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Thesis
Peer Reviewed

The Molecular Mechanisms of Lifespan Extension by Alpha-Ketoglutarate in Caenorhabditis elegans

Chin, Randall
Advisor(s): Huang, Jing

UCLA Electronic Theses and Dissertations (2014)

As the elderly population is now the fastest growing age group in the United States, there is an imminent need to understand the molecular mechanisms of aging and develop new approaches to counter aging and the maladies that come with it. Modulation of metabolism, either by dietary restriction or pharmacologic or genetic alteration of metabolic pathways, can extend lifespan and delay age-related diseases in evolutionarily diverse organisms. Here we demonstrate, for the first time, a role for alpha-ketoglutarate (alpha-KG) in the regulation of aging. Alpha-KG serves many functions in the cell, including being an intermediate of the tricarboxylic acid cycle, a precursor for several amino acids, and a substrate for alpha-KG-dependent oxygenases. We show that alpha-KG extends the mean lifespan of adult Caenorhabditis elegans by about 50%, and provide evidence to suggest that this is not due to the metabolism of alpha-KG into other metabolites. Other metabolites have been shown to extend lifespan as well, but without a well defined mechanism. Research on small molecules in aging has been hindered by the inability to identify drug targets. We use a label-free, unbiased drug target identification strategy termed DARTS (drug affinity responsive target stability) to uncover a new binding target of alpha-KG, the beta subunit of the ATP synthase. We reveal that alpha-KG inhibits ATP synthase in in vitro and in vivo assays using both worm and mammalian models. Consistently, genetic or pharmacological inhibition of ATP synthase yields the same phenotypes as alpha-KG: increased lifespan, decreased ATP levels, lower oxygen consumption rates, elevated levels of reactive oxygen species, and reduced target of rapamycin signaling. However, unlike genetic inhibition of ATP synthase, alpha-KG does not induce the mitochondrial unfolded protein response, cause larval arrest, or slow pharyngeal pumping rates. Remarkably, alpha-KG does not further increase the lifespan of dietary restricted animals and is elevated during starvation, a testament to it being a key regulator of dietary restriction mediated lifespan extension. Like dietary restriction, alpha-KG also induces autophagy, and we provide evidence to suggest that autophagy may play an essential role for the longevity of alpha-KG treated animals. Our findings posit alpha-KG as a key regulator of metabolic pathways in response to nutrient status, and suggest new strategies for the prevention and treatment of aging and age-related diseases.

Cover page: The Molecular Mechanisms of Lifespan Extension by Alpha-Ketoglutarate in Caenorhabditis elegans

Article
Peer Reviewed

2-Hydroxyglutarate Inhibits ATP Synthase and mTOR Signaling.

UCLA Previously Published Works (2015)

We discovered recently that the central metabolite α-ketoglutarate (α-KG) extends the lifespan of C. elegans through inhibition of ATP synthase and TOR signaling. Here we find, unexpectedly, that (R)-2-hydroxyglutarate ((R)-2HG), an oncometabolite that interferes with various α-KG-mediated processes, similarly extends worm lifespan. (R)-2HG accumulates in human cancers carrying neomorphic mutations in the isocitrate dehydrogenase (IDH) 1 and 2 genes. We show that, like α-KG, both (R)-2HG and (S)-2HG bind and inhibit ATP synthase and inhibit mTOR signaling. These effects are mirrored in IDH1 mutant cells, suggesting a growth-suppressive function of (R)-2HG. Consistently, inhibition of ATP synthase by 2-HG or α-KG in glioblastoma cells is sufficient for growth arrest and tumor cell killing under conditions of glucose limitation, e.g., when ketone bodies (instead of glucose) are supplied for energy. These findings inform therapeutic strategies and open avenues for investigating the roles of 2-HG and metabolites in biology and disease.

Cover page: 2-Hydroxyglutarate Inhibits ATP Synthase and mTOR Signaling.

Article
Peer Reviewed

The metabolite α-ketoglutarate extends lifespan by inhibiting ATP synthase and TOR.

UCLA Previously Published Works (2014)

Metabolism and ageing are intimately linked. Compared with ad libitum feeding, dietary restriction consistently extends lifespan and delays age-related diseases in evolutionarily diverse organisms. Similar conditions of nutrient limitation and genetic or pharmacological perturbations of nutrient or energy metabolism also have longevity benefits. Recently, several metabolites have been identified that modulate ageing; however, the molecular mechanisms underlying this are largely undefined. Here we show that α-ketoglutarate (α-KG), a tricarboxylic acid cycle intermediate, extends the lifespan of adult Caenorhabditis elegans. ATP synthase subunit β is identified as a novel binding protein of α-KG using a small-molecule target identification strategy termed drug affinity responsive target stability (DARTS). The ATP synthase, also known as complex V of the mitochondrial electron transport chain, is the main cellular energy-generating machinery and is highly conserved throughout evolution. Although complete loss of mitochondrial function is detrimental, partial suppression of the electron transport chain has been shown to extend C. elegans lifespan. We show that α-KG inhibits ATP synthase and, similar to ATP synthase knockdown, inhibition by α-KG leads to reduced ATP content, decreased oxygen consumption, and increased autophagy in both C. elegans and mammalian cells. We provide evidence that the lifespan increase by α-KG requires ATP synthase subunit β and is dependent on target of rapamycin (TOR) downstream. Endogenous α-KG levels are increased on starvation and α-KG does not extend the lifespan of dietary-restricted animals, indicating that α-KG is a key metabolite that mediates longevity by dietary restriction. Our analyses uncover new molecular links between a common metabolite, a universal cellular energy generator and dietary restriction in the regulation of organismal lifespan, thus suggesting new strategies for the prevention and treatment of ageing and age-related diseases.

Article
Peer Reviewed

The metabolite α-ketoglutarate extends lifespan by inhibiting ATP synthase and TOR

UCLA Previously Published Works (2014)

Metabolism and ageing are intimately linked. Compared with ad libitum feeding, dietary restriction consistently extends lifespan and delays age-related diseases in evolutionarily diverse organisms. Similar conditions of nutrient limitation and genetic or pharmacological perturbations of nutrient or energy metabolism also have longevity benefits. Recently, several metabolites have been identified that modulate ageing; however, the molecular mechanisms underlying this are largely undefined. Here we show that α-ketoglutarate (α-KG), a tricarboxylic acid cycle intermediate, extends the lifespan of adult Caenorhabditis elegans. ATP synthase subunit β is identified as a novel binding protein of α-KG using a small-molecule target identification strategy termed drug affinity responsive target stability (DARTS). The ATP synthase, also known as complex V of the mitochondrial electron transport chain, is the main cellular energy-generating machinery and is highly conserved throughout evolution. Although complete loss of mitochondrial function is detrimental, partial suppression of the electron transport chain has been shown to extend C. elegans lifespan. We show that α-KG inhibits ATP synthase and, similar to ATP synthase knockdown, inhibition by α-KG leads to reduced ATP content, decreased oxygen consumption, and increased autophagy in both C. elegans and mammalian cells. We provide evidence that the lifespan increase by α-KG requires ATP synthase subunit β and is dependent on target of rapamycin (TOR) downstream. Endogenous α-KG levels are increased on starvation and α-KG does not extend the lifespan of dietary-restricted animals, indicating that α-KG is a key metabolite that mediates longevity by dietary restriction. Our analyses uncover new molecular links between a common metabolite, a universal cellular energy generator and dietary restriction in the regulation of organismal lifespan, thus suggesting new strategies for the prevention and treatment of ageing and age-related diseases.