Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P67775 (alpha isoform)
 797 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activity and allosteric properties of plant phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) are controlled posttranslationally by specific reversible phosphorylation of a strictly conserved serine residue near the N-terminus. This up/down-regulation of PEPC is catalyzed by a dedicated and highly regulated serine/threonine (Ser/Thr) kinase (PEPC-kinase) and an opposing type-2A Ser/Thr phosphatase (PP2A). In marked contrast to PEPC-kinase, the PP2A holoenzyme from photosynthetic tissue has been virtually unstudied to date. In the present investigation, we have partially purified and characterized the native form of this PP2A from illuminated leaves of maize (Zea mays L.), a C4 plant, using maize [32P]PEPC as substrate. Various conventional chromatographic matrices, together with thiophosphorylated C4 PEPC-peptide and microcystin-LR affinity-supports, were exploited for the enrichment of this PP2A from soluble leaf extracts. Biochemical and immunological results indicate that the C4-leaf holoenzyme is analogous to other eukaryotic PP2As in being a approximately 170-kDa heteromer comprised of a core PP2Ac-A heterodimer (approximately 38- and approximately 65-kDa subunits, respectively) complexed with a putative, approximately 74-kDa B-type regulatory/targeting subunit. This heterotrimer lacks any strict substrate specificity in that it dephosphorylates C4 PEPC, mammalian phosphorylase a, and casein in vitro. This activity is independent of free Me2+, insensitive to levamisole and the Inhibitor-2 protein that targets PP1, activated by several polycations such as protamine and poly-L-lysine, and highly sensitive to inhibition by microcystin-LR and okadaic acid (IC50 approximately 30 pM), all of which are diagnostic features of yeast and mammalian PP2As. In addition, this C4-leaf PP2A holoenzyme (i) is inhibited in vitro by physiological concentrations of certain C4 PEPC-related metabolites (L-malate, PEP, glucose 6-phosphate, but not the activator glycine) when either 32P-labeled maize PEPC or rabbit muscle phosphorylase a is used as substrate, suggesting a direct effect on this Ser/Thr phosphatase; and (ii) displays, at best, only modest light/dark effects in vivo on its apparent molecular mass, component core subunits and activity against C4 PEPC, in marked contrast to the opposing activity of PEPC-kinase in C4 and Crassulacean acid metabolism leaves. This report represents one of the few studies of a heteromeric PP2A holoenzyme from photosynthetic tissue that dephosphorylates a known target enzyme in plants, such as PEPC, sucrose-phosphate synthase or nitrate reductase.
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PMID:Partial purification and biochemical characterization of a heteromeric protein phosphatase 2A holoenzyme from maize (Zea mays L.) leaves that dephosphorylates C4 phosophoenolpyruvate carboxylase. 1150 60

Neuronal Ca(2+)/calmodulin-dependent protein kinase II (CaM kinase II) plays important roles in the control of nerve functions in response to intracellular Ca(2+) (for reviews [Annu. Rev. Physiol. 57 (1995) 417-445; Trends Neurosci. 17 (1994) 406-412]). Brief Ca(2+) signals activate CaM kinase II, and stimulate an autophosphorylation of Thr-286 which allows the kinase to maintain its activated state even after the Ca(2+) concentration has returned to basal levels [J. Biol. Chem. 264 (1989) 16759-16763; Neuron 3 (1989) 59-70; J. Biochem. 109 (1991) 137-143]. Autophosphorylation of CaM kinase II occurs in situ, but it occurs relatively quickly, within just a few minutes [Endocrinology 134 (1994) 2245-2250; J. Biol. Chem. 268 (1993) 7863-7867; J. Biol. Chem. 265 (1990) 18055-18058]. In the present study, we investigated the involvement of the autophosphorylated/Ca(2+)-independent form of CaM kinase II in neurite outgrowth. When neuroblastoma Neruo2a (Nb2a) cells expressing the alpha isoform of CaM kinase II (Nb2a/alpha cells) were stimulated by plating, they formed neurites. The autophosphorylation of Thr-286 and appearance of Ca(2+)-independent activity preceded the neurite formation. The effect of mutating of the kinase autophosphorylation site replacing Thr-286 with Ala (alpha T286A kinase) or Asp (alpha T286D kinase) was examined. alpha T286A kinase was not converted to a Ca(2+)-independent form, and alpha T286D kinase had Ca(2+)-independent activity significantly as an autophosphorylated kinase. Cells expressing alpha T286D kinase had much longer neurites than Nb2a/alpha cells, whereas cells with alpha T286A kinase did not form neurites. These results indicated that the Ca(2+)-independent form of CaM kinase II autophosphorylated at Thr-286 is involved in neurite outgrowth.
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PMID:Investigation of the Ca(2+)-independent form of Ca(2+)/calmodulin-dependent protein kinase II in neurite outgrowth. 1173 91

