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Frontiers in Plant Science 2022Serine/threonine protein phosphatase 2C (PP2C) dephosphorylates proteins and plays crucial roles in plant growth, development, and stress response. In this study, we...
Serine/threonine protein phosphatase 2C (PP2C) dephosphorylates proteins and plays crucial roles in plant growth, development, and stress response. In this study, we characterized a clade B member of maize PP2C family, i.e., ZmPP2C26, that negatively regulated drought tolerance by dephosphorylating ZmMAPK3 and ZmMAPK7 in maize. The gene generated and isoforms through untypical alternative splicing. ZmPP2C26S lost 71 amino acids including an MAPK interaction motif and showed higher phosphatase activity than ZmPP2C26L. ZmPP2C26L directly interacted with, dephosphorylated ZmMAPK3 and ZmMAPK7, and localized in chloroplast and nucleus, but ZmPP2C26S only dephosphorylated ZmMAPK3 and localized in cytosol and nucleus. The expression of and was significantly inhibited by drought stress. Meanwhile, the maize mutant exhibited enhancement of drought tolerance with higher root length, root weight, chlorophyll content, and photosynthetic rate compared with wild type. However, overexpression of and significantly decreased drought tolerance in and rice with lower root length, chlorophyll content, and photosynthetic rate. Phosphoproteomic analysis revealed that the ZmPP2C26 protein also altered phosphorylation level of proteins involved in photosynthesis. This study provides insights into understanding the mechanism of PP2C in response to abiotic stress.
PubMed: 35463404
DOI: 10.3389/fpls.2022.851531 -
Molecular Cell Oct 2022Neuronal activity induces topoisomerase IIβ (Top2B) to generate DNA double-strand breaks (DSBs) within the promoters of neuronal early response genes (ERGs) and...
Neuronal activity induces topoisomerase IIβ (Top2B) to generate DNA double-strand breaks (DSBs) within the promoters of neuronal early response genes (ERGs) and facilitate their transcription, and yet, the mechanisms that control Top2B-mediated DSB formation are unknown. Here, we report that stimulus-dependent calcium influx through NMDA receptors activates the phosphatase calcineurin to dephosphorylate Top2B at residues S1509 and S1511, which stimulates its DNA cleavage activity and induces it to form DSBs. Exposing mice to a fear conditioning paradigm also triggers Top2B dephosphorylation at S1509 and S1511 in the hippocampus, indicating that calcineurin also regulates Top2B-mediated DSB formation following physiological neuronal activity. Furthermore, calcineurin-Top2B interactions following neuronal activity and sites that incur activity-induced DSBs are preferentially localized at the nuclear periphery in neurons. Together, these results reveal how radial gene positioning and the compartmentalization of activity-dependent signaling govern the position and timing of activity-induced DSBs and regulate gene expression patterns in neurons.
Topics: Animals; Mice; Calcineurin; Calcium; DNA; DNA Breaks, Double-Stranded; DNA Topoisomerases, Type II; DNA-Binding Proteins; Neurons; Receptors, N-Methyl-D-Aspartate
PubMed: 36206766
DOI: 10.1016/j.molcel.2022.09.012 -
Frontiers in Molecular Biosciences 2020Several variant proteins are produced from , including two representative proteins produced via alternative splicing machinery. One protein is the canonical translation...
Several variant proteins are produced from , including two representative proteins produced via alternative splicing machinery. One protein is the canonical translation eukaryotic elongation factor eEF1Bδ1, and the other is the heat shock-responsive transcription factor eEF1BδL. eEF1Bδ1 is phosphorylated by cyclin-dependent kinase 1 (CDK1), but the machinery controlling eEF1BδL phosphorylation and dephosphorylation has not been clarified. In this study, we found that both proteins were dephosphorylated under heat shock and proteotoxic stress, and this dephosphorylation was inhibited by okadaic acid. Using proteins with mutations at putative phosphorylated residues, we revealed that eEF1Bδ1 and eEF1BδL are phosphorylated at S133 and S499, respectively, and these residues are both CDK1 phosphorylation sites. The eEF1BδL S499A mutant more strongly activated promoter-driven reporter than the wild-type protein and S499D mutant. Furthermore, protein phosphatase 1 (PP1) was co-immunoprecipitated with eEF1Bδ1 and eEF1BδL, and PP1 dephosphorylated both proteins . Thus, this study clarified the role of phosphorylation/dephosphorylation in the functional regulation of eEF1BδL during heat shock.
