title: Glycine N-acyltransferase-like 3 is responsible for long-chain N-acylglycine formation in N18TG2 cells.
abstract: Long-chain fatty acid amides are signaling lipids found in mammals and other organisms; however, details of the metabolic pathways for the N-acylglycines and primary fatty acid amides (PFAMs) have remained elusive. Heavy-labeled precursor and subtraction lipidomic experiments in mouse neuroblastoma N18TG2 cells, a model cell line for the study of fatty acid amide metabolism, establish the biosynthetic pathways for the N-acylglycines and the PFAMs. We provide evidence that the N-acylglycines are formed by a long-chain specific glycine-conjugating enzyme, glycine N-acyltransferase-like 3 (GLYATL3). siRNA knockdown of GLYATL3 in the N18TG2 cells resulted in a decrease in the levels of the N-acylglycines and the PFAMs. This is the first report of an enzyme responsible for long-chain N-acylglycine production in cellula. The production of the PFAMs in N18TG2 cells was reported to occur by the oxidative cleavage of the N-acylglycines, as catalyzed by peptidylglycine α-amidating monooxygenase (PAM). siRNA knockdown of PAM resulted in an accumulation of [(13)C18]N-oleoylglycine and decreased levels of [(13)C18]oleamide when the N18TG2 cells were grown in the presence of [(13)C18]oleic acid. The addition of [1-(13)C]palmitate to the N18TG2 cell growth media led to the production of a family of [1-(13)C]palmitoylated fatty acid amides, consistent with the biosynthetic pathways detailed herein.
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Kristen A
authors.initials: KA
authors.lastname: Jeffries
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Dempsey
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Emma K
authors.initials: EK
authors.lastname: Farrell
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Ryan L
authors.initials: RL
authors.lastname: Anderson
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Gabrielle J
authors.initials: GJ
authors.lastname: Garbade
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Tatyana S
authors.initials: TS
authors.lastname: Gurina
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Imran
authors.initials: I
authors.lastname: Gruhonjic
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620.
authors.forename: Carly A
authors.initials: CA
authors.lastname: Gunderson
authors.affiliation: Department of Chemistry, University of South Florida, Tampa, FL 33620 merkler@usf.edu.
authors.forename: David J
authors.identifier: 0000-0001-6659-0005
authors.initials: DJ
authors.lastname: Merkler
mesh_terms: D000217:Acyltransferases / Q000502:physiology*; D000577:Amides / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D005227:Fatty Acids / Q000096:biosynthesis*; D055785:Gene Knockdown Techniques; D050155:Lipogenesis; D051379:Mice
chemical_list: D000577:Amides; D005227:Fatty Acids; D000217:Acyltransferases; C000622213:Glyatl3 protein, mouse
keywords: N-acylamide; arachidonic acid; brain lipids; eicosanoids; lipids; mass spectrometry; neuroblastoma cells; oleamide; palmitoylation; siRNA knockdown