scholarly journals Dephosphorylation of the Core Septin, AspB, in a Protein Phosphatase 2A-Dependent Manner Impacts Its Localization and Function in the Fungal Pathogen Aspergillus fumigatus

2016 ◽  
Vol 7 ◽  
Author(s):  
José M. Vargas-Muñiz ◽  
Hilary Renshaw ◽  
Amber D. Richards ◽  
Greg Waitt ◽  
Erik J. Soderblom ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Protein Phosphatase ◽  
Fungal Pathogen ◽  
Protein Phosphatase 2A ◽  
Dependent Manner ◽  
The Core ◽  
Phosphatase 2A ◽  
And Function

scholarly journals Protein phosphatase 2A regulates life and death decisions via Akt in a context-dependent manner

2007 ◽  
Vol 104 (48) ◽  
pp. 19011-19016 ◽  
Author(s):  
S. Andrabi ◽  
O. V. Gjoerup ◽  
J. A. Kean ◽  
T. M. Roberts ◽  
B. Schaffhausen
Keyword(s):  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Dependent Manner ◽  
Life And Death ◽  
Phosphatase 2A ◽  
Context Dependent

Propofol Inhibits Phosphorylation of N -methyl-d-aspartate Receptor NR1 Subunits in Neurons

2006 ◽  
Vol 104 (4) ◽  
pp. 763-769 ◽  
Author(s):  
Seth Kingston ◽  
Limin Mao ◽  
Lu Yang ◽  
Anish Arora ◽  
Eugene E. Fibuch ◽  
...  
Keyword(s):  
Glutamate Receptors ◽  
Okadaic Acid ◽  
Protein Phosphatase ◽  
Cortical Neurons ◽  
Protein Phosphatase 2A ◽  
Ionotropic Glutamate Receptors ◽  
Dependent Manner ◽  
Calyculin A ◽  
General Anesthetic ◽  
Phosphatase 2A

Background Anesthetics may interact with ionotropic glutamate receptors to produce some of their biologic actions. Cellular studies reveal that the ionotropic glutamate receptors, N-methyl-D-aspartate receptors (NMDARs), can be phosphorylated on their NR1 subunits at the C-terminal serine residues, which is a major mechanism for the regulation of NMDAR functions. It is currently unknown whether anesthetics have any modulatory effects on NMDAR NR1 subunit phosphorylation. Methods The possible effect of a general anesthetic propofol on phosphorylation of NR1 subunits at serine 897 (pNR1S897) and 896 (pNR1S896) was detected in cultured rat cortical neurons. Results Propofol consistently reduced basal levels of pNR1S897 and pNR1S896 in a concentration-dependent manner. This reduction was rapid as the reliable reduction of pNR1S896 developed 1 min after propofol administration. Pretreatment of cultures with the protein phosphatase 2A inhibitors okadaic acid or calyculin A blocked the effect of propofol on the NR1 phosphorylation, whereas okadaic acid or calyculin A alone did not alter basal pNR1S897 and pNR1S896 levels. In addition, propofol decreased tyrosine phosphorylation of protein phosphatase 2A at tyrosine 307, resulting in an increase in protein phosphatase 2A activity. In the presence of propofol, the NMDAR agonist-induced intracellular Ca2+ increase was impaired in neurons with dephosphorylated NR1 subunits. Conclusions Together, these data indicate an inhibitory effect of a general anesthetic propofol on NMDAR NR1 subunit phosphorylation in neurons. This inhibition was mediated through a signaling mechanism involving activation of protein phosphatase 2A.

