Poster Presentation 25th Annual Lorne Proteomics Symposium 2020

Redox modifications of cysteine in the liver of type 2 diabetes mellitus (#110)

Desmond K Li 1 2 , Alexander W Rookyard 2 3 , Lauren E Smith 1 2 , Yen Chin Koay 2 4 , Holly McEwan 2 5 , Anthony Don 2 5 , John F O'Sullivan 2 4 , Stuart J Cordwell 1 2 3 6 , Melanie Y White 1 2 3
  1. Discipline of Pathology, School of Medical Sciences, The University of Sydney, Sydney, NSW, Australia
  2. Charles Perkins Centre, The University of Sydney, Sydney, NSW, Australia
  3. School of Life and Environmental Sciences, The University of Sydney, Sydney, NSW, Australia
  4. Heart Research Institute, Sydney, NSW, Australia
  5. ACRF Centenary Cancer Research Centre, Sydney, NSW, Australia
  6. Sydney Mass Spectrometry, The University of Sydney, Sydney, NSW, Australia

Reactive oxygen species (ROS) and associated redox modifications have normal physiological roles in signalling but are also implicated in a number of pathologies. Oxidative stress associated with type 2 diabetes mellitus (T2DM) has been attributed to increased generation/reduced clearance of ROS in insulin sensitive tissue including the liver. During oxidative stress, proteins undergo redox modifications through the thiols of cysteine, which may alter structure, functionality and signalling pathways. To determine proteins of the liver which undergo redox modifications with T2DM, we investigated a number of associated metabolic disorders using redox proteomics. Rats were subjected to a high fat diet (HFD) to induce peripheral insulin resistance, as well as streptozotocin (STZ) to create β cell dysfunction, thereby generating the T2DM pathology. Rats treated with only HFD or STZ were utilised to analyse the pre-diabetic state. To enrich low abundance redox modified peptides, thiol disulfide exchange was utilised for reversibly modified cysteines with quantitation by iTRAQ and mass spectrometry (MS). Strong cation exchange was used to select for irreversibly modified cysteines, combined with MS based label-free parallel reaction monitoring (PRM) quantitation. We identified 7,635 peptides with reversibly oxidised cysteines, with over 3,000 changing abundance in response to disease progression and quantified 123 PRM amenable peptides with irreversibly oxidised cysteine. Oxidative modifications to proteins which maintain redox balance suggests dysregulation, contributing to oxidative stress in T2DM. Observed changes in enzymes involved in glucose metabolism suggest a role of redox modifications in glucose utilisation/production. Modifications to enzymes in fatty acid metabolism correlate with the hyperlipidemia associated with T2DM. Many proteins showed increased levels of irreversible oxidative modifications contributing to protein damage. Changes in the redox status of these proteins could be indicative of these protein species prone to oxidative damage, contributing to pathogenesis or a level of cellular regulation in response to the metabolic disturbance.