4. Melatonin and mitophagy regulation in mammalian cells
Myriad studies have delineated the preventive role of melatonin against atherosclerosis. To begin with, our group for the first time, reported that melatonin administration impedes activation of NLRP3 inflammasome and consequently ceases secretion of inflammatory factors from macrophages in atherosclerotic lesions in a murine model of atherosclerosis (Ma et al., 2018). The underlying mechanism seems to be related to melatonin-mediated activation of mitophagy via a SIRT3-FOXO3 (forkhead box O3)-PRKN signaling cascade, resulting in ROS scavenging and inhibition of ROS-activated NLRP3 inflammasomes (Ma et al., 2018).
Similar results were noted from examination of melatonin in subarachnoid hemorrhage (Cao et al., 2017). This study revealed that melatonin treatment significantly upregulates autophagy-associated molecules such as ATG5 and MAP1LC3A, and mitophagy molecules such as PRKN and PINK1 in subarachnoid hemorrhage, resulting in ROS scavenging, remarkable inactivation of the NLRP3 inflammasomes, and attenuation of cytokine secretion (Cao et al., 2017). Furthermore, examination of a rat model of radiculopathy showed that melatonin alleviates apoptosis and NLRP3 inflammasome activation through instigating PRKN-dependent mitophagy (Xie et al., 2021).
Conversely, melatonin was shown to induce OPA1 (OPA1 mitochondrial dynamin like GTPase)-mediated mitophagy and mitochondrial fusion via AMP-activated protein kinase (AMPK) under ischemia-reperfusion (I/R) injury in vivo and in vitro , prompting the role of AMPK-OPA1 axis as a new paradigm for melatonin-evoked mitophagy induction (Zhang et al., 2019). Likewise, melatonin significantly attenuates calcium deposition in vascular smooth muscle cells in a pattern dependent on mitophagy via AMPK-OPA1 axis (Chen, Zhou, Yang, Liu, Wu & Sha, 2020). Besides, melatonin downregulates cleaved CASP3 (caspase 3) and RUNX2 (RUNX family transcription factor 2), upregulates MAP1LC3B and MFN2 (mitofusin 2), reduces mitochondrial superoxide, and activates mitophagy via the AMPK-OPA1 signaling axis (Chen, Zhou, Yang, Liu, Wu & Sha, 2020).
In addition, melatonin treatment reactivates mitophagy and boosts mitochondrial function via upregulation of HSPA1L (heat shock protein family A [Hsp70] member 1 like) in senescent mesenchymal stem cells. Mechanistically, HSPA1L forms a complex with cellular PRNP (prion protein) then recruits PRNP to the mitochondria. Afterward, the HSPA1L-PRNP complex binds to COX4I1 (cytochrome c oxidase subunit 4I1), resulting in elevated mitochondrial membrane potential, induced antioxidant enzymes, and mitophagy, validating the role of the HSPA1L-PRNP-COX4I1 axis in melatonin-induced mitophagy induction (Lee, Yoon, Song, Noh & Lee, 2020). Likewise, supplementation of human mesenchymal stem cells with melatonin, causes HSPA1L upregulation, and, enhances HSPA1L-mediated recruitment of PRKN to mitochondria, to favor mitophagy and cell survival (Yoon, Kim, Lee & Lee, 2019). Therefore, the HSPA1L-PRKN axis is modulated by melatonin and promotes mitophagy.
A number of studies have noted PINK1 and PRKN modulation by melatonin. Melatonin supplementation culminates in upregulation of PINK1, PRKN, PPARGC1A (PPARG coactivator 1 alpha), NRF1 (nuclear respiratory factor 1), and TFAM (transcription factor A, mitochondrial) proteins with a role in mitophagy and mitochondrial integrity in rats with liver fibrosis (Kang, Hong & Lee, 2016). Melatonin-mediated upregulation of PRKN is also observed in nucleus pulposus cells in a time- and dose-dependent manner (Chen et al., 2019). In addition, melatonin enhances PRKN mitochondrial translocation via inhibition of MST1 (macrophage stimulating 1) phosphorylation, compromises mitophagy in diabetic cardiomyopathy (Wang et al., 2018).
