Monitoring mitophagy in yeast: the Om45-GFP processing assay.
Macroautophagy (hereafter autophagy) is a ubiquitous degradative process in eukaryotic cells. Mitochondria autophagy (mitophagy) is a type of specific autophagy that degrades mitochondria selectively. Mitophagy is thought to play an important role for maintaining the quality of these organelles by eliminating damaged mitochondria, and it is involved in cellular differentiation, whereas dysfunctional mitophagy is related with neurodegenerative diseases; however, the mechanism of mitophagy is poorly understood. To facilitate the analysis of mitophagy, we recently established a simple method to monitor mitophagy in yeast, the Om45-GFP processing assay. Om45-GFP is a mitochondrial outer membrane protein. Following the uptake of mitochondria into the vacuole, Om45-GFP is degraded, releasing the intact form of GFP, which is detected by immunoblotting. Therefore, the amount of free GFP reflects the level of mitophagy.
Kanki
Tomotake
T
Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Graduate School of Medical Sciences, Fukuoka, Japan. kanki@cclm.med.kyushu-u.ac.jp
Kang
Dongchon
D
Klionsky
Daniel J
DJ
D001343:Autophagy; D001681:Biological Assay / Q000379:methods*; D015153:Blotting, Western; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
D008565:Membrane Proteins; D024101:Mitochondrial Proteins; C547170:Om45-GFP protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins