O-GlcNAc levels are regulated in a tissue and time specific manner during post-natal development, independently of dietary intake

Abstract

O-GlcNAcylation is a well-preserved, ubiquitous, post-translational modification involved in response to stress, considered as a nutrient sensor. GlcNAc moiety is added and removed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), respectively. Metabolic sources vary from carbohydrate in utero, to fatty acid after birth then a mix once adult. Study the evolution of O-GlcNAc levels during the first period of life and decipher the impact of O-GlcNAc on cardiac proteins. Heart and brain were harvested from Wistar rats before and after birth (D-1 and D0), in suckling animals (D12), after weaning (D28) with a standard or a carbohydrate free diet, and when adults (D84). O-GlcNAc levels, GFAT, OGT and OGA protein expression were evaluated by western blot. Mass spectrometry approaches were performed to (i) quantify known regulator of O-GlcNAc and (ii) identify cardiac O-GlcNAcylated proteins at D0 and D28. Cardiac protein O-GlcNAc levels decrease progressively from D-1 to D84 (13 fold, P < 0.05), whereas protein O-GlcNAc levels tend to increase in the brain. Both cardiac and brain protein O-GlcNAc levels were similar between the two weaning diets. Moreover, levels of UDP-GlcNAc and glutamine are not correlated with changes in O-GlcNAc levels. Mass spectrometry analysis allowed us to identify proteins of interest involved in cardiac stress response (including those implicated in protein and RNA biosynthesis, metabolism). Interestingly, several proteins are only O-GlcNAcylated at D28 despite a decrease in O-GlcNAc levels between D0 and D28. We demonstrate for the first time that O-GlcNAc levels are not linked to the dietary sources. Our results suggest that glutamine and UDP-GlcNAc are not the main regulatory element of O-GlcNAc levels. Proteins identified by untargeted mass spectrometry will need to be specifically studied to better understand the impact of O-GlcNAcylation during development, growth or stress response.