Leibniz-Forschungsinstitut für Molekulare Pharmakologie
Inositol Phosphates and Protein Pyrophosphorylation &emdash; Challenges in Signal Transduction
Protein phosphorylation is one of the most commonly used mechanisms in signal transduction. Beyond phosphorylation, protein pyrophosphorylation – a posttranslational modification mediated by inositol pyrophosphate messengers – emerged over ten years ago as a potential additional layer of regulation for cellular signal transduction cascades. 1 However, while the inositol pyrophosphates have been linked to numerous important cellular processes, 2 the function of protein pyrophosphorylation remains poorly characterized because suitable reagents and detection methods are lacking.
Our group recently developed a mass spectrometry, MS, method to directly detect and identify pyrophosphopeptides in complex samples. 3 Excitingly, this MS method has enabled us to uncover the first endogenous pyrophosphorylation sites from mammalian cell lysates. The triggered MS method allowed for the definitive assignment of over 30 sites. The modified proteins span a range of processes and compartments and included histone deacetylase 2, HDAC2, and cytoplasmic phosphoglucomutase 1, PGM1. Our efforts to now to illuminate the effects of this modification on protein structure and function will also be discussed.
In sum, our novel MS method for the analysis of endogenous pyrophosphopeptides will enable the systems-wide study of this modification in a cellular context. Complemented by chemical tools for the generation of stoichiometrically pyrophosphorylated proteins, 4 the function and regulation of this poorly annotated PTM can be elucidated in the future.
Dorothea Fiedler 1,2, Jeremy Morgan 1, Robert Harmel 1,2, Eberhard Krause 1, Fan Liu 1
1. Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany
2. Humboldt Universität zu Berlin, Department of Chemistry, Brook-Taylor-Straße 2, 12489 Berlin, Germany
1. Bhandari, R. et al. Protein pyrophosphorylation by inositol pyrophosphates is a posttranslational event, Proc. Natl. Acad. Sci. U. S. A., 2017 104, 15305–15310
2. Chakraborty, A., Kim, S. & Snyder, S. H. Inositol pyrophosphates as mammalian cell signals, Sci. Signal, 2011 4, re1
3. Penkert, M. et al. Unambiguous identification of serine and threonine pyrophosphorylation using neutral-loss-triggered electron-transfer/higher-energy collision dissociation, Anal. Chem., 2017 89, 3672-3680
4. Marmelstein, A. M. et al. Pyrophosphorylation via selective phosphoprotein derivatization, Chem. Sci., 2018 9, 5929-5936