scholarly journals Reactive oxygen species rescue lysosome coalescence during PIKfyve inhibition

2019 ◽  
Author(s):  
Golam T. Saffi ◽  
Evan Tang ◽  
Aaron Fountain ◽  
Roberto J. Botelho
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Phosphorylation ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Phosphoinositide Kinase ◽  
Photo Damage ◽  
Endosomal Pathway ◽  
Mode Of Production

AbstractLysosomes are terminal, degradative organelles of the endosomal pathway that undergo repeated fusion-fission cycles with themselves and other organelles like endosomes, phagosomes, and autophagosomes. Lysosome number, size and degradative flux depends on the equilibrium between fusion and fission rates. Thus, conditions that favour fusion over fission will reduce lysosome numbers while enlarging remaining lysosomes. Conversely, conditions that favour fission over fusion will cause lysosome fragmentation and increase their numbers. PIKfyve is a phosphoinositide kinase that generates phosphatidylinositol-3,5-bisphosphate to modulate several lysosomal functions. PIKfyve inhibition causes a dramatic increase in lysosome size and reduction in lysosome number, consistent with lysosome coalescence. This is thought to proceed through reduced lysosome reformation and/or fission after fusion with endosomes or other lysosomes. Previously, we observed that photo-damage during live-cell imaging prevented lysosome coalescence during acute PIKfyve inhibition. Thus, we postulated that lysosome fusion and/or fission dynamics are affected by reactive oxygen species (ROS). Here, we show that ROS generated by four independent mechanisms all arrested lysosome coalescence during PIKfyve inhibition and accelerated lysosome fragmentation during PIKfyve re-activation. However, depending on the ROS species and/or mode of production, lysosome dynamics were affected distinctly. H2O2 impaired lysosome motility and reduced lysosome fusion with phagosomes, suggesting that H2O2 prevents lysosome coalescence in PIKfyve-impaired cells by reducing lysosome fusogenecity. In comparison, inhibitors of oxidative phosphorylation, glutathione, and thioredoxin that produce superoxide, did not impair lysosome motility but instead promoted clearance of actin puncta on lysosomes formed during PIKfyve inhibition. Additionally, actin depolymerizing agents prevented lysosome coalescence during PIKfyve inhibition. Thus, we discovered that ROS can generally prevent lysosome coalescence during PIKfyve inhibition using distinct mechanisms.

scholarly journals Reactive oxygen species prevent lysosome coalescence during PIKfyve inhibition

PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0259313
Author(s):  
Golam T. Saffi ◽  
Evan Tang ◽  
Sami Mamand ◽  
Subothan Inpanathan ◽  
Aaron Fountain ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Thiol Groups ◽  
Phosphoinositide Kinase ◽  
Photo Damage ◽  
Endosomal Pathway ◽  
Mode Of Production

Lysosomes are terminal, degradative organelles of the endosomal pathway that undergo repeated fusion-fission cycles with themselves, endosomes, phagosomes, and autophagosomes. Lysosome number and size depends on balanced fusion and fission rates. Thus, conditions that favour fusion over fission can reduce lysosome numbers while enlarging their size. Conversely, favouring fission over fusion may cause lysosome fragmentation and increase their numbers. PIKfyve is a phosphoinositide kinase that generates phosphatidylinositol-3,5-bisphosphate to modulate lysosomal functions. PIKfyve inhibition causes an increase in lysosome size and reduction in lysosome number, consistent with lysosome coalescence. This is thought to proceed through reduced lysosome reformation and/or fission after fusion with endosomes or other lysosomes. Previously, we observed that photo-damage during live-cell imaging prevented lysosome coalescence during PIKfyve inhibition. Thus, we postulated that lysosome fusion and/or fission dynamics are affected by reactive oxygen species (ROS). Here, we show that ROS generated by various independent mechanisms all impaired lysosome coalescence during PIKfyve inhibition and promoted lysosome fragmentation during PIKfyve re-activation. However, depending on the ROS species or mode of production, lysosome dynamics were affected distinctly. H2O2 impaired lysosome motility and reduced lysosome fusion with phagosomes, suggesting that H2O2 reduces lysosome fusogenecity. In comparison, inhibitors of oxidative phosphorylation, thiol groups, glutathione, or thioredoxin, did not impair lysosome motility but instead promoted clearance of actin puncta on lysosomes formed during PIKfyve inhibition. Additionally, actin depolymerizing agents prevented lysosome coalescence during PIKfyve inhibition. Thus, we discovered that ROS can generally prevent lysosome coalescence during PIKfyve inhibition using distinct mechanisms depending on the type of ROS.

