et al., Nat Rev Immunol, 2012
Th2 GATA3
Treg FoxP3
Th17 Ror𝛾t
Th1 T-bet
TGFβ IL-6
IL-23 TGFβ IL-2
Antigen Recognition & Co-Stimulation
IL-12 IFN𝛾 IL-4
IL-10 IL-17A, IL-17F, IL-22
IFN𝛾, TNF IL-4, IL-5, IL-13
POLARIZING MILEU
EFFECTOR MEDIATORS
EFFECTOR FUNCTIONS
Regulation, suppression of inflammatory responses
Inflammation Allergic and helminth
responses
Macrophage activation,
inflammation
Naïve
TCR
![Page 5: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/5.jpg)
CD4 T Cells and Metabolism 5
Adapted from Swain et al., Nat Rev Immunol, 2012
Th2 GATA3
Treg FoxP3
Th17 Ror𝛾t
Th1 T-bet
TGFβ IL-6
IL-23 TGFβ IL-2
Antigen Recognition & Co-Stimulation
IL-12 IFN𝛾 IL-4
IL-10 IL-17A, IL-17F, IL-22
IFN𝛾, TNF IL-4, IL-5, IL-13
POLARIZING MILEU
EFFECTOR MEDIATORS
EFFECTOR FUNCTIONS
Regulation, suppression of inflammatory responses
Inflammation Allergic and helminth
responses
Macrophage activation,
inflammation
Naïve
TCR
Sugiura & Rathmell., J Immunol, 2018
![Page 6: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/6.jpg)
CD4 T Cells and the Immunological Balance 6
Th2 Treg
Tolerance Normal Immunity
Th17
Th1 Th2
Treg
Immunosuppression Cancer
Th17
Th1
Th2
Treg
Inflammation Autoimmunity
Th17
Th1
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Hypothesis
Cytokine signaling during CD4 T cell development induces differential
metabolic rewiring to fulfill biosynthetic, signaling and epigenetic
roles.
These are selectively essential for the development and function of
different CD4 T cell subsets.
7
![Page 8: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/8.jpg)
Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
8
![Page 9: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/9.jpg)
Experimental System for Metabolomics and Proteomics
9
Cytokines added Outcome TGFβ Treg TGFβ + IL-2 Treg proliferative IL-12 Th1 IL-12 + IL-2 Th1 TGFβ + IL-6 Th17 TGFβ + IL-6 + IL-23 More pathogenic Th17 TGFβ + IL-6 + IL-23 + IL-1β
Pathogenic Th17
Nothing Th0
Cytokines added Outcome IL-7 Naïve T cell
Isolated naïve
mouse CD4+ T
cell
anti-CD3 anti-CD28
CD28 CD3
TCR
SIGNAL 1 SIGNAL 2
anti-CD3 and anti-CD28 coated plate (5ug/ml each)
Isolated naïve
mouse CD4+ T
cell
Immune Characterization • Transcription Factor Expression • Cytokine Secretion
Metabolic Characterization • Intracellular Metabolite MS • Medium Analysis by NMR • Proteomics
24h, 48h, 72h
![Page 10: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/10.jpg)
Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
10
![Page 11: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/11.jpg)
Serine/Glycine/One-Carbon Metabolism 11
Glucose
Serine
Methylene-THF
Serine
Extracellular Serine
Methylene-THF
Formate Formate
NADPH NADPH/ NADH PURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Glycine
Glycine
![Page 12: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/12.jpg)
Serine/Glycine/One-Carbon Metabolism 12
Glucose
Serine
Methylene-THF
Serine
Extracellular Serine
Methylene-THF
Formate Formate
MTHFD2
NADPH NADPH/ NADH PURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Glycine
Glycine
![Page 13: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/13.jpg)
Serine/Glycine/One-Carbon Metabolism 13
Glucose
Serine
Methylene-THF
Serine
Extracellular Serine
Methylene-THF
Formate Formate
MTHFD2
NADPH NADPH/ NADH PURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Th17 ? Glycine
Glycine
TReg
Th1
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Possible Mechanisms of MTHFD2 Inhibitor Action
Nucleotide Synthesis Reactive Oxygen Species
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The Model of MTHFD2 Inhibitor Action 15
Glucose
Serine
Methylene-THF
Serine
Extracellular Serine
Methylene-THF
Formate Formate
MTHFD2
NADPH NADPH/ NADH PURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Th17
Glycine
Glycine
TReg
Th1
AICAR
AMPK
mTORC1
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Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
16
![Page 17: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/17.jpg)
Outline
Background and hypothesis
Global metabolomics and proteomics studies of CD4 T cells
Serine pathway in CD4 T cell subsets
Serine pathway and MTHFD2 in EAE and multiple sclerosis
Summary and future directions
17
![Page 18: Metabolomics Analysis Reveals Differential T-Cell Serine ...bioinformatics.cesb.uky.edu/pub/RCSIRM/Workshop...CD4 T Cells and Metabolism 5 Adapted from Swain et al., Nat Rev Immunol,](https://reader030.vdocument.in/reader030/viewer/2022041023/5ed4ae93d18c7b5d8f4baa8d/html5/thumbnails/18.jpg)
Summary and Future Directions
Global metabolomics analysis can reveal pathways that are important in CD4 T cell activation and subset function
Naïve CD4 T cell differentiation with lineage-specific cytokines results in significant alterations of serine metabolism
Targeting MTHFD2 might be a viable strategy to shift the balance between Th1/Th17/Treg in human inflammatory disease In vivo: Does targeting MTHFD2 suppress autoimmunity and promote Treg stability?
