Mitofusin 2-Containing Mitochondrial-Reticular Microdomains Direct Rapid Cardiomyocyte Bioenergetic Responses Via Interorganelle Ca ²⁺ Crosstalk

Abstract

Rationale Mitochondrial Ca2+ uptake is essential for the bioenergetic feedback response through stimulation of Krebs cycle dehydrogenases. Close association of mitochondria to the sarcoplasmic reticulum (SR) may explain efficient mitochondrial Ca2+ uptake despite low Ca2+ affinity of the mitochondrial Ca2+ uniporter. However, the existence of such mitochondrial Ca2+ microdomains and their functional role are presently unresolved. Mitofusin (Mfn) 1 and 2 mediate mitochondrial outer membrane fusion, while Mfn2, but not Mfn1, tethers endoplasmic reticulum to mitochondria in non-cardiac cells. Objective To elucidate roles for Mfn1 and 2 in SR-mitochondrial tethering, Ca2+ signaling and bioenergetic regulation in cardiac myocytes. Methods and Results Fruit fly heart tubes deficient of the Drosophila Mfn ortholog, MARF, had increased contraction-associated and caffeine-sensitive Ca2+ release, suggesting a role for Mfn in SR Ca2+ handling. While cardiac-specific Mfn1 ablation had no effects on murine heart function or Ca2+ cycling, Mfn2 deficiency decreased cardiomyocyte SR-mitochondrial contact length by 30% and reduced the content of SR-associated proteins in mitochondria-associated membranes. This was associated with decreased mitochondrial Ca2+ uptake (despite unchanged mitochondrial membrane potential) but increased steady-state and caffeine-induced SR Ca2+ release. Accordingly, Ca2+-induced stimulation of Krebs cycle dehydrogenases during β-adrenergic stimulation was hampered in Mfn2-, but not Mfn1-KO myocytes, evidenced by oxidation of the redox states of NAD(P)H/NAD(P)+ and FADH2/FAD. Conclusions Physical tethering of SR and mitochondria via Mfn2 is essential for normal inter-organelle Ca2+ signaling in the myocardium, consistent with a requirement for SR-mitochondrial Ca2+ signaling through microdomains in the cardiomyocyte bioenergetic feedback response to physiological stress.

Signalling mechanisms underlying doxorubicin and Nox2 NADPH oxidase‐induced cardiomyopathy: involvement of mitofusin‐2

“…Loss of Mfn2 also delayed membrane depolarization in isolated cardiomyocytes from adult Mfn2 −/− mice, leading to the suggestion that Mfn2 may function to control mitochondrial permeability transition pore opening (Papanicolaou et al, 2011). Similarly, cardiac‐specific deletion of Mfn2 produced dissipation of mitochondrial membrane potential and elevated ROS production (Chen et al, 2012), whilst overexpression of Mfn2 was found to increase the percentage of cells containing elongated mitochondria, thereby reducing mitochondrial permeability transition pore opening and cell death after simulated ischaemia/reperfusion injury (Ong et al, 2010). It also appears that Mfn2 serves an essential role in maintaining mitochondrial coenzyme Q levels in mouse hearts, thereby promoting optimal function of the respiratory chain (Mourier et al, 2015).…”

Strategic Positioning and Biased Activity of the Mitochondrial Calcium Uniporter in Cardiac Muscle

“…One would also expect direct or indirect contribution by mitochondria-jSR tethers that secure the organelles together. Cardiac ablation of mitofusin 2, a multifunctional OMM protein that has been proposed to be (41) or not to be (42, 43) a mitochondria-ER tether in non-muscle cells, causes a decrease in mitochondria-jSR associations and Ca 2ϩ transfer, suggesting a tethering role in cardiomyocytes (44). However, the -fold enrichment of EMRE in the SR versus mitochondrial fractions was not affected significantly by cardiac mitofusin 2 ablation (n ϭ 4 mice), suggesting that either EMRE recruitment to the mitochondria-jSR association did not require mitofusin 2 or alternative mechanism(s) compensated for its loss.…”

“…MCUKO mice were housed/maintained in NIH-NHLBI 1 and used in the Murphy (cardiomyocyte isolation) and Finkel (heart homogenates and membrane fractions) laboratories. The cardiac muscle-specific Mfn2KO (Mfn2 flox/flox, MYH6 Cre) mouse colony originated from Gerald Dorn (Washington University, St. Louis, MO) and was maintained as described (44).…”

Mitochondrial Dynamism and Cardiac Fate

“…57 Together with our collaborators, we proved the existence of this same Mfn2 tethering function between cardiomyocyte mitochondria and the SR, and demonstrated that the calcium microdomains formed by Mfn2 tethers are essential for rapid mitochondrial sensing of increased SR calcium release during the early phase of increased cardiac work (Figure 3). 20 Ablating the Mfn2 mitochondria-SR tether does not adversely affect the young adult mouse heart at rest, but the resulting distortion of the normal, privileged interorganelle calcium export and uptake delays the mitochondrial ATP response to acute stress. Others have suggested that Mfn2 is essential for calcium-stimulated opening of cardiomyocyte MPTPs, presumably also by tethering mitochondria to the SR. 58 Thus, Mfn2 appears to be a centrally positioned orchestrator of both homeostatic mitochondrial fusion and mitochondrion-SR calcium crosstalk, modulating mitochondrial ATP production according to changing physiological demand and facilitating cardiomyocyte death in response to non-lethal stress by inducing programmed necrosis.…”