{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002980","ANALYSIS_ID":"AN004897","VERSION":"1","CREATED_ON":"November 19, 2023, 6:47 pm"},

"PROJECT":{"PROJECT_TITLE":"Glycine homeostasis requires reverse SHMT flux","PROJECT_TYPE":"LCMS metabolomics","PROJECT_SUMMARY":"The folate-dependent enzyme serine hydroxymethyltransferase (SHMT) reversibly converts serine into glycine and a tetrahydrofolate-bound one-carbon unit. Such one-carbon unit production plays a critical role in development, the immune system, and cancer. Using rodent models, here we show that the whole-body SHMT flux acts to net consume rather than produce glycine. Pharmacological inhibition of whole-body SHMT1/2 and genetic knockout of liver SHMT2 elevated circulating glycine levels up to eight-fold. Stable isotope tracing revealed that the liver converts glycine to serine, which is then converted by serine dehydratase into pyruvate and burned in the tricarboxylic acid cycle. In response to diets deficient in serine and glycine, de novo biosynthetic flux was unaltered but SHMT2- and serine dehydratase-mediated catabolic flux was lower. Thus, glucose-derived serine synthesis does not respond to systemic demand. Instead, circulating serine and glycine homeostasis is maintained through variable consumption, with liver SHMT2 a major glycine-consuming enzyme.","INSTITUTE":"Princeton University","DEPARTMENT":"Department of Chemistry","LABORATORY":"Josh Rabinowitz","LAST_NAME":"McBride","FIRST_NAME":"Matthew","ADDRESS":"Carl Icahn Lab, South Drive, Princeton, NJ 08544","EMAIL":"matthewmcbride@princeton.edu","PHONE":"8567457389"},

"STUDY":{"STUDY_TITLE":"Water soluble metabolomics in mice upon loss of SHMT","STUDY_SUMMARY":"The enzyme SHMT interconverts the amino acids serine and glycine as part of the folate cycle. To explore the role of SHMT in amino acid homeostasis, Mice were treated with a small molecule inhibitor of SHMT (SHIN2) or had Shmt2 genetically knocked-out in a liver specific manner. Serum and liver samples were collected and underwent LC-MS metabolomics analysis.","INSTITUTE":"Princeton University","LAST_NAME":"McBride","FIRST_NAME":"Matthew","ADDRESS":"Carl Icahn Lab, South Drive, Princeton, NJ 08544","EMAIL":"matthewmcbride@princeton.edu","PHONE":"8567457389"},

"SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"Veh_1_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_1_serum_NEG.mzXML","RAW_FILE_NAME":"Veh_1_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"Veh_2_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_2_serum_NEG.mzXML","RAW_FILE_NAME":"Veh_2_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"Veh_3_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_3_serum_NEG.mzXML","RAW_FILE_NAME":"Veh_3_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_1_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_1_serum_NEG.mzXML","RAW_FILE_NAME":"SHIN2_1_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_2_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_2_serum_NEG.mzXML","RAW_FILE_NAME":"SHIN2_2_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_3_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_3_serum_NEG.mzXML","RAW_FILE_NAME":"SHIN2_3_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"Veh_1_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_1_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Veh_2_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_2_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Veh_3_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"Vehicle"},
"Additional sample data":{"RAW_FILE_NAME":"Veh_3_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_1_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_1_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_2_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_2_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"SHIN2_3_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"SHIN2"},
"Additional sample data":{"RAW_FILE_NAME":"SHIN2_3_liver_NEG.mzXML","RAW_FILE_NAME":"-"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_15_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_15_serum_NEG.mzXML","RAW_FILE_NAME":"WT_15_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_20_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_20_serum_NEG.mzXML","RAW_FILE_NAME":"WT_20_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_34_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_34_serum_NEG.mzXML","RAW_FILE_NAME":"WT_34_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_37_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_37_serum_NEG.mzXML","RAW_FILE_NAME":"WT_37_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_46_serum",
"Factors":{"Sample type":"Serum","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_46_serum_NEG.mzXML","RAW_FILE_NAME":"WT_46_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_21_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_21_serum_NEG.mzXML","RAW_FILE_NAME":"KO_21_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_22_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_22_serum_NEG.mzXML","RAW_FILE_NAME":"KO_22_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_23_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_23_serum_NEG.mzXML","RAW_FILE_NAME":"KO_23_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_35_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_35_serum_NEG.mzXML","RAW_FILE_NAME":"KO_35_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_43_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_43_serum_NEG.mzXML","RAW_FILE_NAME":"KO_43_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_44_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_44_serum_NEG.mzXML","RAW_FILE_NAME":"KO_44_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_45_serum",
"Factors":{"Sample type":"Serum","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_45_serum_NEG.mzXML","RAW_FILE_NAME":"KO_45_serum_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_15_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_15_liver_NEG.mzXML","RAW_FILE_NAME":"WT_15_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_20_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_20_liver_NEG.mzXML","RAW_FILE_NAME":"WT_20_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_34_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_34_liver_NEG.mzXML","RAW_FILE_NAME":"WT_34_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_37_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_37_liver_NEG.mzXML","RAW_FILE_NAME":"WT_37_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_WT_46_liver",
"Factors":{"Sample type":"Liver","Genotype":"Wild-type","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"WT_46_liver_NEG.mzXML","RAW_FILE_NAME":"WT_46_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_21_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_21_liver_NEG.mzXML","RAW_FILE_NAME":"KO_21_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_22_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_22_liver_NEG.mzXML","RAW_FILE_NAME":"KO_22_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_23_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_23_liver_NEG.mzXML","RAW_FILE_NAME":"KO_23_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_35_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_35_liver_NEG.mzXML","RAW_FILE_NAME":"KO_35_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_43_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_43_liver_NEG.mzXML","RAW_FILE_NAME":"KO_43_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_44_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_44_liver_NEG.mzXML","RAW_FILE_NAME":"KO_44_liver_POS.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"SHMT2flox_KO_45_liver",
"Factors":{"Sample type":"Liver","Genotype":"Liver KO","Treatment":"None"},
"Additional sample data":{"RAW_FILE_NAME":"KO_45_liver_NEG.mzXML","RAW_FILE_NAME":"KO_45_liver_POS.mzXML"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"For pharmacological inhibition of SHMT, mice were treated with Vehicle or SHIN2 for 12 hours and then blood serum and liver samples were collected. For genetic loss of Shmt2, blood serum and liver samples were collected from mice 21 days after liver-specific gene knockout.","SAMPLE_TYPE":"Blood (serum) and Liver"},