PubMed: 33521052
DOI: 10.3389/fmolb.2020.598578 -
Cell Discovery Jul 2023How cells adapt their gene expression to nutritional changes remains poorly understood. Histone H3T11 is phosphorylated by pyruvate kinase to repress gene transcription....
How cells adapt their gene expression to nutritional changes remains poorly understood. Histone H3T11 is phosphorylated by pyruvate kinase to repress gene transcription. Here, we identify the protein phosphatase 1 (PP1), Glc7 as the enzyme that specifically dephosphorylates H3T11. We also characterize two novel Glc7-containing complexes and reveal their roles in regulating gene expression upon glucose starvation. Specifically, the Glc7-Sen1 complex dephosphorylates H3T11 to activate the transcription of autophagy-related genes. The Glc7-Rif1-Rap1 complex dephosphorylates H3T11 to derepress the transcription of telomere-proximal genes. Upon glucose starvation, Glc7 expression is up-regulated and more Glc7 translocates into the nucleus to dephosphorylate H3T11, leading to induction of autophagy and derepressed transcription of telomere-proximal genes. Furthermore, the functions of PP1/Glc7 and the two Glc7-containing complexes are conserved in mammals to regulate autophagy and telomere structure. Collectively, our results reveal a novel mechanism that regulate gene expression and chromatin structure in response to glucose availability.
PubMed: 37433812
DOI: 10.1038/s41421-023-00551-1 -
International Journal of Molecular... Dec 2019Protein phosphorylation affects conformational change, interaction, catalytic activity, and subcellular localization of proteins. Because the post-modification of... (Review)
Review
Protein phosphorylation affects conformational change, interaction, catalytic activity, and subcellular localization of proteins. Because the post-modification of proteins regulates diverse cellular signaling pathways, the precise control of phosphorylation states is essential for maintaining cellular homeostasis. Kinases function as phosphorylating enzymes, and phosphatases dephosphorylate their target substrates, typically in a much shorter time. The c-Jun N-terminal kinase (JNK) signaling pathway, a mitogen-activated protein kinase pathway, is regulated by a cascade of kinases and in turn regulates other physiological processes, such as cell differentiation, apoptosis, neuronal functions, and embryonic development. However, the activation of the JNK pathway is also implicated in human pathologies such as cancer, neurodegenerative diseases, and inflammatory diseases. Therefore, the proper balance between activation and inactivation of the JNK pathway needs to be tightly regulated. Dual specificity phosphatases (DUSPs) regulate the magnitude and duration of signal transduction of the JNK pathway by dephosphorylating their substrates. In this review, we will discuss the dynamics of phosphorylation/dephosphorylation, the mechanism of JNK pathway regulation by DUSPs, and the new possibilities of targeting DUSPs in JNK-related diseases elucidated in recent studies.
Topics: Animals; Dual-Specificity Phosphatases; Humans; JNK Mitogen-Activated Protein Kinases; Mitogen-Activated Protein Kinases; Models, Biological; Phosphorylation; Signal Transduction
PubMed: 31817617
DOI: 10.3390/ijms20246157 -
The Journal of Biological Chemistry Jun 1995Rhodopsin is an important member of the superfamily of G protein-coupled receptors. In vitro studies have suggested that multiphosphorylation of rhodopsin is a pivotal...
