1. SILAC-based phosphoproteomics reveals new PP2A-Cdc55-regulated processes in budding yeast
- Author
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Ministerio de Ciencia e Innovación (España), Ministerio de Economía y Competitividad (España), European Commission, Lundbeck Foundation, Villum Fonden, Instituto de Salud Carlos III, University of Southern Denmark, Baro, Barbara, Játiva, Soraya, Calabria, Inés, Vinaixa, Judith, Bech-Serra, Joan Josep, LaTorre, Carolina de, Rodrigues, João, Hernáez, María Luisa, Gil, Concha, Barceló-Batllori, Silvia, Larsen, Martin R., Queralt, Ethel, Ministerio de Ciencia e Innovación (España), Ministerio de Economía y Competitividad (España), European Commission, Lundbeck Foundation, Villum Fonden, Instituto de Salud Carlos III, University of Southern Denmark, Baro, Barbara, Játiva, Soraya, Calabria, Inés, Vinaixa, Judith, Bech-Serra, Joan Josep, LaTorre, Carolina de, Rodrigues, João, Hernáez, María Luisa, Gil, Concha, Barceló-Batllori, Silvia, Larsen, Martin R., and Queralt, Ethel
- Abstract
Background: Protein phosphatase 2A (PP2A) is a family of conserved serine/threonine phosphatases involved in several essential aspects of cell growth and proliferation. PP2ACdc55 phosphatase has been extensively related to cell cycle events in budding yeast; however, few PP2ACdc55 substrates have been identified. Here, we performed a quantitative mass spectrometry approach to reveal new substrates of PP2ACdc55 phosphatase and new PP2A-related processes in mitotic arrested cells. Results: We identified 62 statistically significant PP2ACdc55 substrates involved mainly in actin-cytoskeleton organization. In addition, we validated new PP2ACdc55 substrates such as Slk19 and Lte1, involved in early and late anaphase pathways, and Zeo1, a component of the cell wall integrity pathway. Finally, we constructed docking models of Cdc55 and its substrate Mob1. We found that the predominant interface on Cdc55 is mediated by a protruding loop consisting of residues 84-90, thus highlighting the relevance of these aminoacids for substrate interaction. Conclusions: We used phosphoproteomics of Cdc55-deficient cells to uncover new PP2ACdc55 substrates and functions in mitosis. As expected, several hyperphosphorylated proteins corresponded to Cdk1-dependent substrates, although other kinases' consensus motifs were also enriched in our dataset, suggesting that PP2ACdc55 counteracts and regulates other kinases distinct from Cdk1. Indeed, Pkc1 emerged as a novel node of PP2ACdc55 regulation, highlighting a major role of PP2ACdc55 in actin cytoskeleton and cytokinesis, gene ontology terms significantly enriched in the PP2ACdc55-dependent phosphoproteome.
- Published
- 2018