Profiling molecular changes to discover new drivers of aging in C. elegans [electronic resource]

Nearly all organisms age and die, but the molecular causes of aging and the factors that determine lifespan are not well understood. Using the model system Caenorhabditis elegans, we have taken the general approach of measuring molecular changes with age, identifying the upstream processes driving these changes, and determining the downstream effects on lifespan. This strategy allows us to identify ways to extend lifespan by reversing detrimental aging changes or enhancing natural protective pathways. In addition, we gain a greater understanding of the mechanisms that drive the normal aging process. In the first part of this work, we identify a conserved C. elegans GATA transcription factor/MTA-1 homolog egr-1 (lin-40) that extends lifespan and promotes resistance to heat and UV stress when overexpressed. Expression of egr-1 increases with age, suggesting that it may function to promote survival during normal aging. This increase in expression is suppressed in germline deficient worms, indicating that egr-1 responds to signals from the germline, and that changes in the germline with age could be a cause of its increasing expression. In addition, we show that egr-1 acts within the insulin signaling pathway and can activate the expression of its paralog egl-27, another factor known to extend lifespan and increase stress resistance. These results identify egr-1 as part of a longevity-promoting circuit that changes with age in a manner that is beneficial for the lifespan of the organism. In the second part of this work, we expand the repertoire of C. elegans inducible systems by adapting destabilization domains (DDs) to C. elegans. C. elegans is a widely used aging model system due to its short lifespan and large genetic and molecular toolkit. However, there are relatively few options for inducible gene expression in this model, and none that work directly at the protein level. DDs are conditionally unstable domains that can be fused to a protein of interest, causing it to be degraded in the absence of stabilizing ligand. We show that DDs engineered for use at room temperature can be used to regulate protein concentrations in C. elegans in a rapid, reversible, and dose-dependent manner. In the last section of this work, we describe the results of a mass-spectrometry based proteomics study to profile changes in protein abundance with age in C. elegans. Assessing changes in the aging proteome directly is important because aging involves dysregulated protein homeostasis: reduced protein synthesis, degradation, and folding capacity, and increased proteome insolubility and protein damage. Using this unbiased approach, we identify a previously uncharacterized class of secreted proteins expressed in the adult uterus that dramatically increase in abundance with age. We find that uterine proteins are not turned over in post-reproductive old animals or in young worms that lack a vulva and cannot lay eggs. Furthermore, the age-dependent accumulation of uterine proteins is partially suppressed in animals with an extended reproductive period, and accelerated in sterile animals that do not lay eggs. In total, these results indicate that age-induced infertility contributes to extracellular protein accumulation in the uterus with age. Finally, we show that knocking down multiple age-increased proteins simultaneously extends lifespan, and overexpression of uterine proteins shortens lifespan. These results provide a mechanistic example of how the cessation of reproduction contributes to detrimental changes in the soma, and demonstrate how the timing of reproductive decline influences the rate of aging.

Keywords: Genetics.
Advisor: Kim Stuart
Brunet Anne
Elias Joshua
Fire Andrew Zachary advisor. Thesis advisor