Protein Phosphatase 2A (PP2A) Associates with Integrin αIIbβ3 and Regulates Adhesion to Fibrinogen.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1644-1644
Author(s):  
Kadekuzhi V. Vijayan ◽  
Yan Liu ◽  
Paul F. Bray
Keyword(s):  
Protein Phosphatase ◽  
Platelet Adhesion ◽  
Protein Phosphatase 2A ◽  
Critical Role ◽  
Specific Interaction ◽  
Cytoskeletal Proteins ◽  
Dependent Manner ◽  
Integrin Αiibβ3 ◽  
Knock Down ◽  
Phosphatase 2A

Abstract We have previously demonstrated that protein phosphatase 1 (PP1) associates constitutively with the integrin αIIb subunit and regulates myosin light chain phosphorylation. In this study, we considered whether other member of the serine/threonine (Ser/Thr) phosphatase family namely, protein phosphatase 2A (PP2A) associates with the integrin αIIbβ3. Co-immunoprecipitation assays using lysates from resting human platelets revealed the presence of a catalytic subunit of PP2A (PP2Ac) in the αIIb immunoprecipitates, and in a reciprocal experiment, αIIb was detected in the PP2Ac immunoprecipitates. In contrast, another platelet abundant Ser/Thr phosphatase, protein phosphatase 2C (PP2C) was not detected in the αIIb immunoprecipitates. Furthermore, the association of PP2Ac with integrin αIIbβ3 was also observed in 293 cells overexpressing αIIbβ3. These results indicate a constitutive and specific interaction of PP2Ac with the integrin αIIbβ3. Polystyrene beads coated with purified PP2Ac but not BSA supported the binding of purified integrin αIIbβ3 in a dose dependent manner, suggesting that the interaction of PP2Ac with αIIbβ3 was direct. Furthermore, purified PP2Ac as well as PP2Ac in lysates obtained from the resting platelets bound specifically to a biotinylated αIIb cytoplasmic peptide encompassing residues 985–995 but not to a scrambled peptide, suggesting that the integrin αIIb is sufficient to mediate the interaction of PP2Ac in vitro. The association of PP2Ac with the platelet integrin αIIbβ3 was not altered during platelet adhesion to fibrinogen (αIIbβ3 outside-in signaling) or during thrombin or ADP stimulation (inside-out signaling to αIIbβ3). In contrast, we have previously shown that integrin-bound PP1 dissociated from the αIIbβ3 complex upon platelet adhesion and thrombin-induced platelet activation. The association of PP2Ac with the integrin αIIbβ3 correlates well with the dephosphorylation of a PP2A substrate, extracellular-signal regulated kinase 2 (ERK2) during thrombin-induced platelet aggregation that we and others have previously demonstrated. More importantly, ERK2 dephosphorylation was not observed in platelets from Glanzmann thrombasthenic patients or in normal platelets pretreated with RGDS or integrilin, suggesting a critical role for integrin αIIbβ3 in the dephosphorylation of ERK2. To ascertain a physiological relevance for the PP2A-αIIbβ3 association, we used short interfering RNAs (siRNAs) to knock down the expression of PP2Ac in the 293/αIIbβ3 cells. Knock down was maximal (~55–70%) and specific to PP2Ac because PP1 and actin protein levels were not different between the control and PP2Ac siRNA treated cells. Consistent with the reduction in the PP2Ac protein level in the PP2Ac knock down cells there was ~70% reduction in the PP2Ac phosphatase activity, and a concomitant increased basal ERK2 phosphorylation. PP2Ac knock down significantly (P≤0.006) increased the adhesion of 293/αIIbβ3 cells to fibrinogen. The adhesion was αIIbβ3 specific because it could be abolished with an αIIbβ3 function blocking antibody (10E5). These findings supports a mechanism whereby the integrin associated Ser/Thr phosphatases might regulate αIIbβ3 adhesive functions via dephosphorylation of key cytoskeletal proteins.

scholarly journals Expression of I2PP2A, an inhibitor of protein phosphatase 2A, induces c-Jun and AP-1 activity

1999 ◽  
Vol 341 (2) ◽  
pp. 293-298 ◽  
Author(s):  
S W.K. AL-MURRANI ◽  
James R. WOODGETT ◽  
Zahi DAMUNI
Keyword(s):  
Dna Binding ◽  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Dependent Manner ◽  
Activator Protein 1 ◽  
Intact Cells ◽  
Hek 293 ◽  
293 Cells ◽  
Site Selectivity ◽  
Phosphatase 2A

