Differential abundance upon SLC overexpression was tested for each metabolite by contrasting its normalized and batch-corrected values in doxycycline induced samples to the corresponding uninduced samples (4 replicates each) in a one-way ANOVA for WT-OE cell lines, or a two-way ANOVA for KO-OE cell lines, modeling the clone effect as additional factor. Resulting p-values were corrected for multiple testing per SLC, using the Benjamini-Hochberg procedure (Benjamini & Hochberg, 1995). Differentially abundant metabolites were called at a false discovery rate of 5%.
"],"repository":["MetaboLights"],"study_status":["Public"],"ptm_modification":[""],"instrument_platform":["Liquid Chromatography MS - negative - reverse phase"],"chromatography_protocol":["A 1290 Infinity II UHPLC system (Agilent Technologies) was used for the chromatographic separation utilizing a ZORBAX RRHD Extend-C18, 2.1 x 150 mm, 1.8 µm analytical column (Agilent Technologies) and a SecurityGuard ULTRA Cartridge UHPLC C18 2.1 mm ID precolumn (Phenomenex). The columns were maintained at a temperature of 40 °C. The mobile phase A was 3% MeOH (v/v), 10 mM tributylamine, 15 mM acetic acid in H2O, and mobile phase B was 10 mM tributylamine, 15 mM acetic acid in MeOH. The gradient elution with a flow rate 0.25 mL/min was performed for a total time of 24 min followed by back flushing of the column using a 6 port 2-position divert valve for 8 min using acetonitrile. At the end of the run 8 min of column equilibration with 100% mobile phase A was performed. Chromatographic separation was coupled to a 6470 triple quadrupole mass spectrometer (Agilent Technologies).
"],"publication":["Data- and knowledge-derived functional landscape of human solute carriers. 10.1038/s44320-025-00108-2.","Metabolic mapping of the human solute carrier superfamily. 10.1101/2024.09.23.614124."],"submitter_affiliation":["CeMM - Research Center for Molecular Medicine of the Austrian Academy of Sciences"],"submitter_name":["Ulrich Goldmann"],"organism_part":["LS-180 cell","HCT-116 cell","mixture","Jump-In-T-REx HEK293 cell","SK-MEL-28 cell","1321-N1 cell","HuH-7 cell"],"technology_type":["mass spectrometry"],"disease":[""],"extraction_protocol":["Cells were plated in quadruplicates in 6-well or 24-well plates in culture medium in the absence or presence of 1 μg/mL doxycycline (150 000 cells per well for HCT 116 and LS180, 100,000 cells per well for 1321N1, 50,000 cells per well for Huh-7, 70,000 cells per well for SK-MEL-28, 750,000 / 150,000 cells per well in poly-L-lysine coated 6-well / 24-well plates for Jump-In T-REx HEK293). After 16-24 h, cells were first gently washed with room temperature ammonium carbonate buffer (75 mM NH4HCO3, pH 7.4). Then, cells were transferred to ice, where 300 µL/well in 24-well plate or 1500 µL/well in 6-well plate of ice-cold 80:20 MeOH:H2O was added. The cells were then scraped and transferred to a pre-cooled Eppendorf tube and immediately snap frozen in liquid nitrogen. Samples were thawed on ice before being centrifuged at 16,000 x g for 10 min at 4 °C. The clarified metabolite-containing supernatants were moved into a high-performance liquid chromatography vial and stored at -80 °C until further preparation for LC-MS/MS analysis. Cell extracts were dried using a nitrogen evaporator. The dry residue was reconstituted in 16 µL H2O and 4 µL of sample extract was used for LC-MS/MS analysis. A mixture of isotopically labelled internal standards (Synthetic heavy isotope labelled metabolites from Sigma-Aldrich and Cambridge Isotope Laboratories as well as Metabolite Yeast Extract (U-13C, 98%) from ISOtopic Solutions, Vienna, Austria) was added to each sample either with the extraction solvent or with the H2O at reconstitution.
