New methods for monitoring mitochondrial biogenesis and mitophagy in vitro and in vivo

Abstract

Removal of damaged mitochondria through mitophagy is critical for maintaining cellular homeostasis and functions. However, reliable quantitative assays to monitor mitophagy, particularly in vivo, are just emerging. This review will summarize the current novel quantitative assays to monitor mitophagy in vivo. AbstractRemoval of damaged mitochondria through mitophagy is critical for maintaining cellular homeostasis and functions. Increasing evidence implicates mitophagy in red blood cell differentiation, neurodegeneration, macrophage-mediated inflammation, ischemia, adipogenesis, drug-induced tissue injury, and cancer. Considerable progress has been made toward understanding the biochemical mechanisms involved in mitophagy regulation. However, few reliable assays to monitor and quantify mitophagy have been developed, particularly in vivo. In this review, we summarize the recent development of three assays, MitoTimer, mt-Keima and mito-QC, for monitoring and quantifying mitophagy in cells and in animal tissues. We also discuss the advantages and limitations of these three assays when using them to monitor and quantify mitophagy.

T-Cell Intracellular Antigens and Hu Antigen R Antagonistically Modulate Mitochondrial Activity and Dynamics by Regulating Optic Atrophy 1 Gene Expression

“…5E), consistent with previous results (29). We assessed mitochondrial dynamics in OPA1v5-expressing cells using the fluorescence marker mitochondrial Keima (mt-Keima) (37,38). OPA1v5 expression promoted mitochondrial clustering (clustering, 80%, versus network, 20%) compared to control cells (clustering, 5%, versus network, 95%) and remodeling around the perinuclear region (Fig.…”

“…Mitochondrial respiration is recognized as an indicator of mitochondrial health (2,38). We show that TIA1b and TIARb expression is associated with a deficiency in respiratory activity, whereas HuR enhances OXPHOS.…”

“…We therefore explored whether the mitochondrial dysfunction in TIA-expressing cells triggered mitophagy. To evaluate mitophagy flux, we used the mt-Keima fluorescence probe (37,38), which emits green fluorescence at 458 nm and at pH 7.0 in mitochondria and red fluorescence at 561 nm and at pH 4.0 in mitochondrion-lysosome fusions (37). TIA1b-and TIARbexpressing cells had a higher rate of mitophagy than GFP-and HuR-expressing cells, estimated as normalized red/green mt-Keima signals (Fig.…”

Alzheimer’s disease-relevant tau modifications selectively impact neurodegeneration and mitophagy in a novelC. eleganssingle-copy transgenic model

“…To investigate whether a causal relationship exists between tau PTMs and mitochondrial morphology, we examined the mitochondrial network in the PLM cell bodies at day 3 and at day 10 of adulthood in our tau PTM mutant models. Touch cell mitochondria were labeled with mito-mKeima, a pH-sensitive fluorescent biosensor (56, 57). Mito-mKeima can be used as a dual excitation ratiometric mitophagy reporter, as we expand upon below (Fig.…”

“…A strain containing mito-mKeima (56, 57) expressed specifically in touch cells was used for assay. Animals were mounted on 2% agarose pads on glass slides and immobilized with 1 mM tetramisole hydrochloride before imaging.…”