Unmasking of Protein Phosphatase 2 Regulatory Subunit B as a crucial factor in the development and progression of dilated cardiomyopathy
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE238141 | GEO | 2024/09/30
REPOSITORIES: GEO
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Project description:Efficient release of promoter-proximally paused Pol II into productive elongation is essential for gene expression. Recently, we reported that the Integrator complex can bind paused Pol II and drive premature transcription termination, potently attenuating the activity of target genes. Premature termination requires RNA cleavage by the endonuclease subunit of Integrator, but the roles of other Integrator subunits in gene regulation have yet to be elucidated. Here, we report that Integrator subunit 8 (IntS8) is critical for transcription repression through its association with Protein Phosphatase 2A (PP2A). We find that Integrator-bound PP2A dephosphorylates the Pol II C-terminal domain and Spt5, and prevents the transition to productive elongation. Blocking PP2A association with Integrator thus stimulates pause release and gene activation. These results reveal a second catalytic function associated with Integrator-mediated transcription termination, and suggest a new model for the control of productive elongation involving active competition between transcriptional kinases and phosphatases.
2020-10-06 | GSE150844 | GEO
Project description:Expressing the type III effector AWR5 of the phytopathogen Ralstonia solanacearum in yeast cells causes cessation of growth and transcriptional changes reminiscent of inhibition of the TOR pathway. Deletion of cdc55, a regulatory subunit of the PP2A Ser/Thr protein phosphatase eliminates the strong growth defect caused by AWR5 expression. This experiments aims to identify the impact of the cdc55 mutation in the transcriptome of AWR5-expressing cells. induction of AWR5 in JA-100 CDC55 tetO:AWR5-GFP and JA-100 cdc55 tetO:AWR5-GFP
2015-12-31 | E-GEOD-70331 | biostudies-arrayexpress
Project description:Protein phosphatase 2A (PP2A) is an essential Ser/Thr phosphatase that regulates a plethora of cellular processes. PP2A operates as a holoenzyme complex, comprising one each of the scaffolding (A), regulatory (B) and catalytic (C) subunits. PPP2CA is the principal catalytic subunit of the PP2A holoenzyme complex. Although previous studies have reported many substrates of specific PP2A holoenzyme complexes, the full scope of PP2A substrates in cells remains to be defined. To address this, we generated HEK293 cells in which PPP2CA was homozygously knocked in with a dTAG, allowing for efficient and selective degradation of dTAG-PPP2CA with proteolysis-targeting chimeras (PROTACs) targeting the dTAG. By employing an unbiased global phospho-proteomic analysis, we identified 6,280 phospho-peptides corresponding to 2,204 proteins that showed a significant increase in abundance upon dTAG-PPP2CA degradation, implicating them as potential PPP2CA substrates. Among these, some were established PP2A substrates, while most were novel. Bioinformatic analyses revealed the involvement of the identified potential PPP2CA substrates in many cellular processes, including spliceosome function, the cell cycle, RNA transport and ubiquitin-mediated proteolysis. We show that a pSP/pTP motif is a predominant target for PPP2CA. We confirmed some of our phosphoproteomic data with immunoblotting, by utilising commercially available phospho-specific antibodies. We provide an in-depth atlas of potential PPP2CA substrates and establish targeted degradation as a robust tool to unveil phosphatase substrates in cells.
2024-03-18 | PXD045779 | Pride
Project description:Expressing the type III effector AWR5 of the phytopathogen Ralstonia solanacearum in yeast cells causes cessation of growth and transcriptional changes reminiscent of inhibition of the TOR pathway. Deletion of cdc55, a regulatory subunit of the PP2A Ser/Thr protein phosphatase eliminates the strong growth defect caused by AWR5 expression. This experiments aims to identify the impact of the cdc55 mutation in the transcriptome of AWR5-expressing cells.
2015-12-31 | GSE70331 | GEO
Project description:Insulin signaling is mediated via a network of protein phosphorylation. Dysregulation of this network is central to obesity, type 2 diabetes and metabolic syndrome. Here we have investigated the role of phosphatase binding protein Alpha4 (α4) that is essential for the Ser/Thr protein phosphatase PP2A in insulin action/resistance in adipocytes. Unexpectedly, adipocyte-specific inactivation of α4 impairs insulin-induced Akt-mediated Ser/Thr phosphorylation despite a decrease in the PP2A levels. Interestingly, loss of α4 also reduces insulin-induced insulin receptor Tyr phosphorylation. This occurs through decreased association of α4 with Y-box protein 1 (YBX1), resulting in the enhancement of the Tyr phosphatase PTP1B expression. Moreover, adipocyte-specific knockout of α4 results in impaired adipogenesis and altered mitochondrial oxidation leading to increased inflammation, systemic insulin resistance, hepatosteatosis, islet hyperplasia, and impaired thermogenesis. Thus, the α4 /YBX1-mediated pathway of insulin receptor signaling is essential for maintaining insulin sensitivity, normal adipose tissue homeostasis and systemic metabolism.
2022-09-21 | GSE210407 | GEO
Project description:As a serine/threonine phosphatase, protein phosphatase 2A (PP2A) is essential in numerous physiological processes. Our previously study confirmed PP2A dysfunction can cause azoospermia by generating catalytic subunit of PP2A (Ppp2ca) conditional knockout (CKO) in C57BL/6J mice. Here, we further explored the possible mechanisms by focusing on meiosis initiation and spermatogenesis. The deficiency of Ppp2ca in germ cells conspicuously disturbed spermatogonial differentiation and lead to pachytene arrest, accompanied by defects in programmed double-strand break (DSB) repair and meiotic sex chromosome inactivation (MSCI). Furthermore, Ppp2ca-deficient spermatocytes exhibited abnormal agglutination and cohesion complex degradation of chromosome, probably contributing to pachytene arrest. Our study demonstrates the irreplaceable role of PP2A in spermatogenesis and provide more evidences on azoospermia etiology.