scholarly journals Trolox-Sensitive Reactive Oxygen Species Regulate Mitochondrial Morphology, Oxidative Phosphorylation and Cytosolic Calcium Handling in Healthy Cells

2012 ◽  
Vol 17 (12) ◽  
pp. 1657-1669 ◽  
Author(s):  
Felix Distelmaier ◽  
Federica Valsecchi ◽  
Marleen Forkink ◽  
Sjenet van Emst-de Vries ◽  
Herman G. Swarts ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Phosphorylation ◽  
Reactive Oxygen ◽  
Cytosolic Calcium ◽  
Calcium Handling ◽  
Mitochondrial Morphology ◽  
Oxygen Species

Mitochondrial-derived reactive oxygen species influence ADP sensitivity, but not CPT-I substrate sensitivity

2018 ◽  
Vol 475 (18) ◽  
pp. 2997-3008 ◽  
Author(s):  
Pierre-Andre Barbeau ◽  
Paula M. Miotto ◽  
Graham P. Holloway
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Phosphorylation ◽  
Acute Exercise ◽  
Adenine Nucleotide ◽  
Mitochondrial Protein ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Emission Rates ◽  
Mitochondrial Ros ◽  
Cpt I

The mechanisms regulating oxidative phosphorylation during exercise remain poorly defined; however, key mitochondrial proteins, including carnitine palmitoyltransferase-I (CPT-I) and adenine nucleotide translocase, have redox-sensitive sites. Interestingly, muscle contraction has recently been shown to increase mitochondrial membrane potential and reactive oxygen species (ROS) production; therefore, we aimed to determine if mitochondrial-derived ROS influences bioenergetic responses to exercise. Specifically, we examined the influence of acute exercise on mitochondrial bioenergetics in WT (wild type) and transgenic mice (MCAT, mitochondrial-targeted catalase transgenic) possessing attenuated mitochondrial ROS. We found that ablating mitochondrial ROS did not alter palmitoyl-CoA (P-CoA) respiratory kinetics or influence the exercise-mediated reductions in malonyl CoA sensitivity, suggesting that mitochondrial ROS does not regulate CPT-I. In contrast, while mitochondrial protein content, maximal coupled respiration, and ADP (adenosine diphosphate) sensitivity in resting muscle were unchanged in the absence of mitochondrial ROS, exercise increased the apparent ADP Km (decreased ADP sensitivity) ∼30% only in WT mice. Moreover, while the presence of P-CoA decreased ADP sensitivity, it did not influence the basic response to exercise, as the apparent ADP Km was increased only in the presence of mitochondrial ROS. This basic pattern was also mirrored in the ability of ADP to suppress mitochondrial H2O2 emission rates, as exercise decreased the suppression of H2O2 only in WT mice. Altogether, these data demonstrate that while exercise-induced mitochondrial-derived ROS does not influence CPT-I substrate sensitivity, it inhibits ADP sensitivity independent of P-CoA. These data implicate mitochondrial redox signaling as a regulator of oxidative phosphorylation.

Effects of short- and medium-term calorie restriction on muscle mitochondrial proton leak and reactive oxygen species production

2004 ◽  
Vol 286 (5) ◽  
pp. E852-E861 ◽  
Author(s):  
Lisa Bevilacqua ◽  
Jon J. Ramsey ◽  
Kevork Hagopian ◽  
Richard Weindruch ◽  
Mary-Ellen Harper
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Phosphorylation ◽  
Metabolic Control ◽  
Calorie Restriction ◽  
Reactive Oxygen ◽  
Proton Leak ◽  
Ros Production ◽  
Oxygen Species ◽  
Medium Term ◽  
Time Points

