1. Regulation of N-glycosylation and secretion of Isthmin-1 by its C-mannosylation.
- Author
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Yoshimoto, Satoshi, Katayama, Kazuhiro, Suzuki, Takehiro, Dohmae, Naoshi, and Simizu, Siro
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CONFOCAL fluorescence microscopy , *SECRETION , *CELL lines - Abstract
C -mannosylation is a type of protein glycosylation. Human Isthmin-1 (ISM1) is a 52-kDa secreted protein with a thrombospondin type 1 repeat (TSR) domain, containing two consensus C -mannosylation sequences at Trp223 and Trp226. In this study, we sought to examine the role of C -mannosylation in the secretion of ISM1. We established and cultured an ISM1-overexpressing HT1080 cell line and purified recombinant ISM1 for analysis from the conditioned medium by LC-MS/MS. Subcellular localization of ISM1 was observed by confocal fluorescence microscopy. We found that ISM1 is C -mannosylated at Trp223 and Trp226 in the TSR domain. To determine the functions of the C -mannosylation of ISM1, we established a C -mannosylation-defective mutant ISM1-overexpressing HT1080 cell line and measured its secretion of ISM1. The secretion of ISM1 decreased significantly in this mutant ISM1-overexpressing line compared with wild-type cells. Furthermore, ISM1 was N -glycosylated only in these C -mannosylation-defective cells. ISM1 is C -mannosylated in its TSR domain, and the status of the C -mannosylation of ISM1 affects its N -glycosylation. The C -mannosylation of ISM1 regulates its N -glycosylation status. • Isthmin-1 (ISM1) is C -mannosylated at Trp223 and Trp226. • C -mannosylation of ISM1 regulates its secretion and subcellular localization. • N -glycosylation of ISM1 is regulated by its C -mannosylation. [ABSTRACT FROM AUTHOR]
- Published
- 2021
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