Transient expression of I2PP2A, a potent inhibitor of protein phosphatase 2A (PP2A), in HEK-293 cells increased the concentration and DNA binding of the proto-oncogene c-Jun. In contrast, expression of the catalytic subunit of PP2A (PP2AC) markedly decreased the concentration and DNA binding of c-Jun. Expression of I2PP2A also increased the transcriptional activity of activator protein-1, and this effect was diminished in a dose-dependent manner by expression of PP2AC. Densitometric analysis following Western blotting of extracts with antibodies specific for phospho-Ser63 and Ser73 suggests that the effects of I2PP2A and PP2AC expression might be mediated, in part, by changes in the phosphorylation of c-Jun at Ser63. The results indicate that I2PP2A elicits effects that are consistent with it acting as an inhibitor of PP2A in intact cells, and suggest that PP2A might exhibit site selectivity with respect to c-Jun phosphorylation.

scholarly journals Specification of the Okadaic Acid Equivalent for Okadaic Acid, Dinophysistoxin-1, and Dinophysistoxin-2 Based on Protein Phosphatase 2A Inhibition and Cytotoxicity Assays Using Neuro 2A Cell Line

2021 ◽  
Vol 9 (10) ◽  
pp. 1140
Author(s):  
Tsuyoshi Ikehara ◽  
Kazuya Chikanishi ◽  
Naomasa Oshiro
Keyword(s):  
Okadaic Acid ◽  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Shellfish Poisoning ◽  
Diarrhetic Shellfish Poisoning ◽  
Oral Toxicity ◽  
Cytotoxicity Assays ◽  
The Core ◽  
Phosphatase 2A ◽  
Core Activity

Diarrhetic shellfish poisoning (DSP) is a globally occurring disease threatening public health and trade. The causative toxins, okadaic acid (OA), dinophysistoxin-1 (DTX1), and dinophysistoxin-2 (DTX2) are collectively called OAs, and are quantified using the LC-MS/MS method. The hazardous effect of total OAs is expressed as the sum of OA equivalents defined for respective OAs based on mouse lethality, produced by either intraperitoneal (OAip) or oral administration (OAor). OAs are potent inhibitors of protein phosphatase 2A (PP2A) and are cytotoxic, necessitating expansion of the concept of OA equivalents to all relevant bioactivities. In this study, we determined OA equivalents for respective OA members in PP2A inhibition and cytotoxicity assays. To secure result credibility, we used certified OAs, reference materials, and PP2A produced using genetic engineering. The relative ratio of the OA equivalents determined by PP2A inhibition assays for OA, DTX1, and DTX2 were 1.0:1.6:0.3, while the ratio determined using the cytotoxicity assays indicated 1.0:1.5:0.5. OA equivalents showed a similar tendency in the PP2A inhibition and cytotoxicity assays, and matched better with oral toxicity data than intraperitoneal toxicity in mice. The PP2A inhibition assay, which measures the core activity of the OAs, suggested a higher OA equivalent for DTX1 than that currently used.

scholarly journals Regulation of Cancerous inhibitor of PP2A (CIP2A) by small molecule inhibitor for c-Jun NH2-Terminal Kinases (JNKs), SP600125, in Human Fibrosarcoma (HT1080) cells

F1000Research ◽  
2013 ◽  
Vol 2 ◽  
pp. 174
Author(s):  
Anchit Khanna
Keyword(s):  
Protein Expression ◽  
Small Molecule ◽  
Protein Phosphatase ◽  
Affinity Purification ◽  
Protein Phosphatase 2A ◽  
Small Molecule Inhibitor ◽  
Dependent Manner ◽  
Molecule Inhibitor ◽  
Ht1080 Cells ◽  
Phosphatase 2A

