Gain-of-function Genetic Screens in Human Cells Identify SLC Transporters Overcoming Environmental Nutrient Restrictions

Overview

Journal Life Sci Alliance

Specialties Biochemistry
Biology
Cell Biology
Molecular Biology

Date 2022 Sep 16

PMID 36114003

Authors

Manuele Rebsamen

Enrico Girardi

Vitaly Sedlyarov

Stefania Scorzoni

Konstantinos Papakostas

Manuela Vollert

Justyna Konecka

Bettina Guertl

Kristaps Klavins

Tabea Wiedmer

Giulio Superti-Furga

Affiliations

Soon will be listed here.

Abstract

Solute carrier (SLC) transporters control fluxes of nutrients and metabolites across membranes and thereby represent a critical interface between the microenvironment and cellular and subcellular metabolism. Because of substantial functional overlap, the interplay and relative contributions of SLCs in response to environmental stresses remain poorly elucidated. To infer functional relationships between SLCs and metabolites, we developed a strategy to identify SLCs able to sustain cell viability and proliferation under growth-limiting concentrations of essential nutrients. One-by-one depletion of 13 amino acids required for cell proliferation enabled gain-of-function genetic screens using a SLC-focused CRISPR/Cas9-based transcriptional activation approach to uncover transporters relieving cells from growth-limiting metabolic bottlenecks. Among the transporters identified, we characterized the cationic amino acid transporter as a gene that, when up-regulated, overcame low availability of arginine and lysine by increasing their uptake, whereas SLC7A5 was able to sustain cellular fitness upon deprivation of several neutral amino acids. Moreover, we identified metabolic compensation mediated by the glutamate/aspartate transporters SLC1A2 and SLC1A3 under glutamine-limiting conditions. Overall, this gain-of-function approach using human cells uncovered functional transporter-nutrient relationships and revealed that transport activity up-regulation may be sufficient to overcome environmental metabolic restrictions.

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References

Chapman N, Boothby M, Chi H . Metabolic coordination of T cell quiescence and activation. Nat Rev Immunol. 2019; 20(1):55-70. DOI: 10.1038/s41577-019-0203-y. View

Alkan H, Walter K, Luengo A, Madreiter-Sokolowski C, Stryeck S, Lau A . Cytosolic Aspartate Availability Determines Cell Survival When Glutamine Is Limiting. Cell Metab. 2018; 28(5):706-720.e6. PMC: 6390946. DOI: 10.1016/j.cmet.2018.07.021. View

Bacci M, Lorito N, Ippolito L, Ramazzotti M, Luti S, Romagnoli S . Reprogramming of Amino Acid Transporters to Support Aspartate and Glutamate Dependency Sustains Endocrine Resistance in Breast Cancer. Cell Rep. 2019; 28(1):104-118.e8. PMC: 6616584. DOI: 10.1016/j.celrep.2019.06.010. View

Palmieri F . The mitochondrial transporter family SLC25: identification, properties and physiopathology. Mol Aspects Med. 2012; 34(2-3):465-84. DOI: 10.1016/j.mam.2012.05.005. View

Hediger M, Clemencon B, Burrier R, Bruford E . The ABCs of membrane transporters in health and disease (SLC series): introduction. Mol Aspects Med. 2013; 34(2-3):95-107. PMC: 3853582. DOI: 10.1016/j.mam.2012.12.009. View

Hsu P, Lander E, Zhang F . Development and applications of CRISPR-Cas9 for genome engineering. Cell. 2014; 157(6):1262-1278. PMC: 4343198. DOI: 10.1016/j.cell.2014.05.010. View

Gilbert L, Horlbeck M, Adamson B, Villalta J, Chen Y, Whitehead E . Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation. Cell. 2014; 159(3):647-61. PMC: 4253859. DOI: 10.1016/j.cell.2014.09.029. View

Labuschagne C, van den Broek N, Mackay G, Vousden K, Maddocks O . Serine, but not glycine, supports one-carbon metabolism and proliferation of cancer cells. Cell Rep. 2014; 7(4):1248-58. DOI: 10.1016/j.celrep.2014.04.045. View

Thoreen C, Kang S, Chang J, Liu Q, Zhang J, Gao Y . An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem. 2009; 284(12):8023-32. PMC: 2658096. DOI: 10.1074/jbc.M900301200. View

10.

Meixner E, Goldmann U, Sedlyarov V, Scorzoni S, Rebsamen M, Girardi E . A substrate-based ontology for human solute carriers. Mol Syst Biol. 2020; 16(7):e9652. PMC: 7374931. DOI: 10.15252/msb.20209652. View

11.

Finicle B, Jayashankar V, Edinger A . Nutrient scavenging in cancer. Nat Rev Cancer. 2018; 18(10):619-633. DOI: 10.1038/s41568-018-0048-x. View

12.

Cormerais Y, Massard P, Vucetic M, Giuliano S, Tambutte E, Durivault J . The glutamine transporter ASCT2 (SLC1A5) promotes tumor growth independently of the amino acid transporter LAT1 (SLC7A5). J Biol Chem. 2018; 293(8):2877-2887. PMC: 5827425. DOI: 10.1074/jbc.RA117.001342. View

13.

van Geldermalsen M, Wang Q, Nagarajah R, Marshall A, Thoeng A, Gao D . ASCT2/SLC1A5 controls glutamine uptake and tumour growth in triple-negative basal-like breast cancer. Oncogene. 2015; 35(24):3201-8. PMC: 4914826. DOI: 10.1038/onc.2015.381. View

14.

Wellberg E, Johnson S, Finlay-Schultz J, Lewis A, Terrell K, Sartorius C . The glucose transporter GLUT1 is required for ErbB2-induced mammary tumorigenesis. Breast Cancer Res. 2016; 18(1):131. PMC: 5168867. DOI: 10.1186/s13058-016-0795-0. View

15.

Gardner D . Changes in requirements and utilization of nutrients during mammalian preimplantation embryo development and their significance in embryo culture. Theriogenology. 2000; 49(1):83-102. DOI: 10.1016/s0093-691x(97)00404-4. View

16.

Konermann S, Brigham M, Trevino A, Joung J, Abudayyeh O, Barcena C . Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex. Nature. 2014; 517(7536):583-8. PMC: 4420636. DOI: 10.1038/nature14136. View

17.

Birsoy K, Possemato R, Lorbeer F, Bayraktar E, Thiru P, Yucel B . Metabolic determinants of cancer cell sensitivity to glucose limitation and biguanides. Nature. 2014; 508(7494):108-12. PMC: 4012432. DOI: 10.1038/nature13110. View

18.

Cesar-Razquin A, Snijder B, Frappier-Brinton T, Isserlin R, Gyimesi G, Bai X . A Call for Systematic Research on Solute Carriers. Cell. 2015; 162(3):478-87. DOI: 10.1016/j.cell.2015.07.022. View

19.

Butler M, Van der Meer L, van Leeuwen F . Amino Acid Depletion Therapies: Starving Cancer Cells to Death. Trends Endocrinol Metab. 2021; 32(6):367-381. DOI: 10.1016/j.tem.2021.03.003. View

20.

Superti-Furga G, Lackner D, Wiedmer T, Ingles-Prieto A, Barbosa B, Girardi E . The RESOLUTE consortium: unlocking SLC transporters for drug discovery. Nat Rev Drug Discov. 2020; 19(7):429-430. DOI: 10.1038/d41573-020-00056-6. View