In humans: Is MTHFD2 expression increased in human multiple sclerosis patient brain lesions?
18
AGXT2L-Cystine
DLD
L-Arginine
CoA
ADC
5,10-Methylenetetrahydrofolate"
CDO1
GOT2
4-Aminobutanoate
MPST
GOT1
2-OxoglutarateMercaptopyruvate
Pyruvate
L-Cysteate
Cyanide
3-Sulfino-L-alanine
GPT
Taurine
GAD1
beta-Alanine
GAD2
GPT2
CSAD
Hypotaurine
SDS
5-FormiminotetrahydrofolateThiocysteine
2-Oxobutanoate
2-Aminoacrylate
L-Glutamate
L-Cysteine
L-Alanine
Oxaloacetate
AGXTCTH
L-Cystathionine
4-Guanidinobutanoate
Succinyl-CoA
GAMT
Guanidinoacetate
Creatine
GSS
DMGDH
S-Adenosyl-L-homocysteine
Betaine aldehyde
CO2
S-Aminomethyldihydrolipoylprotein
L-Pipecolate
PDPR
CHDH
Choline
Sarcosine
GLDC
SARDH
ALAS1
Glutathione
ALAS2
PIPOX
GNMT
GCAT
L-2-Amino-3-oxobutanoic acid
5-Aminolevulinate
MTRBHMT2
"N,N-Dimethylglycine"
BHMT
Selenohomocysteine
AHCYL1
L-Homocysteine
BetaineCob(II)alamin
AHCY
PPCS
ANPEPHydroxypyruvate
CP
Acetyl-CoA
GATM
Se-Adenosylselenomethionine
L-Methionine
MAT1AMAT2B
S-Adenosyl-L-methionine
MAT2A
CBS
PPIGGlycine
IL4I1Cystathionine
ALDH1L2
ALDH1L1
Tetrahydrofolyl-[Glu](n)
Poly-L-glutamate
10-Formyltetrahydrofolylpolyglutamate 10-Formyltetrahydrofolyl L-glutamate
GGH
ATIC
MTHFD2
DHFR
MTHFD1
10-Formyltetrahydrofolate
Tetrahydrofolate
MTHFD1L
"5,10-Methylenetetrahydrofolate"
FPGS
"5,10-Methenyltetrahydrofolate"
Formate
O-Phospho-L-serine
5-MethyltetrahydrofolateMTHFR
MTHFS
FTCD
MTFMT
N-Formimino-L-glutamatePSPH
GART
PHGDH
3-Phosphonooxypyruvate
2-Oxo-3-hydroxy-4-phosphobutanoate
3-Phospho-D-glycerate
O-Phospho-4-hydroxy-L-threonine PSAT1
NAGS
SHMT1
NH3 L-Serine
AMT5-Formyltetrahydrofolate
SHMT2
dUMP dTMP
Folate
DihydropteroateDihydrofolate
TYMS
Glucose
Serine
Methylene-THF
Serine
ExtracelularSerine
Methylene-THF
FormateFormate
MTHFD2
NADPH NADPH/NADHPURINES
GSH
METHIONINE
GSH
MITOCHONDRION
CYTOSOL
Treg
Th17
Th1
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Acknowledgements 19
Rathmells’ labs • Jeffrey Rathmell • Kimryn Rathmell • Kathryn Beckermann • Anna Chytil • Diana Contreras • Aguirre De cubas • Stephanie Dudzinski • Marc Johnson • Aaron Lim • Matthew Madden • Damian Maseda • Frank Mason • Stephen Norris • Bradley Reinfeld • Ayaka Sugiura • Esteban Terzo • Jamie Weyandt • Melissa Wolf • Kirsten Young
Young lab • Jamey Young • Irina Trenary • Ali McAtee
Vanderbilt NMR cores • Donald Stec • Markus Voehler
Sriram lab • Subramaniam Sriram
Aune lab • Thomas Aune
McLean lab • Stacy Sherrod • Alexandra Rutledge • Simona Codreanu
Vanderbilt MS core • Kevin Schey • Wade Calcutt • Emilio Rivera • Kristie Lindsey Rose
Norris lab • Jeremy Norris • Danielle Guttierez