A set of gene knockouts as a resource for global lipidomic changes

Abstract

Enzyme specificity in lipid metabolic pathways often remains unresolved at the lipid species level, which is needed to link lipidomic molecular phenotypes with their protein counterparts to construct functional pathway maps. We created lipidomic profiles of 23 gene knockouts in a proof-of-concept study based on a CRISPR/Cas9 knockout screen in mammalian cells. This results in a lipidomic resource across 24 lipid classes. We highlight lipid species phenotypes of multiple knockout cell lines compared to a control, created by targeting the human safe-harbor locus AAVS1 using up to 1228 lipid species and subspecies, charting lipid metabolism at the molecular level. Lipid species changes are found in all knockout cell lines, however, some are most apparent on the lipid class level (e.g., SGMS1 and CEPT1), while others are most apparent on the fatty acid level (e.g., DECR2 and ACOT7). We find lipidomic phenotypes to be reproducible across different clones of the same knockout and we observed similar phenotypes when two enzymes that catalyze subsequent steps of the long-chain fatty acid elongation cycle were targeted.

Peroxisomal β-oxidation enzyme, DECR2, regulates lipid metabolism and promotes treatment resistance in advanced prostate cancer

“…In a very recent study, Spiegel et al characterised the lipidomic changes of a set of gene knockouts, including DECR2, in a colon cancer cell line. The authors observed elevated levels of long-chain PUFAs in PI, PE-O and PC-O lipid classes (C20 and C22), confirming previous studies that DECR2 can oxidise unsaturated fatty acids and assist in the degradation of PUFAs such as arachidonic acid (AHA) and DHA in peroxisomes 45 .…”

“…Although upon examination of the crystal structure of DECR2 showed selectivity for VLCFAs like docosahexaenoic acid (DHA) 43 , another study reported that DECR2 may be involved in the degradation of short and medium chain substrates 44 . In a very recent study, Spiegel et al and DHA in peroxisomes 45 .…”

Peroxisomal β-oxidation enzyme, DECR2, regulates lipid metabolism and promotes treatment resistance in advanced prostate cancer

“…In a very recent study, Spiegel et al characterised the lipidomic changes of a set of gene knockouts, including DECR2, in a colon cancer cell line. The authors observed elevated levels of long-chain PUFAs in PI, PE-O and PC-O lipid classes (C20 and C22), confirming previous studies that DECR2 can oxidise unsaturated fatty acids and assist in the degradation of PUFAs such as arachidonic acid (AHA) and DHA in peroxisomes [ 37 ]. Accordingly, our data demonstrated that DECR2 knockdown and TDZ treatment increased the levels of several acylcarnitine species in cells supplemented with DHA, indicating inhibition of perFAO.…”

iLipidome: enhancing statistical power and interpretability using hidden biosynthetic interdependencies in the lipidome

“…LPCAT3 has LPEAT and LPCAT activity, while AGPAT3 (LPAAT) catalyzes LPA reactions 33 . To further examine the general applicability of our method, a dataset containing multiple gene knockouts in lipid metabolic pathways was also used to assess the performance of iLipdiome 57 . Overall, the top 5 perturbed pathways across 11 gene knockouts revealed by iLipidome were highly consistent with the enzyme specificities and the expected lipidomic changes affected by each knockout ( Extended Data Table 1 ).…”