Bifunctional DEGS2 has higher hydroxylase activity toward substrates with very-long-chain fatty acids in the production of phytosphingosine-ceramides

Journal home page for Journal of Biological ChemistryAuthor links open overlay panel, , , , , , , Abstract

Phytosphingosine (PHS) is a sphingolipid component present mainly in epithelial tissues, including epidermis and those lining the digestive tract. DEGS2 is a bifunctional enzyme that produces ceramides (CERs) containing PHS (PHS-CERs) via hydroxylation and sphingosine-CERs via desaturation, using dihydrosphingosine-CERs as substrates. Until now, the role of DEGS2 in permeability barrier functioning, its contribution to PHS-CER production, and the mechanism that differentiates between these two activities have been unknown. Here, we analyzed the barrier functioning of the epidermis, esophagus, and anterior stomach of Degs2 knockout (KO) mice and found that there were no differences between Degs2 KO and wild type (WT) mice, indicating normal permeability barriers in the KO mice. In the epidermis, esophagus, and anterior stomach of Degs2 KO mice, PHS-CER levels were greatly reduced relative to WT mice, but PHS-CERs were still present. We obtained similar results for DEGS2 KO human keratinocytes. These results indicate that although DEGS2 plays a major role in PHS-CER production, another synthesis pathway exists as well. Next, we examined the fatty acid (FA) composition of PHS-CERs in various mouse tissues and found that PHS-CER species containing very-long-chain FAs (≥C21) were more abundant than those containing long-chain FAs (C11–C20). A cell-based assay system revealed that the desaturase and hydroxylase activities of DEGS2 toward substrates with different FA chain lengths differed, and that its hydroxylase activity was higher toward substrates containing very-long-chain FAs. Collectively, our findings contribute to the elucidation of the molecular mechanism of PHS-CER production.

Keywords

bifunctional enzyme

ceramide

enzyme mechanism

epidermis

lipid

permeability

sphingolipid

AbbreviationsHexCER

monohexosylceramide

LCFA

long-chain fatty acid

MRM

multiple reaction monitoring

MS/MS

tandem mass spectrometry

SPT

serine palmitoyltransferase

TEWL

transepidermal water loss

VLCFA

very-long-chain fatty acid

© 2023 THE AUTHORS. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology.

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