Background: Protein phosphatase 2A inhibition is one of the pre-requisites for human cell transformation. Previously, we have identified an endogenous inhibitor of PP2A, CIP2A (Cancerous Inhibitor of Protein Phosphatase 2A) in human fibrosarcoma cells (HT1080) using tandem affinity purification. CIP2A over expression has been demonstrated in almost every tumour type studied so far. However, our understanding on the mechanisms regulating CIP2A expression in human cancers, especially in sarcomas, is still emerging. Methods:  Human fibrosarcoma (HT1080) cells were treated with small molecule inhibitors against the three major signalling pathways, namely p38, MEK and JNK pathways to identify the pathway regulating CIP2A expression in the sarcoma cells. This was followed by verification of the results using small interfering RNAs (siRNA) for the kinases. Results:  In line with previous observations, small molecule inhibitor for MEK pathway (PD98059) decreased CIP2A mRNA and protein expression. Interestingly, small molecule inhibitor for the JNK pathway, SP600125 decreased mRNA and protein levels of CIP2A oncoprotein with negligible effect of SB203580 (p38 kinase) inhibitor on CIP2A expression in HT1080 cells. However, siRNAs specific to either JNK1 or JNK2 kinases did not result in decrease in CIP2A expression. Contrarily, two different CIP2A siRNAs, which were used as positive controls, decreased JNK2 expression in HT1080 cells. Conclusion: Although it is well established that SP600125 inhibits JNK kinases, it has also been shown to inhibit a spectra of other kinases. SP600125 inhibits CIP2A protein expression both in time and concentration dependent manner. However, depletion of both JNK1 and JNK2 kinases using specific siRNAs fails to decrease CIP2A protein expression levels, thereby indicating the need to verify the results obtained by treatment with small molecular inhibitors of kinases by independent approaches like two different target specific siRNAs. Finally, fortuitously we identify JNK2 as a CIP2A downstream target in HT1080 cells.

scholarly journals Differential methylation and altered conformation of cytoplasmic and nuclear forms of protein phosphatase 2A during cell cycle progression.

1995 ◽  
Vol 129 (2) ◽  
pp. 397-410 ◽  
Author(s):  
P Turowski ◽  
A Fernandez ◽  
B Favre ◽  
N J Lamb ◽  
B A Hemmings
Keyword(s):  
Cell Cycle ◽  
Conformational Changes ◽  
Protein Phosphatase ◽  
Cell Cycle Progression ◽  
Protein Phosphatase 2A ◽  
Differential Methylation ◽  
Control Mechanisms ◽  
The Core ◽  
Phosphatase 2A ◽  
G1 Cells

Protein phosphatase 2A (PP2A) appears to be involved in the regulation of many cellular processes. Control mechanisms that lead to the activation (and deactivation) of the various holoenzymes to initiate appropriate dephosphorylation events remain obscure. The core components of all PP2A holoenzymes are the catalytic (PP2Ac) and 63-65-kD regulatory (PR65) subunits. Monospecific and affinity-purified antibodies against both PP2Ac and PR65 show that these proteins are ubiquitously localized in the cytoplasm and the nucleus in nontransformed fibroblasts. As determined by quantitative immunofluorescence the core subunits of PP2A are twofold more concentrated in the nucleus than in the cytoplasm. Detailed analysis of synchronized cells reveals striking changes in the nuclear to cytoplasmic ratio of PP2Ac-specific immunoreactivity albeit the total amounts of neither PP2Ac nor PR65 in each compartment alters significantly during the cell cycle. Our results imply that differential methylation of PP2Ac occurs at the G0/G1 and G1/S boundaries. Specifically a demethylated form of PP2Ac is found in the cytoplasm of G1 cells, and in the nucleus of S and G2 cells. In addition nuclear PP2A holoenzymes appear to undergo conformational changes at the G0/G1 and G1/S boundaries. During mitosis PP2A is lost from the nuclear compartment, and unlike protein phosphatase 1 shows no specific association with the condensed chromatin.

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