Aging is a complex biological process and environmental risk factors like pesticide exposure have been implicated in the increased incidence of age-related neurodegenerative diseases like Parkinson’s disease (PD) but the etiology remains unknown. There is also lack of a proper animal model system to study the progressive effect of these environmental toxins on age-associated neurodegeneration. In this study, we established a drosophila model of aging to study the age-dependent vulnerability to the environmental toxin rotenone that has been implicated in sporadic cases of PD. We demonstrate that age plays a determining role in the increased susceptibility to chronic rotenone exposure that is accompanied by severe locomotor deficits, decreased lifespan and loss of dopaminergic (DA) neurons. Chronic low dose exposure to rotenone results in the rapid induction of the neurodegenerative molecule SARM1/dSarm. Further, the age-dependent dSarm induction is accompanied by a heightened inflammatory response (increased expression of Eiger and Relish) that is independent of reactive oxygen species (ROS) generation in the observed rotenone-induced neurotoxicity. dSarm induction and subsequent locomotor deficits is reversed in the presence of the anti-inflammatory molecule resveratrol. Thus, dSarm and heightened inflammatory responses may play a crucial role in age-dependent vulnerability to the pesticide rotenone thus making it an attractive target to help develop cost-effective therapeutic strategies to prevent ongoing dopaminergic neuronal loss as seen in PD.
Sarm1 is an evolutionary conserved innate immune adaptor protein that has emerged as a primary regulator of programmed axonal degeneration over the past decade. In vitro structural insights have revealed that although Sarm1 induces energy depletion by breaking down nicotinamide adenine dinucleotide+ (NAD+), it is also allosterically inhibited by NAD+. However, how NAD+ levels modulate the activation of intracellular Sarm1 has not been elucidated so far. This study focuses on understanding the events leading to Sarm1 activation in both neuronal and non‐neuronal cells using the mitochondrial complex I inhibitor rotenone. Here, we report the regulation of rotenone‐induced cell death by loss of NAD+ that may act as a ‘biological trigger’ of Sarm1 activation. Our study revealed that early loss of endogenous NAD+ levels arising due to PARP1 hyperactivation preceded Sarm1 induction following rotenone treatment. Interestingly, replenishing NAD+ levels by the PARP inhibitor, PJ34 restored mitochondrial complex I activity and also prevented subsequent Sarm1 activation in rotenone‐treated cells. These cellular data were further validated in Drosophila melanogaster where a significant reduction in rotenone‐mediated loss of locomotor abilities, and reduced dSarm expression was observed in the flies following PARP inhibition. Taken together, these observations not only uncover a novel regulation of Sarm1 induction by endogenous NAD+ levels but also point towards an important understanding on how PARP inhibitors could be repurposed in the treatment of mitochondrial complex I deficiency disorders.
The NADase Sarm1 has emerged as an important modulator of programmed axonal degeneration over the past decade but its mode of activation within the cell is not clearly understood. Sarm1 is predominantly expressed in the neurons, kidney and liver but the non-neuronal regulation of Sarm1 remains relatively unexplored. Here we demonstrate that treatment of the human embryonic kidney cell line HEK293 cells with the mitochondrial complex I inhibitor rotenone, induced early loss of NAD+ that preceded induction of Sarm1, a primary mediator of rotenone induced cell death. Interestingly, replenishing NAD+ levels by PARP inhibition, a major NAD+ consumer within the cell, not only restored mitochondrial homeostasis but also prevented subsequent Sarm1 induction by rotenone. These early changes were further marked by a distinct subcellular localization pattern of Sarm1 in the nucleus and the mitochondria that was accompanied by significantly reduced cell death. Taken together, our study provides the first preliminary evidence of temporal regulation of endogenous Sarm1 by fluctuating NAD+ levels induced by rotenone that may act as a biological trigger of Sarm1 activation. This also points towards an important understanding on how PARP inhibitors like PJ34 could be repurposed in the treatment of Sarm1 mediated mitochondrial deficiency disorders.
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