Serine/threonine kinase Akt is a downstream effector protein of phosphatidylinositol-3-kinase (PI-3K). Many integrins can function as positive modulators of the PI-3K/Akt pathway. Integrin alpha 2 beta 1 is a collagen receptor that has been shown to induce specific signals distinct from those activated by other integrins. Here, we found that, in contrast what was found for cells adherent to fibronectin, alpha 2 beta 1-mediated cell adhesion to collagen leads to dephosphorylation of Akt and glycogen synthase kinase 3 beta (GSK3 beta) and concomitantly to the induction of protein serine/threonine phosphatase 2A (PP2A) activity. PP2A activation can be inhibited by mutation in the alpha 2 cytoplasmic domain and by a function-blocking anti-alpha 2 antibody. Akt can be coprecipitated with PP2A, and coexpression of Akt with PP2Ac (catalytic subunit) inhibits Akt kinase activity. Integrin alpha 2 beta 1-related activation of PP2A is dependent on Cdc42. These results indicate that cell adhesion to collagen modulates Akt activity via the alpha 2 beta 1-induced activation of PP2A.
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PMID:Integrin alpha 2 beta 1 promotes activation of protein phosphatase 2A and dephosphorylation of Akt and glycogen synthase kinase 3 beta. 1183 2

Mutations in KRIT1, a protein initially identified based on a yeast two-hybrid interaction with the RAS-family GTPase RAP1A, are responsible for the development of the inherited vascular disorder cerebral cavernous malformations (CCM1). As the function of the KRIT1 protein and its role in CCM pathogenesis remain unknown, we performed yeast two-hybrid screens to identify additional protein binding partners. A fragment containing the N-terminal 272 amino acid residues of KRIT1, a region lacking similarity to any known protein upon database searches, was used as bait. From parallel screens of human fetal brain and HeLa cDNA libraries, we obtained multiple independent isolates of human integrin cytoplasmic domain-associated protein-1 (ICAP-1) as interacting clones. The interaction of KRIT1 and ICAP-1 was confirmed by GST-KRIT1 trapping of endogenous ICAP-1 from 293T cells. The alpha isoform of ICAP-1 is a 200 amino acid serine/threonine-rich phosphoprotein which binds the cytoplasmic tail of beta1 integrins. We show that mutagenesis of the N-terminal KRIT1 NPXY amino acid sequence, a motif critical for ICAP-1 binding to beta1 integrin molecules, completely abrogates the KRIT1/ICAP-1 interaction. The interaction between ICAP-1 and KRIT1, and the presence of a FERM domain in the latter, suggest that KRIT1 might be involved in the bidirectional signaling between integrin molecules and the cytoskeleton. Furthermore, these data suggest that KRIT1 might affect cell adhesion processes via integrin signaling in CCM1 pathogenesis.
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PMID:KRIT1 association with the integrin-binding protein ICAP-1: a new direction in the elucidation of cerebral cavernous malformations (CCM1) pathogenesis. 1185 71

