Dhondt, Ineke et al. (2015) International Worm Meeting "SILeNCe is golden: slow-down of protein turnover in the long-lived Caenorhabditis elegans daf-2 mutant."

Petyuk, Vladislav A., Dhondt, Ineke, Braeckman, Bart P., Smith, Richard D., Depuydt, Geert

[International Worm Meeting, 2015]

Most aging hypotheses revolve around the accumulation of some sort of damage resulting in gradual physiological decline and ultimately death. Avoiding protein damage accumulation by enhanced turnover should slow down the aging process and extend lifespan. However, lowering translational efficiency extends rather than shortens lifespan in C. elegans. We studied turnover of individual proteins in the conserved Insulin/Insulin-like Growth Factor (IGF-1) receptor mutant daf-2 by combining Stable Isotope Labeling by Nitrogen in Caenorhabditis elegans (SILeNCe) and accurate mass spectrometry. Intriguingly, the majority of proteins displayed prolonged half-lives in daf-2, while others remained unchanged, signifying that longevity is not supported by high protein turnover. This slow-down of protein turnover was most prominent for components of the translation machinery and mitochondria. In contrast, the high turnover of lysosomal hydrolases and very low turnover of cytoskeletal proteins remained largely unchanged in daf-2. The slow-down of protein dynamics and low abundance of the translational machinery may point at the importance of anabolic attenuation in lifespan extension as suggested by the hyperfunction theory.

Wang, Wenqing et al. (2015) International Worm Meeting "Metabolic profiling and metabolic flux analyses in NAD+ salvage biosynthesis mutants."

Hanna-Rose, Wendy, Pacella, Marisa, McReynolds, Melanie, Braeckman, Bart, Detwiler, Ariana, Wang, Wenqing, Dhondt, Ineke, Lange, Stephanie, Goncalves, Jimmy, Shu, Muya, Kho, Kelvin

[International Worm Meeting, 2015]

Temporal developmental progression is highly coordinated in C. elegans. However, loss of nicotinamidase PNC-1 activity slows reproductive development, uncoupling it from its typical temporal progression relative to the soma. Nicotinamidases mediate salvage NAD+ synthesis. There has been significant recent interest in boosting NAD+ biosynthesis for therapeutic benefit in aging-related diseases and inhibiting biosynthesis for therapeutic benefit in cancer. However, we haven't fully elucidated how perturbing the availability of NAD+ impacts whole organism physiology. Using LC-MS we show that pnc-1 mutants do not salvage nicotinamide released by NAD+ consumers to resynthesize NAD+, and as a result NAD+ availability is reduced. By manipulating NAD+ levels using genetics, we show that a reduction in NAD+ bioavailability is incompatible with a normal pace of gonad development. We use the pnc-1 mutant phenotype to explore physiological requirements for reproductive development. First we demonstrate that the deficit in NAD+ production compromises NAD+ consumer activity as would be expected. Even though effects on NAD+ consumer activity have been the primary focus for probing the functional consequences of manipulation of NAD+ bioavailability, we revealed no functional link between loss of NAD+ consumer activity and reproductive development in genetic experiments. We used metabolomics profiling to seek explanations for the observed phenotype. The deficit in NAD+ availability from lack of salvage biosynthesis has wide-spread metabolic consequences, including perturbations in glycolysis. Using flux analysis, we demonstrate that glycolysis is blocked at the NAD+-dependent step, and we functionally link the block to the reproductive phenotype using genetics and pharmacological approaches. Interestingly, mitochondria are protected from the deficiency in NAD+ biosynthesis and the effects of reduced glycolytic output, also revealing compartment specific regulation of NAD+ biosynthesis. We suggest that compensatory metabolic processes that maintain mitochondrial activity in the absence of efficient glycolysis are incompatible with the cell division requirements for reproductive development. Our work has implications for understanding mechanisms that mediate therapies that target NAD+ salvage synthesis for the purposes of inhibiting tumor growth.

Pradhan, Sigma et al. (2021) International Worm Meeting "The role of parental diet on progeny's proteome and fitness"

Towbin, Benjamin, Pradhan, Sigma

[International Worm Meeting, 2021]

Animals adjust their phenotype to their environment, which is thought to provide a selective benefit under changing conditions. A famous example of such phenotypic plasticity is the slow down of growth and the delay of aging under dietary restriction (DR). Interestingly, parental DR also affects phenotypes of the F1 generation, for example, DR increases the starvation resistance of progeny (Jordan et al., 2019). Molecularly, DR is associated with a global change in proteome expression, including a reduction in ribosome levels and a proteome-wide slow-down of protein turnover (Dhondt et al., 2016; Visscher et al., 2016). Here, we ask if and how parental diet impacts the proteome of the F1 progeny. We will seek causal relationships between intergenerationally transmitted phenotypes and the inherited proteome. Finally, we aim to determine if the parental control of the proteome provides a selective benefit to their progeny. To address these questions, we have established a tool to precisely quantify ribosomal levels by measuring the luminescence from lysis of HiBit tagged to ribosomal proteins. We plan to globally analyze parental effects on the F1 proteome by proteomics and measure the dynamics of proteome changes, growth rates, and reproduction rates in different nutritional conditions by fluorescence live imaging in microchambers. The project will likely provide important insights about the molecular mechanisms and physiological significance of inter-generational inheritance induced by parental diet.