Mitophagy Selectively Degrades Individual Damaged Mitochondria After Photoirradiation

Center for Cell Death, Injury, and Regeneration, Department of Pharmaceutical and Biomedical Sciences, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Antioxidants & Redox Signaling (Impact Factor: 7.41). 12/2010; 14(10):1919-28. DOI: 10.1089/ars.2010.3768
Source: PubMed


Damaged and dysfunctional mitochondria are proposed to be removed by autophagy. However, selective degradation of damaged mitochondria by autophagy (mitophagy) has yet to be experimentally verified. In this study, we investigated the cellular fate of individual mitochondria damaged by photoirradiation in hepatocytes isolated from transgenic mice expressing green fluorescent protein fused to microtubule-associated protein 1 light chain 3, a marker of forming and newly formed autophagosomes. Photoirradiation with 488-nm light induced mitochondrial depolarization (release of tetramethylrhodamine methylester [TMRM]) in a dose-dependent fashion. At lower doses of light, mitochondria depolarized transiently with re-polarization within 3 min. With greater light, mitochondrial depolarization became irreversible. Irreversible, but not reversible, photodamage induced autophagosome formation after 32±5 min. Photodamage-induced mitophagy was independent of TMRM, as photodamage also induced mitophagy in the absence of TMRM. Photoirradiation with 543-nm light did not induce mitophagy. As revealed by uptake of LysoTracker Red, mitochondria weakly acidified after photodamage before a much stronger acidification after autophagosome formation. Photodamage-induced mitophagy was not blocked by phosphatidylinositol 3-kinase inhibition with 3-methyladenine (10 mM) or wortmannin (100 nM). In conclusion, individual damaged mitochondria become selectively degraded by mitophagy, but photodamage-induced mitophagic sequestration occurs independently of the phosphatidylinositol 3-kinase signaling pathway, the classical upstream signaling pathway of nutrient deprivation-induced autophagy.

