Enhanced inter-compartmental Ca2+ flux modulates mitochondrial metabolism and apoptotic threshold during aging

Abstract

BackgroundSenescence is characterized by a gradual decline in cellular functions, including changes in energy homeostasis and decreased proliferation activity. As cellular power plants, contributors to signal transduction, sources of reactive oxygen species (ROS) and executors of programmed cell death, mitochondria are in a unique position to affect aging-associated processes of cellular decline. Notably, metabolic activation of mitochondria is tightly linked to Ca2+ due to the Ca2+ -dependency of several enzymes in the Krebs cycle, however, overload of mitochondria with Ca2+ triggers cell death pathways. Consequently, a machinery of proteins tightly controls mitochondrial Ca2+ homeostasis as well as the exchange of Ca2+ between the different cellular compartments, including Ca2+ flux between mitochondria and the endoplasmic reticulum (ER).MethodsIn this study, we investigated age-related changes in mitochondrial Ca2+ homeostasis, mitochondrial-ER linkage and the activity of the main ROS production site, the mitochondrial respiration chain, in an in vitro aging model based on porcine aortic endothelial cells (PAECs), using high-resolution live cell imaging, proteomics and various molecular biological methods.ResultsWe describe that in aged endothelial cells, increased ER-mitochondrial Ca2+ crosstalk occurs due to enhanced ER-mitochondrial tethering. The close functional inter-organelle linkage increases mitochondrial Ca2+ uptake and thereby the activity of the mitochondrial respiration, but also makes senescent cells more vulnerable to mitochondrial Ca2+-overload-induced cell death. Moreover, we identified the senolytic properties of the polyphenol resveratrol, triggering cell death via mitochondrial Ca2+ overload exclusively in senescent cells.ConclusionBy unveiling aging-related changes in the inter-organelle tethering and Ca2+ communications we have advanced the understanding of endothelial aging and highlighted a potential basis to develop drugs specifically targeting senescent cells.

ALG-2 and Peflin Stimulate or Inffibit Copii Targeting and Secretion in Response to Calcium Signaling

“…Both lines were treated the same and were grown at 37 o C using high glucose DMEM supplemented with 10% fetal calf serum, and 1% penicillin-streptomycin. Porcine aorta endothelial cells were isolated and cultured to P5 as described before (Madreiter-Sokolowski et al, 2019). For histamine treatments transfected and/or siRNA-transfected NRK cells were grown to 25-75% confluency in 6-well chambers.…”

Mitochondrial Dysfunction in Age‐Related Metabolic Disorders

“…[ 28 ] Aging endothelial cells exhibit enhanced endoplasmic reticulum (ER)–mitochondrial tethering, increased mitochondrial Ca 2+ uptake leading to matrix Ca 2+ overload. [ 29 ] Mitochondrial Ca 2+ overload has also been reported to induce increased hydrogen peroxide (H 2 O 2 ) release in aged muscle tissues. [ 30 ] While overexpression of either pore forming subunit MCU or positive regulator MCUR1 leads to enhanced mitochondrial calcium uptake, [ 16 ] loss of gatekeepers MICU1, or MICU2 results in mitochondrial calcium overload.…”

Tracking intra‐ and inter‐organelle signaling of mitochondria

“…Various proteins stabilizing and modulating mitochondrial–ER interplay, including Ras‐related protein RAB32 , MFN2 , or phosphofurin acidic cluster sorting protein 2 , as well as protein tethering complexes like inositol triphosphate receptor (IP3R), inositol‐requiring enzyme 1 α , glucose‐related protein 75 and VDAC, have been identified and characterized . Disrupted communication between the ER and mitochondria has been associated with pathological conditions and human diseases , such as Alzheimer's disease , Charcot‐Marie Tooth , Parkinson's disease , viral infections , cancer , diabetes mellitus, and age‐related dysfunction .…”

“…Therefore, large efforts have recently been placed to develop methods that allow the visualization of MAMs in living cells and still provide high spatial resolution. As discussed in our ‘Techniques to analyze structure and shape’ section, advancements in high‐resolution fluorescence microscopy and in the design of organelle‐targeted FP‐tagged proteins or sensors allow to visualize and investigate structural and functional mitochondrial and ER interplay in living cells . Moreover, split FP (split‐FP) approaches or so‐called bimolecular fluorescence complementation technology have been used to study mitochondrial–ER interaction .…”