2021-12-01 | GSE181132 | GEO
Project description:Sympathetic activation is the main driver of cardiac inotropy and lusitropy. Stimulation of the -adrenoceptor (AR) signaling cascade leads to activation of cAMP-dependent protein kinase (PKA) and subsequent cardiac protein phosphorylation, which is counteracted by the corresponding protein phosphatases 2A (PP2A). Both, kinase and phosphatase are sensitive to oxidation. For PKA, the oxidant-mediated mode of regulation has been described to occur by interdisulfide formation between type I regulatory subunits (PKA-RI). Furthermore, oxidant-mediated intradisulfide formation in the catalytic subunit of PKA and PP2A was shown to inhibit kinase or phosphatase activity, respectively. Nitroxyl donors exert positive inotropic and lusitropic effects in cardiac myocytes.
2020-10-29 | PXD019808 | Pride
Project description:We report the mRNA expression profile of Streptococcus pneumoniae Type 2 D39 strain-derived mutant lacking the phpP gene encoding eukaryote-type ser/thr phosphatase and the stkP gene encoding Ser/Thr protein kinase.
2019-04-13 | GSE113337 | GEO
Project description:Different transcriptome, metabolome, and phosphoproteome studies have already indicated a regulatory role of Ser/Thr phosphorylation in the central metabolism of S. aureus. However, it remains still unknown how this exactly tunes its metabolism. For that reason, Extreme Pathway Analysis was used to understand metabolic flux regulation by Ser/Thr phosphorylation. YANAsquare software and YANAvergence routine were chosen to calculate pathway activities of WT, ΔpknB, Δstp, and double mutant S. aureus NewmanHG strains. Gene expression data of S. aureus NewmanHG were collected taking the wild type and mutations of kinase (pknB), phosphatase (stp) or both (pknB/stp) phenotypes. The resulting flux changes were modelled with YANAvergence taking the different gene expression data and an established genome scale model of S. aureus metabolic modes as basis. The fluxes of the different strains were next calculated, taking the gene expression data into account. A concerted metabolic change of fluxes was observed. In particular the pathways for peptidoglycan, nucleotide, and aromatic amino acid synthesis as well as catabolism involving aspartate transaminase (GOT) changed significantly compared to the wild type. Single mutations of pknB and stp were compared to illustrate their different contribution to the phenotype, e.g. pyrimidine synthesis is dramatically impaired by pknB deletion but much less by loss of stp. In the double knock out strain, there is higher activity of peptidoglycan, purine, and aromatic amino acids synthesis from glucose, but lower activity of pyrimidine synthesis from glucose compared to WT. The results suggest a probable regulatory role of Ser/Thr phosphorylation in amino acid catabolism and the glycolysis/gluconeogenesis switch to provide the cell with sufficient cell wall components, nucleotides, and aromatic amino acids. Yvck is suggested to serve as regulatory protein linking Ser/Thr phosphorylation to the previous results of S. aureus metabolism. Moreover, Yvck seems in particular to be involved in the switch between glycolysis and gluconeogenesis. However, further studies based on experimental approaches are needed to demonstrate that Ser/Thr phosphorylation and Yvck are necessary for a correct cell growth under gluconeogenic conditions. The metabolic model allows not only a comprehensive representation of these changes for all involved pathways in the four strains, it further points out the function of the previously unannotated ycvk operon.. The relevance of these specific metabolic pathway adaptations regarding the pathogenicity of S.aureus and their contributions to the infection process will be analyzed and discussed.
2019-11-11 | GSE122362 | GEO
Project description:The Ser/Thr protein phosphatase 2A (PP2A) is a highly conserved collection of heterotrimeric holoenzymes responsible for the dephosphorylation of many regulated phosphoproteins. Substrate recognition and the integration of regulatory cues are mediated by B regulatory subunits that are complexed to the catalytic subunit (C) by a scaffold protein (A). PP2A/B55 substrate recruitment was thought to be mediated by charge-charge interactions between the surface of B55a and its substrates. Challenging this view, we recently discovered a conserved SLiM [RK]-V-x-x-[VI]-R in a range of proteins, including substrates such as the retinoblastoma-related protein p107 and TAU (Fowle et al. eLife 2021;10:e63181). Here we report the identification of this SLiM in FAM122A, an inhibitor of B55a/PP2A. This conserved SLiM is necessary for FAM122A binding to B55a in vitro and in cells. Computational structure prediction with AlphaFold-Multimer predicts an interaction consistent with the mutational and biochemical data and supports a mechanism whereby FAM122A uses the 'SLiM' in the form of a short a-helix (helix 1) to dock to the B55a top groove, thereby blocking substrate binding to the same site. In this model, FAM122A also spatially constrains substrate access by occluding the catalytic subunit with a second a-helix immediately adjacent to helix 1. Consistently, FAM122A functions as a competitive inhibitor as it prevents the binding of substrates in in vitro competition assays and the dephosphorylation of CDK substrates by B55a/PP2A in cell lysates. The ablation of FAM122A in human cell lines reduces the rate of proliferation, progression through cell cycle transitions, and abrogates G1/S and intra-S phase cell cycle checkpoints. FAM122A-KO in HEK293 cells results in attenuation of CHK1 and CHK2 activation in response to replication stress. Overall, these data strongly suggest that FAM122A is a short helical motif (SHeM)-dependent, substrate-competitive inhibitor of B55a/PP2A that suppresses multiple functions of B55a in the DNA damage response and in timely progression through the cell cycle interphase.
2024-05-30 | MSV000094900 | MassIVE