RRC ID 63076
著者 Kasai K, Kuroda Y, Takabuchi Y, Nitta A, Kobayashi T, Nozaka H, Miura T, Nakamura T.
タイトル Phosphorylation of Thr328 in hyaluronan synthase 2 is essential for hyaluronan synthesis.
ジャーナル Biochem Biophys Res Commun
Abstract Hyaluronan synthase 2 (HAS2) is an integral membrane protein composed of multi-membrane-spanning regions and a large intracellular loop (HAS2-loop). We previously examined the effect of phorbol 12-myristate 13-acetate (PMA) and/or 4-methylumbelliferone (4-MU) on the synthesis of hyaluronan (HA) in human skin fibroblasts and found that TPA and 4-MU have opposing effects on HA synthesis by phosphorylation and O-linked β-N-acetylglucosaminylation of HAS2, respectively. In this study, we constructed an expression vector for the HAS2-loop and analyzed its post-translational modification by PMA and 4-MU using mass spectrometry. We identified a phosphorylation site at the position corresponding to the Thr328 position of full-length HAS2, which was detected in the cells regardless of the presence of PMA or 4-MU. We next prepared T328A site-directed mutagenesis construct-transfected cells and investigated HA synthesis. The amount of HA was increased in cells expressing full-length HAS2 compared to in mock cells, whereas the amount of HA synthesized by cells transfected with the T328A site-directed mutagenesis construct was the same as that in mock cells. This phosphorylation site corresponded with the casein kinase 1 substrate motif. These results suggest that Thr328 phosphorylation is an essential factor for HA synthesis by HAS2 and the role of HAS2-loop may be useful in analyzing the regulation of HAS2 synthesis in physiological and pathological conditions.
巻・号 533(4)
ページ 732-738
公開日 2020-12-17
DOI 10.1016/j.bbrc.2020.08.093
PII S0006-291X(20)31693-4
PMID 32993960
MeSH Animals COS Cells Casein Kinase I / genetics Chlorocebus aethiops HEK293 Cells Humans Hyaluronan Synthases / genetics Hyaluronan Synthases / metabolism* Hyaluronic Acid / biosynthesis* Hymecromone / pharmacology* Mass Spectrometry Mutagenesis, Site-Directed Phosphorylation Point Mutation Protein Processing, Post-Translational Tetradecanoylphorbol Acetate / pharmacology* Up-Regulation
IF 2.985
リソース情報
ヒト・動物細胞 COS-7(RCB0539)