"],"organism":["mixture","Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/metabolights/MTBLS10077"],"author":["Sabrina Lindinger. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Eirini Christodoulaki. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Shao Thing Teoh. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Abigail Jarret. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Chengzhe Tian. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Kristaps Klavins. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Vitaly Sedlyarov. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Giulio Superti-Furga. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria. GSuperti@cemm.oeaw.ac.at.","Christoph Bueschl. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Daniela Reil. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Tabea Wiedmer. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","J. Thomas Hannich. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Iciar Serrano. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.","Ulrich Goldmann. CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria."],"data_transformation_protocol":["PeakBotMRM (https://github.com/christophuv/PeakBotMRM) was utilized for processing the raw LC-MS/MS data, followed by manual correction of some incorrectly recognized peaks and peak boundaries using the graphical user interface.
Metabolite peak areas were normalized to those of reference internal standards. In total 47 metabolites had internal standards with the same chemical structures, and therefore these respective internal standards were used as references. For the remaining metabolites we performed weighted linear regression on the expected metabolite concentrations against the calculated ratios for each metabolite and each internal standard. When internal standards achieved an adjusted R2 greater or equal to 0.85 in at least 75% of the runs in which the metabolite was measured, the internal standard with the highest percentage was selected as the reference. If multiple internal standards achieved the same highest percentage, all were selected. For 10 metabolites an adjusted R2 greater or equal to 0.85 was achieved in less than 75% and again the internal standard with the highest percentage was chosen.
Individual injections were removed, or a reduced set of internal standards was used for normalization in case certain internal standards were not detected in certain runs. Normalization was performed by calculating the ratio of a metabolite peak area to the corresponding internal standard peak area, and subsequent log-transformation. For metabolites with multiple reference internal standards, the results were averaged. To also enable comparisons between different runs, we corrected for run-to-run differences (batch effects) by centering and scaling the internal-standard normalized values of each metabolite, per run, to its overall (i.e. across runs) median and interquartile range, respectively.
"],"study_factor":["cell line name","replicate","induction","cell line type","protocol detail","gene knock out","run","cell line identifier","gene knock in"],"submitter_email":["ugoldmann@cemm.at"],"sample_collection_protocol":["HCT 116 (RRID:CVCL_0291) and LS180 (RRID:CVCL_0397) cell lines were cultured in RPMI (R8758 Sigma) + 10% Fetal Bovine Serum (10270-106, Lot 42F8381K, Gibco) + Penicillin-Streptomycin (15140-122, Gibco). Jump-In T-REx HEK293 (RRID:CVCL_YL74), HuH-7 (RRID:CVCL_0336), 1321N1 (RRID:CVCL_0110) and SK-MEL-28 (RRID:CVCL_0526) in DMEM (D5796 Sigma) + 10% Fetal Bovine Serum (10270-106, Lot 42F8381K, Gibco) + Penicillin-Streptomycin (15140-122, Gibco). All cell lines were grown at 37 °C and 5% CO2. Transgene expression is induced via doxycycline treatment (1 µg/mL (D9891-1G, Sigma Aldrich; CHEBI:50845).
To generate monoclonal KO clones, cell lines were stably transduced with the lentiviral vector pLentiCRISPRv2 (Addgene 52961) expressing gRNAs targeting SLCs (Table EV1). After 5-10 days puromycin selection, cell pools were sorted to single cells using FACS or a limited dilution series and monoclonal cell lines were expanded and genotyped using an NGS-based approach (Unable to find information for 6872996). At least two independent KO clones were generated for each target. A detailed description of the KO generation can be found here: https://zenodo.org/records/7457297.