"TREATMENT":{"TREATMENT_SUMMARY":"For pharmacological inhibition of SHMT, mice (C57BL/6N) were divided into two groups and received Vehicle (20% 2-hydroxypropyl-β-cyclodextrin) or SHIN2 (200 mg/kg) for 12 hours. For genetic loss of Shmt2, Shmt2(flox/flox) mice and wild-type litter mate controls were injected with AAV8-TBG-Cre viral particles to induce liver-specific gene knockout and samples were harvested 21 days later."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Water soluble metabolites were extracted from serum and liver samples. 3ul of serum was extracted with 120 ul (40X) of 100% methanol, cooled on ice for 10 minutes, centrifuged at 16,000 x g for 30 minutes, and supernatant collected. 20mg of ground liver tissue was extracted with 800ul of 40:40:20 methanol:acetonitrile:water, cooled on ice for 10 minutes, centrifuged at 16,000 x g for 30 minutes, and supernatant collected."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_TYPE":"HILIC","INSTRUMENT_NAME":"Thermo Vanquish","COLUMN_NAME":"Waters XBridge BEH Amide (100 x 2.1mm,2.5um)","SOLVENT_A":"95% water/5% acetonitrile with 20 mM ammonium acetate, 20 mM ammonium hydroxide, pH 9.4","SOLVENT_B":"100% acetonitrile","FLOW_GRADIENT":"0 minutes, 85% B; 2 minutes, 85% B; 3 minutes, 80% B; 5 minutes, 80% B; 6 minutes, 75% B; 7 minutes, 75% B; 8 minutes, 70% B; 9 minutes, 70% B; 10 minutes, 50% B; 12 minutes, 50% B; 13 minutes, 25% B; 16 minutes, 25% B; 18 minutes, 0% B; 23 minutes, 0% B; 24 minutes, 85% B; 30 minutes, 85% B","FLOW_RATE":"150 μl/min","COLUMN_TEMPERATURE":"25°C"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Plus Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_COMMENTS":"MS full scans were in negative ion mode with a resolution of 140,000 at m/z 200 and scan range of 70–1,000 m/z. The automatic gain control (AGC) target was 1 × 10^6. LC-MS peak files were analyzed and visualized with El-MAVEN (Elucidata) using 5 ppm ion extraction window, minimum peak intensity of 1 x 10^5 ions, and minimum signal to background blank ratio of 2.","MS_RESULTS_FILE":"ST002980_AN004897_Results.txt UNITS:Peak area Has m/z:Yes Has RT:Yes RT units:Minutes"}

}