Furthermore, ample evidence has shown that melatonin modulates NFE2L2 (nuclear factor, erythroid 2 like 2), HAS3 (hyaluronan synthase 3), and MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1) to induce mitophagy. Melatonin upregulates NFE2L2 to induce NFE2L2-dependent mitophagy to heal brain injury in a murine model of subarachnoid hemorrhage (Sun, Yang, Li & Hang, 2018). Also, melatonin activates HAS3 and associated mitophagy, in a neuroblastoma N2a cell line (Lee et al., 2019). Furthermore, melatonin activates mitophagy through MTORC1 modulation, which in turn, ceases inflammation by suppressing IL1B (interleukin 1 beta) secretion upon immunopathology of traumatic brain injury (Lin et al., 2016).
Melatonin also modulates other signaling pathways to activate mitophagy. Interestingly, our group revealed that melatonin backs up the CGAS (cyclic GMP-AMP synthase)-STING1 (stimulator of interferon response cGAMP interactor 1)-TBK1 (TANK binding kinase 1) signaling pathway, leading to mitophagy reactivation involving ALDH2 (aldehyde dehydrogenase 2 family member) activation in APP- (amyloid beta precursor protein) and PSEN1 (presenilin 1)-mutant mice (Wang et al., 2020). Our study suggested the role of melatonin in rescuing myopathic changes in the heart via reinstating mitophagy. Moreover, melatonin therapy inhibits mitochondrial fission and boosts mitophagy through inhibiting NR4A1 (nuclear receptor subfamily 4 group A member 1)-PRKDC (protein kinase, DNA-activated, catalytic subunit)-TP53 (tumor protein p53) signaling pathway in nonalcoholic fatty liver disease (Zhou et al., 2018).
Despite mitophagy activation, ample studies have revealed other mechanisms of melatonin regarding mitophagy regulation in mammalian cells. It was suggested that melatonin maintains mild induction of mitophagy through preventing excessive mitophagy induction. This notion was observed in a murine model of microvascular I/R injury, where melatonin activates PRKAA1 (protein kinase AMP-activated catalytic subunit alpha 1) to inhibit DNM1L-based mitochondrial fission. Subsequently, the VDAC1-HK2 (hexokinase 2) interaction is recovered, which results in inhibition of the mitochondrial permeability transition pore opening, and attenuation of PINK1-PRKN-dependent mitophagy (Zhou et al., 2017). It is perceived that melatonin-evoked PRKAA1-DNM1L-VDAC1-HK2-mitochondrial permeability transition pore signaling pathway is a cytoprotective mechanism that blunts excessive mitophagy-evoked cell death ensuing microvascular I/R damage.
More cell signaling pathways are reported to be modulated by melatonin to govern mitophagy including the MT2A (metallothionein 2A)-SIRT3-FOXO3 pathway, which inhibits mitophagy in H9c2 cells (Wu, Yang, Gao, Wang & Ma, 2020), and MAPK8 (mitogen-activated protein kinase 8)-PRKN pathway, which is negatively modulated by melatonin, and causes a cessation of excessive mitophagy induction in human HeLa cells (Chen, Liu, Li & Gao, 2018). In addition, melatonin maintains mild levels of mitophagy-associated proteins such as PRKN, BECN1, SIRT3, FOXO3, and BNIP3L, thus keeping mitophagy in check (Wu, Yang, Gao, Wang & Ma, 2020). Melatonin can also upregulate SIRT1 (sirtuin 1), as a result suppressing excessive PINK1-PRKN mitophagy (Yi, Zheng, Zhu, Cai, Sun & Zhou, 2020). Collectively, these data suggest that melatonin regulates mitophagy by two dogmas: (i) induction of a low-level activation and (ii) prevention of excessive activation, both of which, can be beneficial to cell homeostasis and inflammation suppression.