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Role of Acid Ceramidase in the Hydrolysis of Anti‐inflammatory and Anorexic N ‐Acylethanolamines
Author(s) -
Tsuboi Kazuhito,
Tai Tatsuya,
Yamashita Ryouhei,
Ali Hanif,
Watanabe Takashi,
Uyama Toru,
Okamoto Yoko,
Kitakaze Keisuke,
Takenouchi Yasuhiro,
Go Shinji,
Rahman Iffat,
Houchi Hitoshi,
Tanaka Tamotsu,
Okamoto Yasuo,
Tokumura Akira,
Matsuda Junko,
Ueda Natsuo
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.00235
Subject(s) - anandamide , fatty acid amide hydrolase , ceramide , chemistry , palmitoylethanolamide , biochemistry , lipid signaling , enzyme , sphingosine , microbiology and biotechnology , biology , receptor , apoptosis , cannabinoid receptor , agonist
N ‐Acylethanolamines (NAEs) constitute a class of lipid mediators and include palmitoylethanolamide, oleoylethanolamide and anandamide, which exert anti‐inflammatory/analgesic, anorexic, and cannabimimetic actions, respectively. Two enzymes, fatty acid amide hydrolase and lysosomal NAE acid amidase (NAAA), have been analyzed as NAE hydrolases. On the other hand, acid ceramidase (AC) is known as a lysosomal enzyme degrading ceramide ( N ‐acylsphingosine). AC is abundantly expressed in several human tumors, and its role in chemoresistance is suggested. Since AC shows a high similarity to NAAA in amino acid sequence as well as catalytic mechanism, in the present study we examined the role of AC in the hydrolysis of NAEs. First, we purified recombinant human AC and showed that the purified enzyme hydrolyzed various NAEs as well as ceramides. The most reactive NAE was lauroylethanolamide (C12:0‐NAE), which was consistent with the preference of C12:0‐ceramide by AC. We then metabolically labeled HEK293 cells with [ 14 C]ethanolamine. The overexpression of AC decreased the intracellular level of 14 C‐labeled NAEs. As analyzed by liquid chromatography‐tandem mass spectrometry, the overexpression of AC was found to lower the levels of different NAE species. Furthermore, the knockdown of endogenous AC by siRNA in human prostate LNCaP cells increased the levels of various NAE species. Finally, we analyzed the homogenates of brain and kidney from genetically engineered mice lacking saposin D, which was considered as an activator protein of AC. The results showed that the homogenates from the mutant mice showed much less hydrolyzing activity towards C12:0‐NAE as well as C12:0‐ceramide. These results strongly suggest a role of AC in the degradation of bioactive NAEs as a third NAE hydrolase.