To re-express functional and tagged SLCs in the respective KO background, codon-optimized SLC cDNAs were cloned from pDONR221 gateway entry clones (https://www.addgene.org/depositor-collections/re-solute/) into doxycycline-inducible lentiviral gateway destination vectors containing C- or N-terminal HA-Twin-Strep tags (Addgene 194066, 194065) and stably transduced into KO cell lines. For N-terminally tagged SLCs, cDNAs with STOP codons at the end of the SLC ORF were used. Transgene expression was induced via doxycycline treatment at 1 µg/mL (D9891-1G, Sigma Aldrich; CHEBI:50845) for 24 h. For each SLC, two pooled KO-OE cell lines were generated, each derived from an independent clonal KO cell line. A more detailed description of the KO-OE cell line generation can be found here: https://zenodo.org/records/7457295.
To generate cell lines overexpressing codon-optimized tagged SLCs in a WT background (WT-OE), SLC cDNAs were cloned from the same pDONR221 entry clones into modified pJTI R4 CMV-TO MCS pA vectors (Thermo Fisher Scientific) that contained C- or N-terminal HA-Twin-Strep tags. Jump-In T-REx HEK293 cells (Thermo Fisher Scientific) were stably transfected with these constructs as recommended by the manufacturers protocol. Transgene expression was induced via doxycycline treatment at 1 µg/mL for 24 h or the duration indicated. A more detailed description of the pooled WT-OE cell line generation and quality control can be found here: https://zenodo.org/records/7457221 and https://zenodo.org/records/5566805.
"],"omics_type":["Metabolomics"],"study_design":["ultra-performance liquid chromatography-mass spectrometry","Transmembrane Transport","tandem mass spectrometry","analyte assay","Solute Carrier Family","targeted metabolites"],"curator_keywords":["ultra-performance liquid chromatography-mass spectrometry","Transmembrane Transport","tandem mass spectrometry","analyte assay","Solute Carrier Family","targeted metabolites"],"mass_spectrometry_protocol":["The triple quadrupole mass spectrometer was operated in electrospray ionization negative mode, spray voltage 2 kV, gas temperature 150 °C, gas flow 1.3 L/min, nebulizer 45 psi, sheath gas temperature 325 °C, and sheath gas flow 12 L/min. The metabolites of interest were detected using a dynamic Multiple Reaction Monitoring (dMRM) mode.
"],"metabolite_name":["Phenylpyruvic acid","Taurine","Hydroxyphenylacetic acid","Allantoin","Deoxyribose","Dihydroxyacetone phosphate","Pentahydroxyhexanoic acid","Riboflavin","Phospho-Serine","Deoxyinosine","Histidine","Hydroxynicotinic acid","Pentose","Ketobutyrate","Kynurenic acid","Ketovaleric acid","Dehydroshikimic acid","alpha-Ketoglutaric acid","Muconic acid","Folic Acid","Galactosamine","Taurocholic acid","Xylulose 5-phosphate","Guanosine cyclic monophosphate","Methoxytryptamine","Threonine","Guanidobutyric acid","Methionine","Inosine triphosphate","Citrulline","Deoxyadenosine","Hydroxyglutaric acid","Propionylcarnitine","Hydroxyethylindole","Hexose 1-phosphate","Lysine","Homocitrate","Nicotinamide adenine dinucleotide","O-Phosphorylethanolamine","AICAR","5 Methyl tetrahydrofolate","Uridine diphosphohexose","Glutamic acid","Deoxycytidine triphosphate","N-Acetyl galactosamine","Methylglutaric acid","Pyruvic acid","Inosine monophosphate","Butyrylcarnitine","Glycine","Guanine","Cystine","Mevalonic acid","Arginine","Cystathionine","Indoleacetic acid","Kynurenine","Cytidine triphosphate","Mevalonolactone","Pyridoxal 5 phosphate","Pyridoxamine","Glucosamine 6-phosphate","Creatinine","Dimethyl Succinic acid","Creatine phosphate","Dihydrofolate","N-Acetylglutamic acid","Salicylic acid","Alanine","Phosphoenolpyruvic