The pivotal role of mitochondrial Ca2+, reactive oxygen species (ROS), and morphology in controlling cell fate is well recognized. In cardiac muscle cells, it has been proposed that increases in mitochondrial Ca2+ concentrations ([Ca2+]m) enhance ATP and ROS generation as well as mitochondrial fission. However, the precise contribution of mitochondrial Ca2+ uniporter (mtCU), the primary mechanism for mitochondrial Ca2+ influx, in regulating mitochondrial ATP, ROS, and fission is still inconclusive mostly due to the lack of its molecular identity. Furthermore, without the molecular information, i has been challenging to study the molecular mechanisms of how mtCU is regulated in the physiological and pathological conditions. In 2011, two ground-breaking studies have elucidated the molecular components of the mtCU complexes including the pore forming unit (MCU), the coiled-coil domain-containing protein 109A (CCDC109A), and regulatory components (MICU1-3). Meanwhile, it has gained appreciation that Ca2+-dependent redox-sensitive proline-rich tyrosine kinase 2 (Pyk2) functions as a key transducer of stress stimuli involved in pathological cardiac remodeling and the progression of heart failure (HF). Intriguingly, basal tyrosine phosphorylation of CCDC109A was reported from mass spectroscopy analyses of human and mouse samples. Finally, mitochondrial Ca2+ overload can cause HF through events (e.g. oxidative stress and energy depletion) associated with the opening of mitochondrial permeability transition pores (mPTP). We hypothesize that Pyk2 phosphorylates MCU that increases the number of tetrametric channels by oligomerization so that mitochondrial Ca2+ uptake is enhanced. The increases in [Ca2+]m augments ROS generation. This increase in ROS promotes mitochondrial fission. Physiologically, mitochondrial Ca2+ and fission work in concert to increase ATP production efficiently. However, under stress, excessive Pyk2 and MCU activation leads to pathologically high levels of mitochondrial Ca2+, fission, and ROS, which cause prolonged mPTP opening, resulting in cell injury/death and subsequent HF. To test this hypothesis, we will employ multiple techniques including biochemistry (from in vitro to in situ assays), molecular biology (gene knock in or knock out, overexpression, RNA interference), cell biology (confocal, fluorescence resonance energy transfer, electron microscopy), biophysics (single channel recordings with lipid bilayer or mitoplast), cardiac physiology (echocardiogram), and phenylephrine infusion mouse model of HF, to obtain experimental results that will lead to mechanistic insights. The feature of pinpointing the precise phosphorylation sites of MCU by Pyk2 and demonstrating the formation of functional Ca2+ permeable channels through MCU oligomerization is unique. The elucidation of molecular mechanisms how increases in [Ca2+]m induce fission will significantly add novel insights regarding crosstalk signaling between mitochondrial form and function. Finally, the prospect of tweaking Pyk2/MCU signaling pathways for treating human diseases will be encouraging because the destruction of mitochondrial Ca2+ homeostasis is a key element for leading to mitochondrial dysfunction-associated clinical phenotypes including heart diseases (e.g. HF), neurodegenerative diseases, metabolic diseases (diabetes), and aging.

Public Health Relevance

Failure to provide sufficient cellular energy by mitochondria can cause numerous human diseases including ischemic heart disease, sudden cardiac death, neurodegenerative diseases, diabetes, and aging. One of the key regulators for the mitochondrial energy production is calcium. The proposed research will explore the mechanisms of post translational modification of mitochondrial calcium influx transporting protein. Moreover, we will investigate how the disturbances of this mitochondrial influx mechanism can result in cell injury and death, which is one of the major causes for heart failure. The completion of this research work not only will provide new and fundamental principles about mitochondrial calcium controls the function and dysfunction of the heart, but also will shed the light about how to develop possible therapeutic means for treating the above-mentioned debilitating disorders.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL093671-08
Application #
9261558
Study Section
Cardiac Contractility, Hypertrophy, and Failure Study Section (CCHF)
Program Officer
Wong, Renee P
Project Start
2010-01-01
Project End
2019-04-30
Budget Start
2017-05-01
Budget End
2019-04-30
Support Year
8
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Thomas Jefferson University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
053284659
City
Philadelphia
State
PA
Country
United States
Zip Code
19107
De La Fuente, Sergio; Lambert, Jonathan P; Nichtova, Zuzana et al. (2018) Spatial Separation of Mitochondrial Calcium Uptake and Extrusion for Energy-Efficient Mitochondrial Calcium Signaling in the Heart. Cell Rep 24:3099-3107.e4
Wang, Wang; Fernandez-Sanz, Celia; Sheu, Shey-Shing (2018) Regulation of mitochondrial bioenergetics by the non-canonical roles of mitochondrial dynamics proteins in the heart. Biochim Biophys Acta Mol Basis Dis 1864:1991-2001
Hurst, Stephen; Hoek, Jan; Sheu, Shey-Shing (2017) Mitochondrial Ca2+ and regulation of the permeability transition pore. J Bioenerg Biomembr 49:27-47
Zhang, Huiliang; Wang, Pei; Bisetto, Sara et al. (2017) A novel fission-independent role of dynamin-related protein 1 in cardiac mitochondrial respiration. Cardiovasc Res 113:160-170
Mishra, Jyotsna; Jhun, Bong Sook; Hurst, Stephen et al. (2017) The Mitochondrial Ca2+Uniporter: Structure, Function, and Pharmacology. Handb Exp Pharmacol 240:129-156
Wang, Wang; Gong, Guohua; Wang, Xianhua et al. (2016) Mitochondrial Flash: Integrative Reactive Oxygen Species and pH Signals in Cell and Organelle Biology. Antioxid Redox Signal 25:534-49
Jhun, Bong Sook; Mishra, Jyotsna; Monaco, Sarah et al. (2016) The mitochondrial Ca2+ uniporter: regulation by auxiliary subunits and signal transduction pathways. Am J Physiol Cell Physiol 311:C67-80
O-Uchi, Jin; Sorenson, Jaime; Jhun, Bong Sook et al. (2015) Isoform-specific dynamic translocation of PKC by ?1-adrenoceptor stimulation in live cells. Biochem Biophys Res Commun 465:464-70
Fujiwara, Keigi; Sheu, Shey-Shing (2015) Mitochondrial dynamics regulate neointima formation. Cardiovasc Res 106:175-7
Gomez, L; Thiebaut, P-A; Paillard, M et al. (2015) The SR/ER-mitochondria calcium crosstalk is regulated by GSK3? during reperfusion injury. Cell Death Differ 22:1890

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