Oral Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Subcellular distribution of post-translational modifications in human proteome (#13)

Anna Andrejeva 1 , Tom S Smith 1 , Rayner M L Queiroz 1 , Eneko Villanueva 1 , Kathryn Lilley 1
  1. University of Cambridge, Cambridge, CAMBRIDGESHIRE, United Kingdom

Protein post-translational modifications (PTMs) regulate many processes, including subcellular localisation, protein function and protein-protein interaction. Phosphorylation on serine, threonine and tyrosine remains the most studied PTM but more than 200 PTMs have been discovered. Lysine succinylation is a relatively understudied modification despite it being widespread and expected to have large effect on protein function and activity through the change in lysine charge.1,2 Succinylation of mitochondrial proteins may be explained by the production of succinyl-CoA as an intermediate in the mitochondrial citric acid cycle. Yet, so far there is no study addressing the true subcellular localisation of succinylated proteins and the hypothesised correlation of succinyl-CoA subcellular concentration and protein succinylation. Furthermore, the discovery of biologically relevant PTMs is greatly assisted through the establishment of their subcellular distributions, especially where the modified proteins are found at distinct locations from their unmodified counterparts.

We have pioneered the development of the novel high-resolution subcellular proteomics technique LOPIT (Localisation of Organelle Proteins by Isotope Tagging), as well as machine‑learning based approaches to address protein high‑throughput subcellular classification.3,4 LOPIT-DC utilises 10-plex TMT tags to quantify peptide profiles over cellular fractions and enables protein classification to subcellular niches.5 In the work presented here, we combine LOPIT-DC with sequential enrichment of Lysine succinylated and phosphorylated peptides to address fundamental questions including: Which modifications co‑occur with differential localisation? Where in the cell are the proteins with stable succinylated sites?

Here we present data of subcellular localisation of the phosphorylated and succinylated peptides alongside the unmodified proteome derived from the same sample. Our preliminary experiments suggest that the majority of succinylated proteins are not mitochondrial and identify succinylated proteins which are differently located relative to their unmodified proteins. This study will be critical to further our understanding of the role of these modifications and their interplay on protein localisation.

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