Rhodopsin is an important member of the superfamily of G protein-coupled receptors. In vitro studies have suggested that multiphosphorylation of rhodopsin is a pivotal step in phototransduction. Because the in vitro biochemical experiments were conducted under nonphysiological conditions, we investigated the phosphorylation of mouse rhodopsin in vivo and determined the sites of phosphorylation and the time course of dephosphorylation. We found that a single phosphate group is incorporated into the rhodopsin molecule in a light-dependent manner, primarily at Ser338 after flashes and at Ser334 after continuous illumination. Dephosphorylation of these sites had different kinetics and spatial distribution in rod outer segments. Dephosphorylation of Ser338 was complete within 30 min, while Ser334 was dephosphorylated much slower (requiring up to 60 min), correlating with the regeneration of rhodopsin. These results suggest that phosphorylation of Ser338 and Ser334 plays different roles in phototransduction.
Topics: Amino Acid Sequence; Animals; Dark Adaptation; Light; Mice; Molecular Sequence Data; Phosphorylation; Rhodopsin; Rod Cell Outer Segment; Signal Transduction
PubMed: 7782279
DOI: 10.1074/jbc.270.24.14259 -
Biochimica Et Biophysica Acta.... Jun 2021PGAM5 is a protein phosphatase located in the inner mitochondrial membrane through its transmembrane (TM) domain and is cleaved within the TM domain upon mitochondrial...
PGAM5 is a protein phosphatase located in the inner mitochondrial membrane through its transmembrane (TM) domain and is cleaved within the TM domain upon mitochondrial dysfunction. We found previously that cleaved PGAM5 is released from mitochondria, following proteasome-mediated rupture of the outer mitochondrial membrane during mitophagy, a selective form of autophagy specific to mitochondria. Here, we examined the role of cleaved PGAM5 outside mitochondria. Deletion mutants that mimic cleaved PGAM5 existed not only in the cytosol but also in the nucleus, and a fraction of cleaved PGAM5 translocated to the nucleus during mitophagy induced by the uncoupler CCCP. We identified serine/arginine-related nuclear matrix protein of 160 kDa (SRm160)/SRRM1, which contains a highly phosphorylated domain rich in arginine/serine dipeptides, called the RS domain, as a nuclear protein that interacts with PGAM5. PGAM5 dephosphorylated SRm160, and incubation of lysates from WT cells, but not of those from PGAM5-deficient cells, induced dephosphorylation of SRm160 and another RS domain-containing protein SRSF1, one of the most characterized serine/arginine-rich (SR) proteins. Moreover, phosphorylation of these proteins and other SR proteins, which are commonly reactive toward the 1H4 monoclonal antibody that detects phosphorylated SR proteins, decreased during mitophagy, largely because of PGAM5 activity. These results suggest that PGAM5 regulates phosphorylation of these nuclear proteins during mitophagy. Because SRm160 and SR proteins play critical roles in mRNA metabolism, PGAM5 may coordinate cellular responses to mitochondrial stress at least in part through post-transcriptional and pre-translational events.
Topics: Antigens, Nuclear; Cell Nucleus; Cytosol; HeLa Cells; Humans; Membrane Proteins; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Membranes; Mitochondrial Proteins; Mitophagy; Nuclear Matrix-Associated Proteins; Phosphoprotein Phosphatases; Phosphorylation; RNA-Binding Proteins; Serine-Arginine Splicing Factors; Ubiquitin-Protein Ligases
PubMed: 33872670
DOI: 10.1016/j.bbamcr.2021.119045 -
Science Signaling Feb 2021, an important human parasite, has a flagellum that controls cell motility, morphogenesis, proliferation, and cell-cell communication. Inheritance of the newly assembled...
, an important human parasite, has a flagellum that controls cell motility, morphogenesis, proliferation, and cell-cell communication. Inheritance of the newly assembled flagellum during the cell cycle requires the Polo-like kinase homolog TbPLK and the kinetoplastid-specific protein phosphatase KPP1, although whether TbPLK acts on KPP1 or vice versa has been unclear. Here, we showed that dephosphorylation of TbPLK on Thr by KPP1 maintained low TbPLK activity in the flagellum-associated hook complex structure, thereby ensuring proper flagellum positioning and attachment. This dephosphorylation event required the recognition of phosphorylated Thr in the activation loop of TbPLK by the N-terminal Plus3 domain of KPP1 and the dephosphorylation of phosphorylated Thr in TbPLK by the C-terminal catalytic domain of KPP1. Dephosphorylation of TbPLK by KPP1 prevented hyperphosphorylation of the hook complex protein TbCentrin2, thereby allowing timely dephosphorylation of phosphorylated TbCentrin2 for hook complex duplication and flagellum positioning and attachment. Thus, KPP1 attenuates TbPLK activity by dephosphorylating TbPLK to facilitate flagellum inheritance.
