DEGS1-associated aberrant sphingolipid metabolism impairs nervous system function in humans

Abstract

Volume 129 Number 3 March 2019 conjugation of L-serine and palmitoyl-CoA, the rate-limiting step catalyzed by serine palmitoyltransferase (SPT). The immediate product 3-keto-sphinganine is reduced to sphinganine (SA), which is then N-acylated to dihydroceramide (dhCer) by 1 of 6 ceramide synthase isoforms (CerS1-6) (4). In the final step, dhCer is converted to ceramide by the insertion of a Δ4,5 trans (Δ4E) double bond into the SA backbone. This final conversion is catalyzed by the Δ4-dihydroceramide desaturase DEGS1 (5). On the catabolic side, ceramides are deacylated by ceramidases to form sphingosine (SO), which can be either recycled back to ceramides (sal-BACKGROUND. Sphingolipids are important components of cellular membranes and functionally associated with fundamental processes such as cell differentiation, neuronal signaling, and myelin sheath formation. Defects in the synthesis or degradation of sphingolipids leads to various neurological pathologies; however, the entire spectrum of sphingolipid metabolism disorders remains elusive. METHODS.A combined approach of genomics and lipidomics was applied to identify and characterize a human sphingolipid metabolism disorder. RESULTS.By whole-exome sequencing in a patient with a multisystem neurological disorder of both the central and peripheral nervous systems, we identified a homozygous p.Ala280Val variant in DEGS1, which catalyzes the last step in the ceramide synthesis pathway. The blood sphingolipid profile in the patient showed a significant increase in dihydro sphingolipid species that was further recapitulated in patient-derived fibroblasts, in CRISPR/Cas9-derived DEGS1-knockout cells, and by pharmacological inhibition of DEGS1. The enzymatic activity in patient fibroblasts was reduced by 80% compared with wild-type cells, which was in line with a reduced expression of mutant DEGS1 protein. Moreover, an atypical and potentially neurotoxic sphingosine isomer was identified in patient plasma and in cells expressing mutant DEGS1. CONCLUSION.We report DEGS1 dysfunction as the cause of a sphingolipid disorder with hypomyelination and degeneration of both the central and peripheral nervous systems.(OMIM #617575) (19-21), but also with axonal peripheral neuropathy without renal or adrenal deficiencies (22).Here, we identify DEGS1 dysfunction as the cause of an SL disorder with leukodystrophy and hypomyelination of the peripheral nervous system.

Biosynthesis of the anti‐lipid‐microdomain sphingoid base 4,14‐sphingadiene by the ceramide desaturase FADS3

“…D, The FADS3-catalyzed reaction only between C14 and C15 is detected in healthy individuals, whereas it is found in patients with a DEGS1 mutation. 51 We, therefore, speculate that C4 desaturation reaction by DEGS1 occurs earlier than the C14 desaturation reaction by FADS3 and that under physiological conditions ceramide synthesis proceeds in the order DHS ceramides, SPH ceramides, and finally SPD ceramides.…”

FADS3 is a Δ14Z sphingoid base desaturase that contributes to gender differences in the human plasma sphingolipidome

“…Enzyme activity was tested in human and mouse FADS3expressing cells supplemented either with isotope-labeled (d7)d18:0 or (d3)m18:0. The free LCBs were absorbed by the cells (13) and metabolized to (d7)ceramide and downstream products, such as (d7)SM and (d7)HexCer (Fig. S2A).…”

“…We recently reported that DEGS1 deficiency causes leukodystrophy and peripheral hypomyelination, which was associated with a pathologically increased formation of saturated dihydroSL species (13) and the formation of an atypical monounsaturated d18:1 isomer. Further analysis showed that this isomer contained a ⌬14Z DB but lacked the canonical ⌬4E DB.…”

“…Lipidomics analysis was performed as described previously (13,28) using the internal standards (100 pmol/ml, Avanti Polar Lipids): (d5) 1-desoxymethylsphinganine (m17:0), dihydroceramide (d18:0:12:0), ceramide (d18:1/12:0), 1-deoxydihydroceramide (m18:0/12:0), 1-deoxyceramide (m18:1/12:0), glucosylceramide (d18:1/8:0), and sphingomyelin (d18:1/12:0).…”

Long-read sequencing in human genetics

“…This unique feature of TGS enables SV, SNV, and the methylation status of genomic loci to be analyzed in parallel and may improve the molecular diagnostics, for example, of cancer and imprinting disorders. Not only the landscape of alternative splicing can be investigated by reading through entire isoforms [33], but the various base modifications present on native RNA molecules can also be detected using this PCR-free method [18]. Moreover, native CpG methylation and chromatin accessibility can be studied in parallel using long reads [38].…”