An evolutionary proteomics approach identifies substrates of the cAMP-dependent protein kinase
- 19 September 2005
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 102 (39), 13933-13938
- https://doi.org/10.1073/pnas.0501046102
Abstract
Protein kinases are important mediators of much of the signal transduction that occurs in eukaryotic cells. Unfortunately, the identification of protein kinase substrates has proven to be a difficult task, and we generally know few, if any, of the physiologically relevant targets of any particular kinase. Here, we describe a sequence-based approach that simplified this substrate identification process for the cAMP-dependent protein kinase (PKA) in Saccharomyces cerevisiae. In this method, the evolutionary conservation of all PKA consensus sites in the S. cerevisiae proteome was systematically assessed within a group of related yeasts. The basic premise was that a higher degree of conservation would identify those sites that are functional in vivo. This method identified 44 candidate PKA substrates, 5 of which had been described. A phosphorylation analysis showed that all of the identified candidates were phosphorylated by PKA and that the likelihood of phosphorylation was strongly correlated with the degree of target site conservation. Finally, as proof of principle, the activity of one particular target, Atg1, a key regulator of autophagy, was shown to be controlled by PKA phosphorylation in vivo. These data therefore suggest that this evolutionary proteomics approach identified a number of PKA substrates that had not been uncovered by other methods. Moreover, these data show how this approach could be generally used to identify the physiologically relevant occurrences of any protein motif identified in a eukaryotic proteome.Keywords
This publication has 52 references indexed in Scilit:
- The Ras/cAMP-dependent Protein Kinase Signaling Pathway Regulates an Early Step of the Autophagy Process in Saccharomyces cerevisiaePublished by Elsevier BV ,2004
- Yeast autophagosomes: de novo formation of a membrane structureTrends in Cell Biology, 2002
- The C Terminus of the Vps34p Phosphoinositide 3-Kinase Is Necessary and Sufficient for the Interaction with the Vps15p Protein KinasePublished by Elsevier BV ,2002
- Convergence of Multiple Autophagy and Cytoplasm to Vacuole Targeting Components to a Perivacuolar Membrane Compartment Prior tode Novo Vesicle FormationPublished by Elsevier BV ,2002
- Molecular Evidence for the Early Colonization of Land by Fungi and PlantsScience, 2001
- The Vesicle Transport Protein Vps33p Is an ATP-binding Protein That Localizes to the Cytosol in an Energy-dependent MannerPublished by Elsevier BV ,1998
- Novel System for Monitoring Autophagy in the Yeast Saccharomyces cerevisiaeBiochemical and Biophysical Research Communications, 1995
- Use of an oriented peptide library to determine the optimal substrates of protein kinasesCurrent Biology, 1994
- Isolation and characterization of autophagy‐defective mutants of Saccharomyces cerevisiaeFEBS Letters, 1993
- Embryonic ε and γ globin genes of a prosimian primate (Galago crassicaudatus)Journal of Molecular Biology, 1988