Topics: Cell Cycle; Flagella; Phosphoprotein Phosphatases; Protozoan Proteins; Trypanosoma brucei brucei
PubMed: 33563698
DOI: 10.1126/scisignal.abc6435 -
Cell Chemical Biology Jun 2022PDE3A-SLFN12 complex formation activates the SLFN12 RNase, but the biochemical details of RNase activation remain mysterious. In this issue of Cell Chemical Biology, Yan...
PDE3A-SLFN12 complex formation activates the SLFN12 RNase, but the biochemical details of RNase activation remain mysterious. In this issue of Cell Chemical Biology, Yan and colleagues report that two phosphoserines on SLFN12 are dephosphorylated in response to PDE3A binding, and this dephosphorylation is required for activation of the SLFN12 RNase.
Topics: Cell Line, Tumor; Phosphorylation; Ribonucleases
PubMed: 35714590
DOI: 10.1016/j.chembiol.2022.05.010 -
The Biochemical Journal Dec 1988Calmodulin-dependent protein phosphatase has been proposed to be an important phosphotyrosyl-protein phosphatase. The ability of the enzyme to attack autophosphorylated...
Calmodulin-dependent protein phosphatase has been proposed to be an important phosphotyrosyl-protein phosphatase. The ability of the enzyme to attack autophosphorylated insulin receptor was examined and compared with the known ability of the enzyme to act on autophosphorylated epidermal-growth-factor (EGF) receptor. Purified calmodulin-dependent protein phosphatase was shown to catalyse the complete dephosphorylation of phosphotyrosyl-(insulin receptor). When compared at similar concentrations, 32P-labelled EGF receptor was dephosphorylated at greater than 3 times the rate of 32P-labelled insulin receptor; both dephosphorylations exhibited similar dependence on metal ions and calmodulin. Native phosphotyrosyl-protein phosphatases in cell extracts were also characterized. With rat liver, heart or brain, most (75%) of the native phosphatase activity against both 32P-labelled insulin and EGF receptors was recovered in the particulate fraction of the cell, with only 25% in the soluble fraction. This subcellular distribution contrasts with results of previous studies using artificial substrates, which found most of the phosphotyrosyl-protein phosphatase activity in the soluble fraction of the cell. Properties of particulate and soluble phosphatase activity against 32P-labelled insulin and EGF receptors are reported. The contribution of calmodulin-dependent protein phosphatase activity to phosphotyrosyl-protein phosphatase activity in cell fractions was determined by utilizing the unique metal-ion dependence of calmodulin-dependent protein phosphatase. Whereas Ni2+ (1 mM) markedly activated the calmodulin-dependent protein phosphatase, it was found to inhibit potently both particulate and soluble phosphotyrosyl-protein phosphatase activity. In fractions from rat liver, brain and heart, total phosphotyrosyl-protein phosphatase activity against both 32P-labelled receptors was inhibited by 99.5 +/- 6% (mean +/- S.E.M., 30 observations) by Ni2+. Results of Ni2+ inhibition studies were confirmed by other methods. It is concluded that in cell extracts phosphotyrosyl-protein phosphatases other than calmodulin-dependent protein phosphatase are the major phosphotyrosyl-(insulin receptor) and -(EGF receptor) phosphatases.
Topics: Animals; Calcineurin; Calmodulin-Binding Proteins; ErbB Receptors; Liver; Phosphoprotein Phosphatases; Phosphorylation; Protein Tyrosine Phosphatases; Rats; Receptor, Insulin
PubMed: 2852008
DOI: 10.1042/bj2560893