Protein phosphorylation, encompassing the addition or removal of phosphate moieties on the functional groups of amino acid side chains (typically Ser, Thr, Tyr), is ubiquitously found in eukaryotes and revealed to be critical for growth, adaptation and survival in photosynthetic organisms. Catalyzed by a class of enzymes known as kinases, these phosphorylation events serve as activators of protein function within the cell through induction of conformational change, and these kinases form networks that propagate phosphorylation signals through specific biological pathways as a mechanism for generating cellular responses from various stimuli.
The study of these phosphorylation events in the cellular proteome, or phosphoproteomics, in photosynthetic organisms has the potential to not only inform on how life is sustained but also how an organism responds to their environment, with insights into broad ranging fields from agriculture, biofuels, and medicine. Our lab seeks to elucidate these phosphorylation events through complementary in vivo and in vitro platforms that utilize enrichment strategies specific to phosphorylation coupled with liquid chromatography and high mass accuracy, high resolution mass spectrometry (LC-MS) for broad, quantitative coverage of the phosphoproteome.
To assess changes caused by chemical inhibition of protein kinases and the subsequent effect on the phosphoproteome, the in vivo quantitative phosphoproteomics platform aims to detect and quantify phosphorylation status occurring on the -omics level in response to treatments. Through investigating the modulation of phosphosites found on direct and indirect kinase targets, the objective is to gain insight into signaling pathways and their involvement in specific cellular functions. To identify the direct targets of specific kinases, the in vitro kinase screening platform aims to detect newly generated phosphopeptides and their associated proteins in the natural protein complement screened against individual active kinases. In combination, these two approaches will allow for a comprehensive understanding of signaling pathways within photosynthetic organisms and identify substrates regulated by kinase activity.
Couso, I., Smythers, A. L., Ford, M. M., Umen, J. G., Crespo, J. L., Hicks, L. M. Inositol polyphosphates and target of rapamycin kinase signalling govern photosystem II protein phosphorylation and photosynthetic function under light stress in Chlamydomonas. New Phytologist, 232, 5, 2011-2025, December 2021.
Iannetta, A. A., Minton, N. E., Uitenbroek, A. A., Little, J. L., Stanton, C. R., Kristich, C. J., Hicks, L. M. IreK-Mediated, Cell Wall-Protective Phosphorylation in Enterococcus faecalis. Journal of Proteome Research, 20, 11, 5131–5144, October 2021.
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