Biology subjects

Hildebrand, A. N.

Publications and source records attributed to Hildebrand, A. N..

3 recordsLinked to original sources

bioRxiv2023

Genetic ablation of neuronal mitochondrial calcium uptake halts Alzheimer's disease progression

Loss of mCa2+ efflux capacity contributes to the pathogenesis and progression of Alzheimers disease (AD) by promoting mitochondrial Ca2+ (mCa2+) overload. Here, we utilized loss-of-function genetic mouse models to causally evaluate the role of mCa2+ uptake by conditionally deleting the mitochondrial calcium uniporter channel (mtCU) in a robust mouse model of AD. Loss of neuronal mCa2+ uptake reduced A{beta} and tau-pathology, synaptic dysfunction, and cognitive decline in 3xTg-AD mice. Knockdown of Mcu in an in vitro model of AD significantly reduced matrix Ca2+ content, redox imbalance, and mitochondrial dysfunction. The preservation of mitochondrial function rescued the AD-dependent decline in autophagic capacity and protected neurons against amyloidosis and cell death. This was corroborated by in vivo data showing improved mitochondrial structure and apposition in AD mice with loss of neuronal Mcu. These results suggest that inhibition of neuronal mCa2+ uptake represents a powerful therapeutic target to impede AD progression.

neuroscience↗

bioRxiv2023

TMEM65 regulates NCLX-dependent mitochondrial calcium efflux

The balance between mitochondrial calcium (mCa2+) uptake and efflux regulates ATP production, but if perturbed causes energy starvation or mCa2+ overload and cell death. The mitochondrial sodium-calcium exchanger, NCLX, is a critical route of mCa2+ efflux in excitable tissues, such as the heart and brain, and animal models support NCLX as a promising therapeutic target to limit pathogenic mCa2+ overload. However, the mechanisms that regulate NCLX activity remain largely unknown. We used proximity biotinylation proteomic screening to identify the NCLX interactome and define novel regulators of NCLX function. Here, we discover the mitochondrial inner membrane protein, TMEM65, as an NCLX-proximal protein that potently enhances sodium (Na+)-dependent mCa2+ efflux. Mechanistically, acute pharmacologic NCLX inhibition or genetic deletion of NCLX ablates the TMEM65-dependent increase in mCa2+ efflux. Further, loss-of-function studies show that TMEM65 is required for Na+-dependent mCa2+ efflux. Co-fractionation and in silico structural modeling of TMEM65 and NCLX suggest these two proteins exist in a common macromolecular complex in which TMEM65 directly stimulates NCLX function. In line with these findings, knockdown of Tmem65 in mice promotes mCa2+ overload in the heart and skeletal muscle and impairs both cardiac and neuromuscular function. We further demonstrate that TMEM65 deletion causes excessive mitochondrial permeability transition, whereas TMEM65 overexpression protects against necrotic cell death during cellular Ca2+ stress. Collectively, our results show that loss of TMEM65 function in excitable tissue disrupts NCLX-dependent mCa2+ efflux, causing pathogenic mCa2+ overload, cell death and organ-level dysfunction, and that gain of TMEM65 function mitigates these effects. These findings demonstrate the essential role of TMEM65 in regulating NCLX-dependent mCa2+ efflux and suggest modulation of TMEM65 as a novel strategy for the therapeutic control of mCa2+ homeostasis.

physiology↗

bioRxiv2023

MCU gain- and loss-of-function models define the duality of mitochondrial calcium uptake in heart failure

BackgroundMitochondrial calcium (mCa2+) uptake through the mitochondrial calcium uniporter channel (mtCU) stimulates metabolism to meet acute increases in cardiac energy demand. However, excessive mCa2+ uptake during stress, as in ischemia-reperfusion, initiates permeability transition and cell death. Despite these often-reported acute physiological and pathological effects, a major unresolved controversy is whether mtCU-dependent mCa2+ uptake and long-term elevation of cardiomyocyte mCa2+ contributes to the hearts adaptation during sustained increases in workload. ObjectiveWe tested the hypothesis that mtCU-dependent mCa2+ uptake contributes to cardiac adaptation and ventricular remodeling during sustained catecholaminergic stress. MethodsMice with tamoxifen-inducible, cardiomyocyte-specific gain (MHC-MCM x flox-stop-MCU; MCU-Tg) or loss (MHC-MCM x Mcufl/fl; Mcu-cKO) of mtCU function received 2-wk catecholamine infusion. ResultsCardiac contractility increased after 2d of isoproterenol in control, but not Mcu-cKO mice. Contractility declined and cardiac hypertrophy increased after 1-2-wk of isoproterenol in MCU-Tg mice. MCU-Tg cardiomyocytes displayed increased sensitivity to Ca2+- and isoproterenol-induced necrosis. However, loss of the mitochondrial permeability transition pore (mPTP) regulator cyclophilin D failed to attenuate contractile dysfunction and hypertrophic remodeling, and increased isoproterenol-induced cardiomyocyte death in MCU-Tg mice. ConclusionsmtCU mCa2+ uptake is required for early contractile responses to adrenergic signaling, even those occurring over several days. Under sustained adrenergic load excessive MCU-dependent mCa2+ uptake drives cardiomyocyte dropout, perhaps independent of classical mitochondrial permeability transition pore opening, and compromises contractile function. These findings suggest divergent consequences for acute versus sustained mCa2+ loading, and support distinct functional roles for the mPTP in settings of acute mCa2+ overload versus persistent mCa2+ stress.

physiology↗