Doctor of Philosophy

dissertationThe gastrointestinal (GI) tract is a central player in the regulation of development and aging. FoxA factors are master regulators of the GI tract during embryogenesis and after birth but little is know about their regulation. I use the simple digestive tract of C. elegans to investigate the role of pha-4/FoxA in whole organism responses to nutrients. My search for pha-4/FoxA regulators has lead to insights regarding how an animal responds to nutrients during aging in the adult and postembryonic development. I have examined the role of pha-4/FoxA during adult lifespan extension due to loss of nutrient signaling through the Target of Rapamycin (TOR) pathway. To search for regulators of pha-4/FoxA, Dustin Updike performed a genetic screen to search for suppressors of the larval lethality associated with the pha-4 mutation and discovered an AAA ATPase, ruvb-1, as apotent suppressor. I showed that the ruvb-1 mutant phenocopies mutations in the TOR pathway suggesting that these genes have a similar function in protein biosynthesis. I found that pha-4/FoxA is required for extension of adult lifespan by loss of CeTOR signaling; however, only mutations in one of the predicted CeTOR targets, rsks-l/S6K, required pha-4/FoxA for lifespan regulation. These data suggest a model where TOR signaling through S6 kinase antagonizes pha-4/FoxA factor activity. Secondly, I have investigated the role of pha-4/FoxA during L1 diapause induced by starvation. Proper developmental arrest in response to starvation has been shown to be required for survival however little is known about factors that are required during L1 diapause. I showed that pha-4/FoxA is require d for L1 diapause survival post embryonically. I showed that levels of PHA-4 are important for starvation survival. pha-4/FoxA is required for initiation of development after periods of starvation and is not required for developmental arrest during starvation. We a recurrently using genomewide approaches to determine targets of pha-4/FoxA that a reimportant for L1 diapause recovery. This work investigates how nutrients regulate whole body responses through the GI tract. I find that pha-4/FoxA plays critical roles in coupling low nutrient intake to aging and development

Genome-Wide Identification of Binding Sites Defines Distinct Functions for Caenorhabditis elegans PHA-4/FOXA in Development and Environmental Response

Transcription factors are key components of regulatory networks that control development, as well as the response to environmental stimuli. We have established an experimental pipeline in Caenorhabditis elegans that permits global identification of the binding sites for transcription factors using chromatin immunoprecipitation and deep sequencing. We describe and validate this strategy, and apply it to the transcription factor PHA-4, which plays critical roles in organ development and other cellular processes. We identified thousands of binding sites for PHA-4 during formation of the embryonic pharynx, and also found a role for this factor during the starvation response. Many binding sites were found to shift dramatically between embryos and starved larvae, from developmentally regulated genes to genes involved in metabolism. These results indicate distinct roles for this regulator in two different biological processes and demonstrate the versatility of transcription factors in mediating diverse biological roles.Molecular and Cellular Biolog

The Target of Rapamycin pathway antagonizes pha-4/FoxA to control development and aging

FoxA factors are critical regulators of embryonic development and postembryonic life, but little is know about the upstream pathways that modulate their activity. C. elegans pha-4 encodes a FoxA transcription factor that is required to establish the foregut in embryos and to control growth and longevity after birth. We previously identified the AAA+ ATPase homolog ruvb-1 as a potent suppressor of pha-4 mutations.; Here we show that ruvb-1 is a component of the Target of Rapamycin (TOR) pathway in C. elegans (CeTOR). Both ruvb-1 and let-363/TOR control nucleolar size and promote localization of box C/D snoRNPs to nucleoli, suggesting a role in rRNA maturation. Inactivation of let-363/TOR or ruvb-1 suppresses the lethality associated with reduced pha-4 activity. The CeTOR pathway controls protein homeostasis and also contributes to adult longevity. We find that pha-4 is required to extend adult lifespan in response to reduced CeTOR signaling. Mutations in the predicted CeTOR target rsks-1/S6 kinase or in ife-2/eIF4E also reduce protein biosynthesis and extend lifespan, but only rsks-1 mutations require pha-4 for adult longevity. In addition, rsks-1, but not ife-2, can suppress the larval lethality associated with pha-4 loss-of-function mutations.; The data suggest that pha-4 and the CeTOR pathway antagonize one another to regulate postembryonic development and adult longevity. We suggest a model in which nutrients promote TOR and S6 kinase signaling, which represses pha-4/FoxA, leading to a shorter lifespan. A similar regulatory hierarchy may function in other animals to modulate metabolism, longevity, or disease