Project Summary
Cellular stress responses play essential roles in cell and organismal survival and contribute to a
wide range of physiological processes and diseases in humans. The molecular architecture of
most stress response pathways are well defined. A striking exception to this is osmotic stress
response, where the relevant stress sensors and signaling mechanisms in animals are poorly
understood. Most studies of the osmotic stress response use cultured cells, where in vivo
complexities, i.e. the extracellular matrix, tissue mechanical properties, etc., are not replicated.
To better mimic these conditions, we study the osmotic stress response in a live animal, the
nematode C. elegans. Like humans, C. elegans responds to osmotic stress by metabolizing
glucose to produce organic osmolytes, such as glycerol. We performed an unbiased forward
genetic screen to identify mutants that exhibit no induction of osmolyte biosynthesis genes (Nio
genes) and discovered multiple alleles of nio-2, which encodes the sole C. elegans homolog of
the O-GlcNAc transferase (ogt-1; OGT in humans). OGT post-translationally O-GlcNAcylates
Ser/Thr residues of cytosolic and nuclear proteins but also exhibits important GlcNAcylation
independent functions. Mammalian cells lacking OGT do not survive, but C. elegans lacking ogt-
1 are viable and fertile, providing a unique opportunity to study the role of ogt-1 in cellular
physiology. ogt-1 mutants are unable to adapt and grow in hypertonic environments and exhibit
reduced organic osmolyte levels and no induction of the osmolyte biosynthesis protein GPDH-1.
However, osmotic induction of osmolyte biosynthesis gene mRNAs is normal, suggesting that
ogt-1 functions post-transcriptionally. These defects can be rescued by expression of wild type
or catalytically inactive human OGT, showing that non-canonical functions of OGT in the osmotic
stress response are conserved from C. elegans to humans. We also discovered mutations in
interacting components of a conserved 3’ mRNA processing complex that phenocopy ogt-1. We
hypothesize that non-canonical functions of ogt-1 facilitate upregulation of stress-induced mRNA
translation via interactions with 3’ RNA processing complex proteins during osmotic stress. To
test this hypothesis, we will determine the temporal, functional, and regulatory requirements for
ogt-1 in the osmotic stress response (Aim 1), identify the specific gene expression mechanism(s)
that are affected by ogt-1 (Aim 2), and determine if ogt-1 regulates the osmotic stress response
via interactions with 3’ mRNA cleavage and polyadenylation components also identified in our Nio
screen. (Aim 3). Our studies will delineate a novel paradigm in stress signaling and reveal new
mechanisms by which OGT impacts cell physiology.