We provide evidence of a cross-talk between nuclear receptor and Ser/Thr protein phosphatases and show that vitamin D receptor (VDR) interacts with the catalytic subunit of protein phosphatases, PP1c and PP2Ac, and induces their enzymatic activity in a ligand-dependent manner. PP1c specifically interacts with VDR but not retinoic acid receptor alpha and retinoid X receptor alpha in yeast. Although VDR-PP1c and VDR-PP2Ac interaction is ligand-independent in vivo, 1alpha,25-dihydroxy-vitamin D(3) induces VDR-associated phosphatase activity. Further, VDR modulation of PP1c/PP2Ac activity results in a rapid and specific dephosphorylation and inactivation of their substrate, p70 S6 kinase (p70(S6k)). Finally, we demonstrate that the endogenous VDR, PP1c or PP2Ac, and p70(S6k) are present in a ternary complex in vivo, and the interaction of p70(S6k) with the VDR-PP complex is modulated by the phosphorylation state of the kinase. Since p70(S6k) is essential for G(1)-S transition, our results provide a molecular basis of 1alpha,25-dihydroxyvitamin D(3)-induced G(1) block in colon cancer cells.
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PMID:A vitamin D receptor-Ser/Thr phosphatase-p70 S6 kinase complex and modulation of its enzymatic activities by the ligand. 1203 52

Casein kinase Ialpha (CKIalpha) belongs to a family of serine/threonine protein kinases involved in membrane trafficking, RNA processing, mitotic spindle formation and cell cycle progression. In this report, we identified several CKIalpha interacting proteins including RCC1, high mobility group proteins 1 and 2 (HMG1, HMG2), Erf, centaurin-alpha1, synaptotagmin IX and CPI-17 that were isolated from brain as CKIalpha co-purifying proteins. Actin, importin-alpha(1), importin-beta, PP2Ac, centaurin-alpha1, and HMG1 were identified by affinity chromatography using a peptide column comprising residues 214-233 of CKIalpha. We have previously shown that centaurin-alpha1 represents a CKIalpha partner both in vitro and in vivo. The nuclear protein regulator of chromosome condensation 1 (RCC1) is a guanosine nucleotide exchange factor for Ran which is involved in nuclear transport and mitotic spindle formation. Here we show that CKIalpha and RCC1 interact in brain and in cultured cells. However, the interaction does not involve residues 217-233 of CKIalpha which are proposed from X-ray structures to represent an anchoring site for CKI partners. Formation of the RCC1/CKIalpha complex is consistent with the association of the kinase with mitotic spindles. In conclusion, we have identified a number of novel CKIalpha protein partners and their relations to CKI are discussed.
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PMID:Identification of casein kinase Ialpha interacting protein partners. 1206 30

T lymphocyte activation signals regulate the expression and transactivation function of retinoid X receptor (RXR) alpha through an interplay of complex signaling cascades that are not yet fully understood. We show that cellular Ser/Thr protein phosphatases (PPs) play an important role in mediating these processes. Inhibitors specific for PP1 and PP2A decreased basal expression of RXR alpha RNA and protein in T lymphocyte leukemia Jurkat cells and prevented activation-induced RXR alpha accumulation in these cells. In addition, these inhibitors attenuated the RXR responsive element (RXRE)-dependent transcriptional activation in transient transfection assays. Inhibitors of calcineurin (CN), by contrast, did not have any effect on the basal RXR alpha expression and even augmented activation-induced RXR alpha expression. Expression of a dominant-active (DA) mutant of CN together with a DA mutant of protein kinase C (PKC)theta;, a novel PKC isoform, significantly increased RXRE-dependent transcription. Expression of catalytically inactive PKC theta; or a dominant-negative mutant of PKC theta; failed to synergize with CN and did not increase RXRE-dependent transcription. Expression of a DA mutant of PKC alpha or treatment with PMA was found to attenuate PKC theta; and CN synergism. We conclude that PP1, PP2A, and CN regulate levels and transcriptional activation function of RXR alpha in T cells. In addition, CN synergizes with PKC theta; to induce RXRE-dependent activation, a cooperative function that is antagonized by the activation of the conventional PKC alpha isoform. Thus, PKC theta; and PKC alpha may function as positive and negative modulators, respectively, of CN-regulated RXRE-dependent transcription during T cell activation.
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PMID:Regulation of retinoid X receptor responsive element-dependent transcription in T lymphocytes by Ser/Thr phosphatases: functional divergence of protein kinase C (PKC)theta; and PKC alpha in mediating calcineurin-induced transactivation. 1209 75

