METHOD DEVELOPMENT FOR MEASURING RELATIVE CHANGES IN PROTEIN LEVEL

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are developing metabolic labeling methodologies for measuring relative changes in protein level. The initial application of this technology will be in the measurement of changes in protein level between the daf-2 single mutant and the daf-2; daf-16 double mutant of C. elegans. In C. elegans the insulin/IGF-like pathway regulates metabolism, growth and longevity. Weak loss-of-function mutations in daf-2, the insulin/IGF-like receptor, have been found to have an increased lifespan. DAF-2 ultimately acts to inhibit the DAF-16 transcription factor downstream in the pathway. Without DAF-2 inhibition, DAF-16 regulates the increase of fat and glycogen storage. The increased lifespan phenotype of the daf-2 mutant is thought to be a result of caloric restriction and requires wild-type DAF-16. Hence, differences between the level of proteins in the long-lived single daf-2 mutant and the normal-lived daf-16; daf-2 double mutant should reveal proteins that are transcriptionally regulated by DAF-16 and therefore may be involved in the process of aging. The insulin/IGF-like signaling pathway is highly conserved between higher eukaryotes and therefore this approach will likely be valuable for studying insulin signaling and sugar metabolism in other organisms.