A protein phosphatase 1 specific phosphatase targeting peptide (PhosTAP) to identify the PP1 phosphatome

• 成果类型: Article
• 署名作者: Choy, Meng S.; Nguyen, Hieu T.; Kumar, Ganesan S.; Peti, Wolfgang; Kettenbach, Arminja N.; Page, Rebecca
• 署名单位: Dartmouth College; Department of Biotechnology (DBT) India; National Institute of Immunology (NII)
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-12054
• DOI: 10.1073/pnas.2415383121
• 发表日期: 2024-10-29
• 关键词: histone h3; structural basis; thr-3 phosphorylation; kinase haspin; aurora b; substrate; binding; dephosphorylation; inhibitor-2; calcineurin

摘要

Phosphoprotein phosphatases (PPPs) are the key serine/threonine phosphatases that regulate all essential signaling cascades. In particular, Protein Phosphatase 1 (PP1) dephosphorylates similar to 80% of all ser/thr phosphorylation sites. Here, we developed a phosphatase targeting peptide (PhosTAP) that binds all PP1 isoforms and does so with a stronger affinity than any other known PP1 regulator. This PhosTAP can be used as a PP1 recruitment tool for Phosphorylation Targeting Chimera (PhosTAC)-type recruitment in in vitro and cellular experiments, as well as in phosphoproteomics experiments to identify PP1- specific substrates and phosphosites. The latter is especially important to further our understanding of cellular signaling, as the identification of substrates and especially phosphosites that are targeted by specific phosphatases lags behind that of their kinase counterparts. Using PhosTAP-based proteomics, we show that, counter to our current understanding, many PP1 regulators are also substrates, that the number of residues between regulator PP1- binding and phosphosites vary significantly, and that PP1 counteracts the activities of mitotic kinases. Finally, we also found that Haspin kinase is a direct substrate of PP1 and that its PP1- dependent dephosphorylation modulates its activity during anaphase. Together, we show that PP1- specific PhosTAPs are a powerful tool for +studying PP1 activity in vitro and in cells.