Reduction of Elevated Proton Leak Rejuvenates Mitochondria in the Aged Cardiomyocyte

Abstract

17Aging-associated diseases, including cardiac dysfunction, are increasingly common in the 18population. However, the mechanisms of physiologic aging in general, and cardiac aging in 19 particular, remain poorly understood. While effective medical interventions are available for 20 some kinds of heart failure, one age-related impairment, diastolic dysfunction in Heart Failure 21with Preserved Ejection Fraction (HFpEF) is lacking a clinically effective treatment. Using the 22 model of naturally aging mice and rats, we show direct evidence of increased proton leak in the 23 aged heart mitochondria. Moreover, we identified ANT1 as mediating the increased proton 24 permeability of old cardiomyocytes. Most importantly, the tetra-peptide drug SS-31 25(elamipretide) prevents age-related excess proton entry, decreases the mitochondrial flash 26 activity and mitochondrial permeability transition pore (mPTP) opening and rejuvenates 27 mitochondrial function by direct association with ANT1 and the mitochondrial ATP synthasome. 28Our results uncover a novel mechanism of age-related cardiac dysfunction and elucidate how 29 SS-31 is able to reverse this clinically important complication of cardiac aging. 30 31 Significance 32Aging is the greatest risk factor for cardiac dysfunction, including Heart Failure with Preserved 33Ejection Fraction (HFpEF). Unfortunately, the mechanisms of cardiac aging remain elusive, and 34there are no effective pharmacologic therapies for HFpEF. Here, we show direct evidence of 35 increased proton leak in aged cardiac mitochondria and have identified ANT1 as mediating the 36 increased proton permeability of old cardiomyocytes. Moreover, the mitochondrial-targeted 37 tetra-peptide SS-31 (elamipretide) prevents the age-related excess proton entry and 38 rejuvenates mitochondrial function by direct association with ANT1 and the mitochondrial ATP 39 synthasome, resulting in alleviation of diastolic dysfunction in old mice. Our results unmask a 40 novel mechanism of cardiac aging and elucidate how SS-31 reverses this clinically important 41 complication of aging. 42

Mitochondrial protein interaction landscape of SS-31

“…In addition to direct binding of SS-31 with ADT, the same group also reported that SS-31 associated with ATP synthasome and stabilized it [70]. Our crosslink data provides direct evidences that bSS-31 binds with two members of ATP synthasome supercomplex, CV and ADT.…”

“…Studies have shown that proton leak through ADT was elevated in aged mitochondria and impairment of ADT was believed to play a central mechanism causing aged-related mitochondria defects [66][67][68]. More significantly, a recent report showed that mitochondrial trifunctional enzyme-deficient cardiac model system that simulates SIDS disease had a higher level proton leak and SS-31 treatment rescued the increased proton leak [69]; another recent report showed that SS-31 can suppress proton leak and rejuvenate mitochondrial function through direct binding with ADT [70]. SS-31 shares many common properties with inhibitors BKA and CATR such as larger molecular volume than ADP or ATP, and binding with ADT via multiple polar interactions [64]; and additionally, all three molecules were reported to block proton leak in ADT [70].…”

“…More significantly, a recent report showed that mitochondrial trifunctional enzyme-deficient cardiac model system that simulates SIDS disease had a higher level proton leak and SS-31 treatment rescued the increased proton leak [69]; another recent report showed that SS-31 can suppress proton leak and rejuvenate mitochondrial function through direct binding with ADT [70]. SS-31 shares many common properties with inhibitors BKA and CATR such as larger molecular volume than ADP or ATP, and binding with ADT via multiple polar interactions [64]; and additionally, all three molecules were reported to block proton leak in ADT [70]. However unlike SS-31, BKA and CATR decreased the maximal respiratory rate and failed to enhance respiratory control ratio [70].…”

“…SS-31 shares many common properties with inhibitors BKA and CATR such as larger molecular volume than ADP or ATP, and binding with ADT via multiple polar interactions [64]; and additionally, all three molecules were reported to block proton leak in ADT [70]. However unlike SS-31, BKA and CATR decreased the maximal respiratory rate and failed to enhance respiratory control ratio [70]. SS-31 binding to the ADT channel yet not functioning as an ADT inhibitor opens the possibility of therapeutic alteration of the balance of conformational states, potentially modulating proton leak and ADP/ATP exchange.…”

SS‐31 and NMN: Two paths to improve metabolism and function in aged hearts

“…Other reasons why ATP production may be enhanced by SS-31 lie in its direct effects on the mitochondrial inner membrane and inner membrane proteins. We have recently demonstrated that SS-31 is highly effective at preventing an age-associated excess protein leak across the mitochondrial inner membrane, which appears to be mediated by the adenine nucleotide translocase (Zhang et al, 2020). Reduced production of reactive oxygen species, which otherwise cause damage to mitochondrial proteins and consume NADPH through antioxidant regeneration, also occurs and is another likely mechanism for the improved efficiency (Chiao et al, 2020).…”