Post translational modifications in the type 2 diabetic liver and their contribution to the diseased state — ASN Events
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Post translational modifications in the type 2 diabetic liver and their contribution to the diseased state (#203)

Desmond K Li 1 2 3 , Lauren E Smith 1 3 , Joel A Cain 2 3 , Jana Paulech 2 , Stuart J Cordwell 1 2 3 4 , Melanie Y White 1 2 3
  1. Discipline of Pathology, School of Medical Sciences, The University of Sydney, Sydney, NSW, Australia
  2. School of Life and Environmental Sciences, The University of Sydney, Sydney, NSW, Australia
  3. Charles Perkins Centre, The University of Sydney, Sydney, NSW, Australia
  4. Mass Spectrometry Core Facility, The University of Sydney, Sydney, NSW, Australia

Insulin resistance and oxidative stress are hallmarks of type 2 diabetes (T2D), commonly observed in insulin sensitive tissues including the liver. Insulin resistance has been attributed to impaired insulin signalling which normally promotes glucose uptake and utilisation through the IRS/PI3K pathway. Oxidative stress involves the increased generation/reduced clearance of reactive oxygen species (ROS), with high levels of ROS oxidising proteins producing redox modifications which may alter structure, functionality and signalling pathways. To investigate the changes in signalling mediated by phosphorylation and/or redox modifications with T2D, we investigated a rat model using global phosphoproteomics and redox proteomics. Rats were subjected to a high fat diet and streptozotocin to generate the T2D pathology. To quantify alterations in PTM status, samples were isobarically tagged and subjected to various enrichment strategies prior to mass spectrometry (MS). Phosphopeptides were enriched by a combination of TiO2 and IMAC while redox modified peptides were enriched by thiol disulfide exchange for reversibly oxidised cysteines. Enriched and non-captured fractions were subjected to hydrophilic interaction chromatography prior to MS analysis. The insulin signalling pathway showed significant reduction in phosphorylation confirming impairment in T2D. Pathways regulating energy metabolism also showed significant perturbations in phosphorylation, possibly in response to or contributing to the energy imbalance. Oxidative modifications to the insulin receptor suggests contribution of redox modifications to insulin resistance, while modifications to proteins which maintain redox balance suggests dysregulation, contributing to the oxidative stress in T2D. The current study has identified changes in protein phosphorylation and redox state indicative of dysregulated signalling and energy utilisation while isolating species prone to oxidative damage, all able to contribute to the pathogenesis of T2D.