Nutrient Flux and Development

National Institute Of Diabetes And Digestive And Kidney Diseases

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Abstract

A dynamic cycle of O-linked N-acetylglucosamine (O-GlcNAc) addition and removal acts on nuclear pore proteins, transcription factors, and kinases to modulate cellular signaling cascades. This nutrient sensing hexosamine signaling pathway is conserved from nematodes to man. A single nucleotide polymorphism in the human O-GlcNAcase gene is linked to type 2 diabetes, suggesting that perturbation of this pathway results in disease. In collaboration the Hanover lab (NIDDK), we showed that the C. elegans genome encodes the two evolutionarily conserved enzymes that mediate O-GlcNAc cycling, with the genes called ogt-1 and oga-1. We previously characterized a knockout alleles of ogt-1 and oga-1 genes. Using a combination of genomic expression arrays and chromatin immunoprecipitation (ChIP) we are looking for genes that respond to nutrient flux differently in the mutants with the hope of identifying pathways of importance. The expression analysis has revealed widespread de-regulation of gene expression in the mutants, identify affected pathways including longevity and aging. We have tested these pathways in the mutants and find alteration in function that are consistent with the gene expression patterns we observe. From the ChIP studies, we have identified a discrete number of genes associated with O-GlcNAcylated proteins. These associations are pronounced at the promoters of the genes and show some overlap with ChIP signals using RNA PolII antibodies. We are currently investigating the function role, if any, of these restricted O-GlcNAc chromatin marks. These marks have the potential to link nutritional flux in the cell directly to gene regulation, offering a novel insight into the role of O-GlcNAc cycling in animal physiology and development. In collaboration with the Notkins lab (NIDCR) we have followed up on our earlier studies of a protein called IA-2. This membrane protein associated with dense core vesicles plays an important role in neurosecretion from a variety of tissues. Using an in vivo reporter for IA-2, we have screened for C. elegans mutants that can not properly control IA-2 levels. One such mutant turned out to be in the gene called pag-3 encoding a transcription factor. Our studies have demonstrated that mis-regulation of IA-2 in pag-3 mutants results in a disruption of dense core vesicle synthesis or stability resulting in neurosecretory defects. Our work demonstrated a novel layer of transcriptional regulation impacting dense core vesicle dynamics with implications for diseases of neurosecretion, including diabetes.