Reductions in cellular oxygen consumption (V̇o2) and reactive oxygen species (ROS) production have been proposed as mechanisms underlying the anti-aging effects of calorie restriction (CR). Mitochondria are a cell's greatest “sink” for oxygen and also its primary source of ROS. The mitochondrial proton leak pathway is responsible for 20–30% of V̇o2 in resting cells. We hypothesized that CR leads to decreased proton leak with consequential decreases in V̇o2, ROS production, and cellular damage. Here, we report the effects of short-term (2-wk, 2-mo) and medium-term (6-mo) CR (40%) on rat muscle mitochondrial proton leak, ROS production, and whole animal V̇o2. Whole body V̇o2 decreased with CR at all time points, whereas mass-adjusted V̇o2 was normal until the 6-mo time point, when it was 40% lower in CR compared with control rats. At all time points, maximal leak-dependent V̇o2 was lower in CR rats compared with controls. Proton leak kinetics indicated that mechanisms of adaptation to CR were different between short- and medium-term treatments, with the former leading to decreases in protonmotive force (Δp) and state 4 V̇o2 and the latter to increases in Δp and decreases in state 4 V̇o2. Results from metabolic control analyses of oxidative phosphorylation are consistent with the idea that short- and medium-term responses are distinct. Mitochondrial H2O2 production was lower in all three CR groups compared with controls. Overall, this study details the rapid effects of short- and medium-term CR on proton leak, ROS production, and metabolic control of oxidative phosphorylation. Results indicate that a reduction in mitochondrial V̇o2 and ROS production may be a mechanism for the actions of CR.

scholarly journals Targeting Mitochondrial Metabolism as a Strategy to Treat Senescence

Cells ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 3003
Author(s):  
Yun Haeng Lee ◽  
Ji Yun Park ◽  
Haneur Lee ◽  
Eun Seon Song ◽  
Myeong Uk Kuk ◽  
...  
Keyword(s):  
Energy Source ◽  
Reactive Oxygen Species ◽  
Oxidative Phosphorylation ◽  
Reactive Oxygen ◽  
Mitochondrial Metabolism ◽  
Metabolic Reprogramming ◽  
Oxygen Species ◽  
Metabolic Alterations ◽  
Age Related

Mitochondria are one of organelles that undergo significant changes associated with senescence. An increase in mitochondrial size is observed in senescent cells, and this increase is ascribed to the accumulation of dysfunctional mitochondria that generate excessive reactive oxygen species (ROS). Such dysfunctional mitochondria are prime targets for ROS-induced damage, which leads to the deterioration of oxidative phosphorylation and increased dependence on glycolysis as an energy source. Based on findings indicating that senescent cells exhibit mitochondrial metabolic alterations, a strategy to induce mitochondrial metabolic reprogramming has been proposed to treat aging and age-related diseases. In this review, we discuss senescence-related mitochondrial changes and consequent mitochondrial metabolic alterations. We assess the significance of mitochondrial metabolic reprogramming for senescence regulation and propose the appropriate control of mitochondrial metabolism to ameliorate senescence. Learning how to regulate mitochondrial metabolism will provide knowledge for the control of aging and age-related pathologies. Further research focusing on mitochondrial metabolic reprogramming will be an important guide for the development of anti-aging therapies, and will provide novel strategies for anti-aging interventions.

scholarly journals 3D imaging of ZnO NP distribution and ROS accumulation in MCF-7 cells and quantification of retention dynamics using laser scanning confocal microscopy

2020 ◽  
Author(s):  
Aishee Dey ◽  
Gare Suman ◽  
Sarpras Swain ◽  
Proma Bhattacharya ◽  
Vaibhav Dhyani ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Live Cell Imaging ◽  
Laser Scanning ◽  
Cell Imaging ◽  
Time Lapse ◽  
Confocal Microscope ◽  
Oxygen Species ◽  
Laser Scanning Confocal Microscope ◽  
Time Lapse Microscopy ◽  
Mcf 7

Abstract Generally, investigations on nanomedicine involve conventional imaging techniques for obtaining static images on nanoparticle internalization at a single time point where various phases can be overlooked. In contrast, 3D live-cell imaging can be used for obtaining cellular retention of drugs at various phases, and cells can be followed for days. This article demonstrates the application of time-lapse microscopy in the investigation of Poly-L-lysine coated ZnO nanoparticle dynamics. In this work, a laser scanning confocal microscope has been employed to quantify the dynamics of internalization particles and reactive oxygen species generation (ROS) using volumetric imaging. Firstly, we show that simultaneous spatial mapping of nanoparticle uptake in MCF-7 cells and ROS in a single cell can be used to identify the interdependence between the accumulation of particles and ROS generation. Secondly, monitoring of ROS formation and cytotoxicity using the same imaging platform offers an advantage over monitoring these parameters using various instruments. Finally, the ability of the fluorescent particles in inducing a significant reduction in cell viability suggests its potential to be used as a therapeutic agent. The proposed framework opens up a new avenue of research for investigating mechanistic aspects of ZnO particle adsorption in vitro through long term imaging. Keywords: Fluorescent ZnO particle, Time-lapse microscopy, 3D Live-cell imaging, laser scanning confocal microscope, Reactive oxygen species

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