O-glycosylation of mucin-like domain retains the neutral ceramidase on the plasma membranes as a type II integral membrane protein

Motohiro Tani, Hiroshi Iida, Makoto Ito

研究成果: ジャーナルへの寄稿学術誌査読

56 被引用数 (Scopus)

抄録

Ceramidase is a key enzyme involved in regulating cellular levels of ceramide, sphingosine, and possibly sphigosine 1-phosphate and thus could modulate sphingolipid signaling. Here we report that O-glycosylation of the mucin-like domain of neutral ceramidases was required for localization to the surface of plasma membranes. The deduced amino acid sequences of the mammalian enzymes contain a serine-threonine-rich domain (mucin box), which follows the signal/anchor sequence, whereas those of bacterial and invertebrate enzymes completely lack a mucin box, suggesting that the specific domain has been acquired during evolution. In HEK293 cells overexpressing ceramidase, the enzyme was not only secreted into the medium after cleavage of the NH2-terminal signal/anchor sequence but also localized at the plasma membrane as a type II integral membrane protein. Lectin blot analysis using peanut agglutinin revealed that the mucin box of the enzyme is highly glycosylated with O-glycans. Interestingly, a mutant lacking the mucin box or possible O-glycosylation sites in the mucin box was secreted into the medium but not localized at the surface of the cells. Furthermore, a mucin box-fused chimera green fluorescent protein (GFP), but not GFP itself, with the signal/anchor sequence was distributed on the surface of the cells. These results suggest that O-glycosylation of the mucin box retains proteins on the plasma membranes. We also found that the 112-kDa membrane-bound enzyme from mouse kidney is O-glycosylated, whereas the 94-kDa soluble enzyme from liver is not. These results clearly indicate that post-translational modification of the enzyme with O-glycans is tissue-specific and helps the enzyme to localize at the surface of plasma membranes as a type II membrane protein.

本文言語英語
ページ(範囲)10523-10530
ページ数8
ジャーナルJournal of Biological Chemistry
278
12
DOI
出版ステータス出版済み - 3月 21 2003

!!!All Science Journal Classification (ASJC) codes

  • 生化学
  • 分子生物学
  • 細胞生物学

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