acid","Phenylalanine","Deoxyuridine","Pyridoxic acid","Tryptophan","Xanthosine","Pyridinedicarboxylic acid","Pyridoxine","Xanthine","Prephenic acid","Dihydroxybenzoic acid","Guanosine triphosphate","Oxamic acid","Deoxycytidine monophosphate","Deoxyguanosine","Quinic acid","Valine","Asparagine","Lipoamide","Hexose 6-phosphate","Indoline-2-carboxylate","5,10 Methenyl tetrahydrofolate","N-Acetyl-glucosamine phosphate","Uracil","N-Acetylneuraminic acid","Pantothenic acid","Hexose","Isopentyl acetate","Cytidine monophosphate","Guanosine","Ribonic acid gamma lactone","Nicotinic acid","Thymidine","Aspartic Acid","Creatine","Hydroxyanthranilic acid","Shikimic acid","N-Carbamyl-glutamic acid","S-Adenosyl-homocysteine","Aminobenzoic acid","Itaconic acid","Orotic acid","Aminobutyric acid","Glutathione oxidized","Octanoylcarnitine","Glucoheptonic acid","Maleic acid","Malic acid","Tauro-muricholic acid","Vanillic acid","Cytosine","Glyceric acid","Hydroxy-indoleacetic acid","Flavin adenine dinucleotide","Deoxyribose phosphate","Adenosine cyclic monophosphate","Tyrosine","Deoxythymidine triphosphate","Deoxyuridine triphosphate","Hydroxyphenyl-pyruvic acid","Aminoadipic acid","Fructose 1,6-biphosphate","Dihydroxyisovalerate","Palmitoylcarnitine","Cytidine","Carnitine","Muricholic acid","Arabitol","Succinic acid","Uridine","Hypoxanthine","Adipic acid","Ribose 5-phosphate","Homoserine","Malonic acid","Uridine monophosphate","Hydroxy-glutamic acid","Canavanine","Hydroxy-kynurenine","Isovalerylcarnitine","Serine","Proline","Leucine","Hydroxy-proline","Citramalic acid","Epicatechin","Nicotinic acid mononucleotide","Hydroxybenzoic acid","Pyridoxal hydrochloride","Citric acid+Isocitric acid","cis-Aconitic acid","Adenosine triphosphate","Thiamine","Uridine triphosphate","Isoleucine","gamma-Glu-Cys","Adenosine","Adenosine monophosphate","Deoxyadenosine monophosphate","Sedoheptulose-7-phosphate","Deoxy-(methylthio)adenosine","Thymine","Adenine","Deoxyadenosine triphosphate","Uric acid","Aconitic acid","N-Formyl-Tyrosine","Xylitol","Deoxycytidine","Acetylcarnitine","Dihydroorotic acid","N-Carbamoyl-aspartic acid","Argininosuccinic acid","Inosine","Isopropylmalic acid","Phosphoglyceric acid","Lactic acid","Anthranilic acid","Homocystine","Glutamine"],"additional_accession":[]},"is_claimable":false,"name":"Metabolic mapping of the entire human solute carrier superfamily","description":"Solute carrier (SLC) transporters govern most of the chemical exchange across cellular membranes and are integral to metabolic regulation, which in turn is linked to cellular function and identity. Despite their key role, individual functions of the SLC superfamily members were not evaluated systematically. We determined the metabolic and transcriptional profiles upon SLC overexpression in knock-out or wild-type isogenic cell backgrounds for 378 SLCs and 441 SLCs, respectively. Targeted metabolomics provided a fingerprint of 189 intracellular metabolites, while transcriptomics offered insights into cellular programs modulated by SLC expression. Beyond the metabolic profiles of 102 SLCs directly related to their known substrates, we identified putative substrates or metabolic pathway connections for 71 SLCs without previously annotated bona fide substrates, including SLC45A4 as a new polyamine transporter. By comparing the molecular profiles, we identified functionally related SLC groups, including some with distinct impacts on osmolyte balancing and glycosylation. The assessment of functionally related human genes presented here may serve as a blueprint for other systematic studies and supports future investigations into the functional roles of SLCs.
