O-linked-N-acetyl glucosamine (O-GlcNAc) is a post-translational modification implicated in cellular processes including transcription, translation, proteasome activity, and the stress response. Over 500 proteins are O-GlcNAc modified, including components of the insulin signaling pathway. Over expression of the O-GlcNAc transferase OGT-1 or inhibition of the O-GlcNAcase OGA-1 causes insulin resistance in mammalian cells, and OGA-1 is a human type II diabetes susceptibility locus. Glucose levels influence O-GlcNAc modification levels, suggesting that this pathway acts as a nutrient sensor. Although these enzymes are essential in mammals and highly conserved in C. elegans, knockouts of
oga-1 and
ogt-1 are viable in the worm, making it a good model system for genetic studies of the O-GlcNAc modification and its effect on nutrient and insulin signaling. Previous work in our laboratory demonstrated that loss of OGT-1 in a
daf-2 mutant partially suppresses dauer formation. We find that loss of OGA-1 results in a similar decrease in dauer formation in
daf-2 mutants, suggesting that O-GlcNAc cycling is important in insulin signaling. We also find that dauer formation in
daf-2 ogt-1 and
daf-2;
oga-1 double mutants is DAF-16 dependent. In order to define the role of O-GlcNAc modification in insulin signaling, we tested how
ogt-1 and
oga-1 mutations affect other DAF-16-dependent
daf-2 phenotypes. We find that
oga-1 and
ogt-1 have different effects on different
daf-2 phenotypes: although both mutations suppress dauer formation, they have no effect on
daf-2 fertility and have opposing effects on
daf-2 lifespan (collaboration with CA Wolkow). Since the mammalian DAF-16 homolog FOXO1 is O-GlcNAc modified, we are testing whether DAF-16 is O-GlcNAc modified and how the modification could affect its localization, activity, and transcriptional program. Since O-GlcNAc levels respond to nutrient levels, we also tested whether this pathway is involved in the response to excess nutrition. We find that
ogt-1 mutants, but not
oga-1 or
daf-2 mutants, show decreased fertility on high glucose (3-5X reduced vs. N2). These data suggest that O-GlcNAc modification, but not insulin signaling, is necessary for the response to nutrient stress. However, high glucose suppresses dauer formation in
daf-2,
daf-2 ogt-1 and
daf-2;
oga-1 mutants, but not in the TGF-b mutants
daf-1 and
daf-7, indicating that this effect is specific to insulin signaling but independent of O-GlcNAc. We plan to exploit this phenotype in a genetic screen for novel factors that connect nutrient stress and insulin signaling.