PNPLA1 is a transacylase essential for the generation of the skin barrier lipid ω-O-acylceramide

Abstract

Lipids are the primary components of the skin permeability barrier, which is the body's most powerful defensive mechanism against pathogens. Acylceramide (ω-O-acylceramide) is a specialized lipid essential for skin barrier formation. Here, we identify PNPLA1 as the long-sought gene involved in the final step of acylceramide synthesis, esterification of ω-hydroxyceramide with linoleic acid, by cell-based assays. We show that increasing triglyceride levels by overproduction of the diacylglycerol acyltransferase DGAT2 stimulates acylceramide production, suggesting that triglyceride may act as a linoleic acid donor. Indeed, the in vitro analyses confirm that PNPLA1 catalyses acylceramide synthesis using triglyceride as a substrate. Mutant forms of PNPLA1 found in patients with ichthyosis exhibit reduced or no enzyme activity in either cell-based or in vitro assays. Altogether, our results indicate that PNPLA1 is directly involved in acylceramide synthesis as a transacylase, and provide important insights into the molecular mechanisms of skin barrier formation and of ichthyosis pathogenesis.

PNPLA1 has a crucial role in skin barrier function by directing acylceramide biosynthesis

“…Our genetic approach using knockout mice and the biochemical approach by Ohno et al 49. complement each other by providing different lines of evidence that prove that PNPLA1 catalyses the ω- O -esterification in acylceramide biosynthesis.…”

“…The corresponding accumulation of putative precursors of these three lipid groups, namely ω-OH Cer, ω-OH GlcCer and ω-OH ULCFA, in PNPLA1-deficient epidermis provides strong evidence that PNPLA1 acts as an ω- O -acyltransferase or transacylase required for acylceramide synthesis. In this regard, the accompaning study by Ohno et al 49. has clearly shown that exogenous overexpression of PNPLA1 in cells or PNPLA1-reconstituted proteoliposomes promotes acylceramide formation likely as a transacylase and that PNPLA1 mutations associated with ARCI inactivate this transacylase activity.…”

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

“…Fragment (product) ions were detected in the scanning range m/z 100‐600. In MRM, ceramides were detected by setting the Q1 m/z value at [M + H − H 2 O] + for the precursor ions and the Q3 m/z value as that specific to SPH ( m/z 264.3) or SPD‐specific fragment ions ( m/z 262.3), using the appropriate collision energies (Tables ), as described previously . Data analyses were performed using MassLynx software (Waters).…”

Reduced chain length in myelin sphingolipids and poorer motor coordination in mice deficient in the fatty acid elongaseElovl1

“…Lipids corresponding to 5 µg of brain tissue [for GalCer, 2‐hydroxy (2‐OH) GalCer, sulfatide, and ceramide] or 0.5 µg of brain tissue (for sphingomyelin) were subjected to liquid chromatography (LC)–tandem mass spectrometry (MS/MS) analysis using an ultra‐performance LC–triple‐quadrupole mass spectrometry system (Xevo TQ‐S; Waters), as described previously . Lipids were resolved on a reverse‐phase column (ACQUITY UPLC CSH C18 column; 1.7 µm particle size, 2.1 × 100 mm; Waters), ionized by electrospray ionization in positive ion mode, and detected using multiple reaction monitoring by selecting the specific m/z at quadrupole mass filters Q1 and Q3 and using the appropriate collision energy for each GalCer, 2‐OH GalCer, sulfatide, sphingomyelin, and ceramide species (Tables ).…”