","dates":{"publication":"2025-04-08","submission":"2024-04-29"},"accession":"MTBLS10077","cross_references":{"MetaboLights":["MTBLC15641","MTBLC12071","MTBLC73024","MTBLC32806","MTBLC16708","MTBLC16335","MTBLC17489","MTBLC16027","MTBLC15422","MTBLC30832","MTBLC28498","MTBLC16449","MTBLC15676","MTBLC30915","MTBLC37024","MTBLC22495","MTBLC16865","MTBLC30754","MTBLC22605","MTBLC29016","MTBLC15682","MTBLC22653","MTBLC22660","MTBLC7676","MTBLC609827","MTBLC17126","MTBLC32805","MTBLC15584","MTBLC30769","MTBLC18211","MTBLC16919","MTBLC17287","MTBLC16737","MTBLC17755","MTBLC17376","MTBLC17562","MTBLC17361","MTBLC17677","MTBLC16040","MTBLC30918","MTBLC17509","MTBLC17256","MTBLC17713","MTBLC16284","MTBLC15698","MTBLC15918","MTBLC16311","MTBLC17172","MTBLC28997","MTBLC28816","MTBLC19565","MTBLC18077","MTBLC16450","MTBLC17625","MTBLC15633","MTBLC30865","MTBLC16108","MTBLC18026","MTBLC15689","MTBLC86537","MTBLC23053","MTBLC16238","MTBLC27470","MTBLC16905","MTBLC33033","MTBLC60312","MTBLC17515","MTBLC67270","MTBLC47987","MTBLC18237","MTBLC28300","MTBLC17858","MTBLC33508","MTBLC15428","MTBLC86564","MTBLC16235","MTBLC16750","MTBLC16356","MTBLC15996","MTBLC18133","MTBLC24588","MTBLC47877","MTBLC27570","MTBLC17852","MTBLC17485","MTBLC30653","MTBLC15793","MTBLC24676","MTBLC17890","MTBLC32811","MTBLC17084","MTBLC27823","MTBLC1547","MTBLC16168","MTBLC17445","MTBLC15999","MTBLC18240","MTBLC17368","MTBLC16411","MTBLC167877","MTBLC17596","MTBLC17202","MTBLC16039","MTBLC24898","MTBLC31725","MTBLC28635","MTBLC73025","MTBLC30838","MTBLC30831","MTBLC28683","MTBLC18344","MTBLC78320","MTBLC25017","MTBLC17460","MTBLC25094","MTBLC18300","MTBLC6650","MTBLC30794","MTBLC16811","MTBLC2089","MTBLC68566","MTBLC25351","MTBLC67849","MTBLC38407","MTBLC134216","MTBLC28800","MTBLC15784","MTBLC17533","MTBLC17012","MTBLC64850","MTBLC71028","MTBLC50603","MTBLC44215","MTBLC15940","MTBLC15763","MTBLC73039","MTBLC17553","MTBLC16742","MTBLC18058","MTBLC73067","MTBLC7916","MTBLC24266","MTBLC25901","MTBLC28044","MTBLC30851","MTBLC44897","MTBLC24344","MTBLC37712","MTBLC16666","MTBLC26271","MTBLC28867","MTBLC16675","MTBLC18405","MTBLC17310","MTBLC16410","MTBLC17405","MTBLC16709","MTBLC32816","MTBLC17521","MTBLC17015","MTBLC74168","MTBLC52742","MTBLC16680","MTBLC16914","MTBLC15721","MTBLC17822","MTBLC16119","MTBLC15741","MTBLC15891","MTBLC28865","MTBLC133057","MTBLC18385","MTBLC26986","MTBLC17748","MTBLC17821","MTBLC27897","MTBLC18186","MTBLC17568","MTBLC27226","MTBLC16704","MTBLC18066","MTBLC16695","MTBLC15713","MTBLC27266","MTBLC30816