Untangling the Dynamics of Lysine Acetylation and Phosphorylation in Adipogenesis in the Established Human and Mouse Adipocyte Cell Lines SGBS and 3T3L1

Obesity is one of the most pressing global public health challenges of our time and prevalence continues to increase. It is characterized by enlarged and dysfunctional adipose tissue and is often associated with the development of metabolic or cardiovascular comorbidities. Post-translational protein modifications (PTMs) are important determinants of health and disease and enable vibrant cellular signalling. However, sound knowledge about how PTMs affect adipogenesis is still scarce. In addition to protein phosphorylation, which plays a crucial role in insulin signalling, lysine acetylation provides a direct link between cellular metabolism and signal transduction and is therefore of particular importance for the study of metabolic diseases. Stable cell lines are an integral part of research into the development and physiology of adipocytes in health and disease and various models have been used to date. Here, we investigate the temporal dynamics of the transcriptome, proteome, the levels of central carbon metabolites as well as the acetyl and phosphoproteome in adipogenesis using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in combination with PTM enrichment strategies on the established human (SGBS) and mouse (3T3-L1) adipocyte cell culture models. Both cell lines showed unique temporal PTM profiles during adipogenesis, with a large number of acetylated and phosphorylated sites being significantly regulated. Enriched pathways of acetylated proteins were predominantly associated with central energy metabolism, while phosphorylated proteins were enriched in the areas of insulin/glucagon signalling, cell adhesion processes and organization of cellular structure. We identified several key driver candidates with strong correlation to the time course of adipogenesis, including CD44 and the acetylation site FASN K673 (negative) or IDH K272 (positive). The general agreement between results from SGBS and 3T3L1 cells was high, however details appeared cell line specific. We provide a detailed data resource on the transcriptome, proteome, the levels of central carbon metabolites, the acetylome and phosphoproteome during adipogenesis of the established human and mouse adipocyte cell lines SGBS and 3T3L1, accessible for further mining. Human SGBS cells were provided by the laboratory of Prof. Dr. Wabitsch at the University Clinic Ulm. Cells were differentiated according to the standard protocol described previously (Wabitsch et al., 2001). The cell line was maintained under 5 % CO2 at 37°C and 95 % humidity. Cell culture experiments for transcriptomic (SGBS) data analysis were prepared in four replicates. Differential expression was determined with the R package DESeq2 (Love et al., 2014).