期刊
JOURNAL OF DIABETES INVESTIGATION
卷 13, 期 7, 页码 1134-1139出版社
WILEY
DOI: 10.1111/jdi.13800
关键词
Exocytosis; Glucagon-like peptide-1; Total internal reflection fluorescence microscope
资金
- MEXT [20H05872]
- JSPS [19KK0198, 21K1934701, 21H00413]
- AMED-CREST from AMED [JP21gm161000]
- Photo-excitonix Project in Hokkaido University
- Grants-in-Aid for Scientific Research [20H05872, 21H00413, 19KK0198] Funding Source: KAKEN
FP-tagged GLP-1 allows visualization of the secretion process of GLP-1 and displays high efficiency in exocytosis and colocalization with secretory vesicle markers. This tagging method facilitates the study of GLP-1 secretion mechanisms and the discovery of new antidiabetic drugs.
Live-cell imaging with fluorescent proteins (FPs) is a powerful tool for investigating the exocytosis processes of hormones. However, the secretion process of glucagon-like peptide-1 (GLP-1) has not been visualized by FPs, which might be because tagging FPs inhibits GLP-1 synthesis through the post-translational processing from proglucagon. Here, we have developed FP-tagged GLP-1 by inserting FPs into the middle of GLP-1 and adding the proglucagon signal peptide. Confocal imaging confirmed that GLP-1 fused to FPs with high folding efficiency showed granular structure, in which secretory vesicle markers colocalized. The fluorescence intensity of FP in the culture supernatant from cells treated with KCl or forskolin was significantly increased compared with those from untreated cells. Furthermore, FP-tagged GLP-1 enables direct visualization of stimulation-dependent exocytosis of GLP-1 at a single granule resolution with total internal reflection fluorescence microscopy. FP-tagged GLP-1 might facilitate the screening of GLP-1 secretagogues and the discovery of new antidiabetic drugs.
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