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Bandhuvula, P., Tam, Y. Y., Oskouian, B. & Saba, J. D. The immune modulator FTY720 inhibits sphingosine-1-phosphate lyase activity. J. Biol. Chem. 280, 33697–33700. https://doi.org/10.1074/jbc.C500294200 (2005). The International Standards Organisation (ISO) standard, ISO 3601 comprehensively defines both the o-ring and hardware dimensions for rod, piston, face seal grooves. are usually free or discounted: Lawyer Referral & Information Service (LRIS) Committed to Public Service Human S1P lyase cDNA (SGPL1, Genbank Accession number NM_003901) was amplified from placenta cDNA and FLAG epitope-tagged at the 3′ end by PCR with oligonucleotide primers 5′-TATATAGAATTCGCCACCATGCCTAGCACAGACCTTCT-3′ and 5′-TATATAGAATTCACTTGTCATCGTCGTCCTTGTAGTCGTGGGGTTTTGGAGAACCAT-3′. The PCR product was digested with EcoRI and cloned into pcDNA3 (Invitrogen) for expression in mammalian cells. Sequencing verified the orientation and integrity of the cDNA. Quantitative RT-PCR Chen, M. H. et al. Identification of SPHK1 as a therapeutic target and marker of poor prognosis in cholangiocarcinoma. Oncotarget 6, 23594–23608. https://doi.org/10.18632/oncotarget.4335 (2015).

Roberts, J. L. et al. An assay for sphingosine kinase activity using biotinylated sphingosine and streptavidin-coated membranes. Anal. Biochem. 331, 122–129. https://doi.org/10.1016/j.ab.2004.03.030 (2004). Even in times of a national crisis, fraudsters will go to extraordinary lengths to divert funds away from the public sector. The Counter Fraud Functional Standard was launched in October 2018, and is being implemented across government. It applies to all government departments and their arms-length bodies. As of February 2020, 123 public bodies had adopted the functional standard as the basis for managing the risk of fraud, bribery and corruption in the public sector. Newton, J., Lima, S., Maceyka, M. & Spiegel, S. Revisiting the sphingolipid rheostat: Evolving concepts in cancer therapy. Exp. Cell Res. 333, 195–200. https://doi.org/10.1016/j.yexcr.2015.02.025 (2015). The publication of the Counter Fraud Functional Standard reinforces the government’s commitment to fighting fraud. By meeting the requirements of this standard, public bodies are better prepared, and more able to respond to the risk of fraud against the public sector.MV411 cells (1.5 × 10 7) were seeded at 1 × 10 6 per mL and treated with DMSO (0.1% final) or 10 µM JTE-013 for 6 h. Cells were pelleted by centrifugation, washed in PBS and snap frozen. The cell pellets were suspended in 1 mL of chilled PBS and centrifuged at 2000× g for 5 min at 4 °C. The supernatant was discarded and the remaining solid was resuspended in 450 µL of extraction mix [chloroform: methanol: H 2O, 2:6:1 (v/v)]. Odd chain lipid standards mix was added to give final concentrations of 241 nM sphingosine (d17:1), 250 nM dihydosphingosine (d17:0), 253 nM S1P (d17:1), 235 nM dihydrosphingosine 1-phosphate (d17:0), 250 nM sphingomyelin (d18:1/17:0), and 450 nM C17 ceramide (d18:1/17:0). The samples were frozen/thawed three times, then centrifuged at 14,800× g for 5 min at 4 °C. 400 µL of supernatant was then transferred to new tubes, with some remaining sample combined to make a pooled QC sample. To reconstitute, the solvent was evaporated to dryness keeping the samples in a centrifugal evaporator at 55 °C for 50 min. Dried extracts were frozen at – 80 °C until LC–MS analysis was performed. On the day of analysis, the samples were dissolved in 180 µL of butanol: methanol (v/v 1:1) mixture and 20 µL of water. The samples were vortexed on a rotary vortex for 10 min and sonicated in a sonicator bath for 1 h keeping the temperature below 25 °C. The samples were centrifuged at 14,800× g for 10 min at 20 °C and transferred to LC–MS vials. Cirillo, F. et al. The antithetic role of ceramide and sphingosine-1-phosphate in cardiac dysfunction. J. Cell Physiol. 236, 4857–4873. https://doi.org/10.1002/jcp.30235 (2021). Osada, M., Yatomi, Y., Ohmori, T., Ikeda, H. & Ozaki, Y. Enhancement of sphingosine 1-phosphate-induced migration of vascular endothelial cells and smooth muscle cells by an EDG-5 antagonist. Biochem. Biophys. Res. Commun. 299, 483–487. https://doi.org/10.1016/s0006-291x(02)02671-2 (2002).

McNaughton, M., Pitman, M., Pitson, S. M., Pyne, N. J. & Pyne, S. Proteasomal degradation of sphingosine kinase 1 and inhibition of dihydroceramide desaturase by the sphingosine kinase inhibitors, SKi or ABC294640, induces growth arrest in androgen-independent LNCaP-AI prostate cancer cells. Oncotarget 7, 16663–16675. https://doi.org/10.18632/oncotarget.7693 (2016). Saba, J. D. Fifty years of lyase and a moment of truth: Sphingosine phosphate lyase from discovery to disease. J. Lipid. Res. 60, 456–463. https://doi.org/10.1194/jlr.S091181 (2019). The standard was developed by counter fraud experts (including from across the public and private sectors, banking and academia) to help guide a whole of government approach. It has been extensively tested in government before its formal release, and represents the minimum that all public bodies are expected to have in place. Drexler, Y., Molina, J., Mitrofanova, A., Fornoni, A. & Merscher, S. Sphingosine-1-phosphate metabolism and signaling in kidney diseases. J. Am. Soc. Nephrol. 32, 9–31. https://doi.org/10.1681/ASN.2020050697 (2021).French, K. J. et al. Pharmacology and antitumor activity of ABC294640, a selective inhibitor of sphingosine kinase-2. J. Pharmacol. Exp. Ther. 333, 129–139. https://doi.org/10.1124/jpet.109.163444 (2010). Long, J. S. et al. Sphingosine 1-phosphate receptor 4 uses HER2 (ERBB2) to regulate extracellular signal regulated kinase-1/2 in MDA-MB-453 breast cancer cells. J. Biol. Chem. 285, 35957–35966. https://doi.org/10.1074/jbc.M110.117945 (2010). Powell, J. A. et al. Targeting sphingosine kinase 1 induces MCL1-dependent cell death in acute myeloid leukemia. Blood 129, 771–782. https://doi.org/10.1182/blood-2016-06-720433 (2017).