Feasibility of phosphoproteomics on left-over sample after RNA extraction (RNA-Bee)

In daily practice biomolecules are usually extracted with their own specific protocols for downstream omics analysis. However, when sample material is limited, an all-in-one strategy is preferable. DNA, RNA and proteins can be isolated with phenol guanidine thiocyanate based extraction, yet lysis with Urea is the accepted standard for phosphoproteomic applications. Here we compared Urea with RNA-Bee mediated protein extraction for use in mass spectrometry (MS) based phosphoproteomics analysis of cells and tissues. We profile the DNA damage response after ionizing irradiation of U2OS cells as proof of principle for cultured human cells as well as mouse liver and three different human tissues. We show high overlap and similarity of phosphosite data and kinase activity for RNA-bee extraction when compared to standard Urea approach. Phenol guanidine thiocyanate lysis might thus be an appropriate way for multi-omics or MS profiling workflows in order to obtain several data types from a single sample.