","MTBLC15318","MTBLC18107","MTBLC17151","MTBLC27354"],"ChEBI":["CHEBI:15641","CHEBI:12071","CHEBI:73024","CHEBI:32806","CHEBI:16708","CHEBI:16335","CHEBI:17489","CHEBI:16027","CHEBI:15422","CHEBI:30832","CHEBI:28498","CHEBI:16449","CHEBI:15676","CHEBI:30915","CHEBI:37024","CHEBI:22495","CHEBI:16865","CHEBI:30754","CHEBI:22605","CHEBI:29016","CHEBI:15682","CHEBI:22653","CHEBI:22660","CHEBI:7676","CHEBI:609827","CHEBI:17126","CHEBI:32805","CHEBI:15584","CHEBI:30769","CHEBI:18211","CHEBI:16919","CHEBI:17287","CHEBI:16737","CHEBI:17755","CHEBI:17376","CHEBI:17562","CHEBI:17361","CHEBI:17677","CHEBI:16040","CHEBI:30918","CHEBI:17509","CHEBI:17256","CHEBI:17713","CHEBI:16284","CHEBI:15698","CHEBI:15918","CHEBI:16311","CHEBI:17172","CHEBI:28997","CHEBI:28816","CHEBI:19565","CHEBI:18077","CHEBI:16450","CHEBI:17625","CHEBI:15633","CHEBI:30865","CHEBI:16108","CHEBI:18026","CHEBI:15689","CHEBI:86537","CHEBI:23053","CHEBI:16238","CHEBI:27470","CHEBI:16905","CHEBI:33033","CHEBI:60312","CHEBI:17515","CHEBI:67270","CHEBI:47987","CHEBI:18237","CHEBI:28300","CHEBI:17858","CHEBI:33508","CHEBI:15428","CHEBI:86564","CHEBI:16235","CHEBI:16750","CHEBI:16356","CHEBI:15996","CHEBI:18133","CHEBI:24588","CHEBI:47877","CHEBI:27570","CHEBI:17852","CHEBI:17485","CHEBI:30653","CHEBI:15793","CHEBI:24676","CHEBI:17890","CHEBI:32811","CHEBI:17084","CHEBI:27823","CHEBI:1547","CHEBI:16168","CHEBI:17445","CHEBI:15999","CHEBI:18240","CHEBI:17368","CHEBI:16411","CHEBI:167877","CHEBI:17596","CHEBI:17202","CHEBI:16039","CHEBI:24898","CHEBI:31725","CHEBI:28635","CHEBI:73025","CHEBI:30838","CHEBI:30831","CHEBI:28683","CHEBI:18344","CHEBI:78320","CHEBI:25017","CHEBI:17460","CHEBI:25094","CHEBI:18300","CHEBI:6650","CHEBI:30794","CHEBI:16811","CHEBI:2089","CHEBI:68566","CHEBI:25351","CHEBI:67849","CHEBI:38407","CHEBI:134216","CHEBI:28800","CHEBI:15784","CHEBI:17533","CHEBI:17012","CHEBI:64850","CHEBI:71028","CHEBI:50603","CHEBI:44215","CHEBI:15940","CHEBI:15763","CHEBI:73039","CHEBI:17553","CHEBI:16742","CHEBI:18058","CHEBI:73067","CHEBI:7916","CHEBI:24266","CHEBI:25901","CHEBI:28044","CHEBI:30851","CHEBI:44897","CHEBI:24344","CHEBI:37712","CHEBI:16666","CHEBI:26271","CHEBI:28867","CHEBI:16675","CHEBI:18405","CHEBI:17310","CHEBI:16410","CHEBI:17405","CHEBI:16709","CHEBI:32816","CHEBI:17521","CHEBI:17015","CHEBI:74168","CHEBI:52742","CHEBI:16680","CHEBI:16914","CHEBI:15721","CHEBI:17822","CHEBI:16119","CHEBI:15741","CHEBI:15891","CHEBI:28865","CHEBI:133057","CHEBI:18385","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