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    • "The study of mitochondrial quality control in neurons has been hampered by the lack of methods to induce levels of mi­ tochondrial damage that are likely to approximate the extent of damage that might normally arise in a neuron. Other stud­ ies had previously used activation of mt­KR (Yang and Yang, 2011; Wang et al., 2012; Ertürk et al., 2014) or irradiation without a photosensitizer (Kim and Lemasters, 2011) to trigger mitophagy in nonneuronal cells. However, such methods were not applied to the sparsely distributed mitochondria in neuronal processes. "
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    ABSTRACT: To minimize oxidative damage to the cell, malfunctioning mitochondria need to be removed by mitophagy. In neuronal axons, mitochondrial damage may occur in distal regions, far from the soma where most lysosomal degradation is thought to occur. In this paper, we report that PINK1 and Parkin, two Parkinson's disease-associated proteins, mediate local mitophagy of dysfunctional mitochondria in neuronal axons. To reduce cytotoxicity and mimic physiological levels of mitochondrial damage, we selectively damaged a subset of mitochondria in hippocampal axons. Parkin was rapidly recruited to damaged mitochondria in axons followed by formation of LC3-positive autophagosomes and LAMP1-positive lysosomes. In PINK1(-/-) axons, damaged mitochondria did not accumulate Parkin and failed to be engulfed in autophagosomes. Similarly, initiation of mitophagy was blocked in Parkin(-/-) axons. Our findings demonstrate that the PINK1-Parkin-mediated pathway is required for local mitophagy in distal axons in response to focal damage. Local mitophagy likely provides rapid neuroprotection against oxidative stress without a requirement for retrograde transport to the soma.
    The Journal of Cell Biology 08/2014; 206(5). DOI:10.1083/jcb.201401070 · 9.83 Impact Factor
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    • "Mitophagy plays a role in segregating and degrading dysfunctional mitochondria that might otherwise release some adverse factor such as reactive oxygen species, pro-apoptotic proteins, or other toxic mediators. Studies show that factors released from the mitochondrial intermembranous space may provide the signal that not only leads to a degradation of individual mitochondria, but also stimulate autophagy [28], [29]. We showed that cell apoptosis ratios were increased in the mineral oil treatment group and that HO-1 siRNA treatment before IR increases the rate of apoptosis compared with the IR group (P<0.05). "
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    ABSTRACT: Background Growing evidence has linked autophagy to a protective role of preconditioning in liver ischemia/reperfusion (IR). Heme oxygenase-1 (HO-1) is essential in limiting inflammation and preventing the apoptotic response to IR. We previously demonstrated that HO-1 is up-regulated in liver graft after remote ischemic preconditioning (RIPC). The aim of this study was to confirm that RIPC protects against IR via HO-1-mediated autophagy. Methods RIPC was performed with regional ischemia of limbs before liver ischemia, and HO-1 activity was inhibited pre-operation. Autophagy was assessed by the expression of light chain 3-II (LC3-II). The HO-1/extracellular signal-related kinase (ERK)/p38/mitogen-activated protein kinase (MAPK) pathway was detected in an autophagy model and mineral oil-induced IR in vitro. Results In liver IR, the expression of LC3-II peaked 12–24 h after IR, and the ultrastructure revealed abundant autophagosomes in hepatocytes after IR. Autophagy was inhibited when HO-1 was inactivated, which we believe resulted in the aggravation of liver IR injury (IRI) in vivo. Hemin-induced autophagy also protected rat hepatocytes from IRI in vitro, which was abrogated by HO-1 siRNA. Phosphorylation of p38-MAPK and ERK1/2 was up-regulated in hemin-pretreated liver cells and down-regulated after treatment with HO-1 siRNA. Conclusions RIPC may protect the liver from IRI by induction of HO-1/p38-MAPK-dependent autophagy.
    PLoS ONE 06/2014; 9(6):e98834. DOI:10.1371/journal.pone.0098834 · 3.23 Impact Factor
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    • "Current advances in the field have shown that mitophagy activity is highly sensitive to a wide array of cellular and mitochondrial cues. Indeed, perturbation of the cellular bioenergetic status, changes of oxygen tension, loss of mitochondrial membrane potential, an increase in cellular ROS69 (either derived from the cytosol or mitochondria), a disturbance of Ca2+ signaling, defects in mitochondrial protein import or export70, mtDNA damages71, and perturbation of the mitochondrial protein quality control system or accumulation of protein aggregates in mitochondria can all activate mitophagy72. "
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    ABSTRACT: Mitophagy, or mitochondria autophagy, plays a critical role in selective removal of damaged or unwanted mitochondria. Several protein receptors, including Atg32 in yeast, NIX/BNIP3L, BNIP3 and FUNDC1 in mammalian systems, directly act in mitophagy. Atg32 interacts with Atg8 and Atg11 on the surface of mitochondria, promoting core Atg protein assembly for mitophagy. NIX/BNIP3L, BNIP3 and FUNDC1 also have a classic motif to directly bind LC3 (Atg8 homolog in mammals) for activation of mitophagy. Recent studies have shown that receptor-mediated mitophagy is regulated by reversible protein phosphorylation. Casein kinase 2 (CK2) phosphorylates Atg32 and activates mitophagy in yeast. In contrast, in mammalian cells Src kinase and CK2 phosphorylate FUNDC1 to prevent mitophagy. Notably, in response to hypoxia and FCCP treatment, the mitochondrial phosphatase PGAM5 dephosphorylates FUNDC1 to activate mitophagy. Here, we mainly focus on recent advances in our understanding of the molecular mechanisms underlying the activation of receptor-mediated mitophagy and the implications of this catabolic process in health and disease.Cell Research advance online publication 6 Jun 2014; doi:10.1038/cr.2014.75.
    Cell Research 06/2014; 24(7). DOI:10.1038/cr.2014.75 · 12.41 Impact Factor
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