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Determinants of site-specific dephosphorylation by PP2A-B56

ABSTRACT: The tumor suppressor PP2A is a major cellular protein phosphatase that regulates numerous cellular processes through the formation of holoenzymes containing distinct regulatory B-subunits. However, the determinants of differential substrate dephosphorylation by PP2A are poorly understood. Here, we develop a specific genetically encoded inhibitor of PP2A-B56 and perform global phosphoproteomic studies to identify hundreds of regulated phosphorylation sites. We show that PP2A-B56 substrate specificity is controlled by affinity and position of B56 binding motifs and that PP2A active site preference is determined by the B-subunit. These insights uncover PP2A-B56 as a novel negative regulator of ADAM17 mediated growth factor signalling and tumor development. Collectively our work identifies basic principles of PP2A-B56 specificity with broad implications for understanding signalling in eukaryotes.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Fusion, Q Exactive Plus

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Arminja Kettenbach

PROVIDER: MSV000084245 | MassIVE | Wed Aug 28 14:13:00 BST 2019

SECONDARY ACCESSION(S): PXD015205

REPOSITORIES: MassIVE

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A dynamic charge:charge interaction modulates PP2A:B56 interactions

Project description:The recruitment of specific substrates by ser/thr protein phosphatase 2A (PP2A) regulatory B-subunits is poorly understood, limiting our understanding of PP2A-regulated signaling. Recently, the first PP2A:B56 consensus binding motif, LxxIxE, which confers substrate specificity to PP2A:B56 holoenzymes, was identified. However, most validated LxxIxE motifs interact with PP2A:B56 with low micromolar affinities, suggesting that additional recognition motifs exist that function cooperatively to enhance PP2A:B56 binding. Here, we report the requirement of a positively charged motif in a subset of PP2A:B56 interactors that facilitates B56 binding via dynamic charge:charge interactions. Using molecular and cellular approaches, we show that a highly conserved, negatively charged groove on B56 mediates this dynamic interaction. We also show that this motif is required for efficient binding and dephosphorylation of KIF4A in cells, confirming the functional relevance of this motif for PP2A:B56 dephosphorylation. Collectively our results provide an important framework for understanding PP2A regulation of cellular signaling.

2019-05-16 | MSV000083785 | MassIVE

Structural mechanism for inhibition of PP2A-B56 and oncogenicity by CIP2A

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Project description:Meiosis creates genetic diversity by recombination and segregation of chromosomes. The synaptonemal complex assembles during meiotic prophase I and assists faithful exchanges between homologous chromosomes, but how its assembly/disassembly is regulated remains to be understood. Here we report how two major post-translational modifications, phosphorylation and ubiquitination, co-operate to promote synaptonemal complex assembly. We found that the ubiquitin ligase complex SCF is important for assembly and maintenance of the synaptonemal complex in Drosophila female meiosis. This function of SCF is mediated by two substrate-recognising F-box proteins, Slmb/βTrcp and Fbxo42. SCF-Fbxo42 downregulates the phosphatase subunit PP2A-B56, which is important for synaptonemal complex assembly and maintenance.

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