| RRC ID |
50105
|
| 著者 |
Katayama H, Kogure T, Mizushima N, Yoshimori T, Miyawaki A.
|
| タイトル |
A sensitive and quantitative technique for detecting autophagic events based on lysosomal delivery.
|
| ジャーナル |
Chem Biol
|
| Abstract |
We sought to develop a sensitive and quantitative technique capable of monitoring the entire flux of autophagy involving fusion of lysosomal membranes. We observed the accumulation inside lysosomal compartments of Keima, a coral-derived acid-stable fluorescent protein that emits different-colored signals at acidic and neutral pHs. The cumulative fluorescent readout can be used to quantify autophagy at a single time point. Remarkably, the technique led us to characterize an autophagy pathway in Atg5-deficient cells, in which conventional LC3-based autophagosome probes are ineffective. Due to the large Stokes shift of Keima, this autophagy probe can be visualized in conjunction with other green-emitting fluorophores. We examined mitophagy as a selective autophagic process; time-lapse imaging of mitochondria-targeted Keima and GFP-Parkin allowed us to observe simultaneously Parkin recruitment to and autophagic degradation of mitochondria after membrane depolarization.
|
| 巻・号 |
18(8)
|
| ページ |
1042-52
|
| 公開日 |
2011-8-26
|
| DOI |
10.1016/j.chembiol.2011.05.013
|
| PII |
S1074-5521(11)00204-3
|
| PMID |
21867919
|
| MeSH |
Animals
Anthozoa / chemistry
Anthozoa / genetics
Autophagy*
Autophagy-Related Protein 5
Cell Line
Gene Deletion
Green Fluorescent Proteins / analysis
Green Fluorescent Proteins / genetics
Luminescent Proteins / analysis*
Luminescent Proteins / genetics
Lysosomes / metabolism*
Mice
Microscopy, Confocal / methods*
Microtubule-Associated Proteins / genetics
Mitochondria / metabolism
Mutagenesis, Site-Directed
Recombinant Fusion Proteins / analysis
Recombinant Fusion Proteins / genetics
|
| IF |
5.915
|
| 引用数 |
237
|
| リソース情報 |
| 遺伝子材料 |
mKeima (h)/pCS2 (RDB15236)
dKeima (h)/pCS2 (RDB15237)
mt-mKeima (h)/pIND(SP1) (RDB15238)
mt-mKeima/pcDNA3 (RDB15456). |