The cellular localization of the 35 kDa, low molecular mass acid metallophosphatase (LMW AcPase) from the frog (Rana esculenta) liver and its activity towards P-Ser and P-Tyr phosphorylated peptides were studied. This enzyme was localized to the cytoplasm of hepatocytes but did not appear in other cells of liver tissue (endothelium, macrophages, blood cells). This LMW AcPase does not display activity towards (32)P-phosphorylase a under conditions standard for the enzymes of PPP family. Proteins containing P-Ser: rabbit (32)P-phosphorylasea and phosvitin are hydrolysed only at acidic pH and are poor substrates for this enzyme. The frog AcPase is not inhibited by okadaic acid and F(-) ions, the Ser/Thr protein phosphatase inhibitors. Moreover, the frog enzyme does not cross-react with specific antisera directed against N-terminal fragment of human PP2A and C-terminal conserved fragment of the eukaryotic PP2A catalytic subunits. These results exclude LMW AcPase from belonging to Ser/Thr protein phosphatases: PP1c or PP2Ac. In addition to P-Tyr, this enzyme hydrolyses efficiently at acidic pH P-Tyr phosphorylated peptides (hirudin and gastrin fragments). K(m) value for the hirudin fragment (7.55 +/- 1.59 x 10(-6) M) is 2-3 orders of magnitude lower in comparison with other substrates tested. The enzyme is inhibited competitively by typical inhibitors of protein tyrosine phosphatases (PTPases): sodium orthovanadate, molybdate and tungstate. These results may suggest that the LMW AcPase of frog liver can act as PTPase in vivo. A different cellular localization and different response to inhibition by tetrahedral oxyanions (molybdate, vanadate and tungstate) provide further evidence that LMW AcPase of frog liver is distinct from the mammalian tartrate-resistant acid phosphatases.
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PMID:The 35 kDa acid metallophosphatase of the frog Rana esculenta liver: studies on its cellular localization and protein phosphatase activity. 1283 81

The beta1 subunit of integrin is serine/threonine phosphorylated in growth arrested human breast cancer MCF-7 cells, while it is not in quiescent normal human breast epithelial (HBE) cells. Using the affinity-purified antibodies PB788-9 against the synthetic oligopeptide that contained phosphothreonines corresponding to threonines 788 and 789 on beta1 integrin, beta1 integrin in MCF-7 cells, but not in HBE cells, was found to react with PB788-9. The beta1 integrin immunoprecipitates from HBE cells co-immunoprecipitated the core enzyme of serine/threonine protein phosphatase (PP) 2A, consisting of the regulatory A (PP2A-A) and the catalytic C (PP2A-C) subunits, with the protein phosphatase activity susceptible to okadaic acid (OA), an inhibitor of PP2A and PP1, but not to a PP1 inhibitor. In contrast, beta1 integrin from MCF-7 cells co-immunoprecipitated PP2A-C, but not PP2A-A, with no protein phosphatase activity. Immunoblotting of whole cell lysates revealed that a comparable amount of PP2A-C was present in either HBE or MCF-7 cells, but the amount of PP2A-A was significantly reduced in MCF-7 cells compared to that in HBE cells. The results suggest that the failure of beta1 integrin dephosphorylation at threonines 788 and 789 may be due to a significant reduction in the PP2A-A expression in MCF-7 cells.
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PMID:Reduced expression of the regulatory A subunit of serine/threonine protein phosphatase 2A in human breast cancer MCF-7 cells. 1453 64

The biochemistry of the mitochondrial production of nitric oxide is reviewed to gain insight into the basic role of this radical in mitochondrial and cellular oxidative metabolism. The mitochondrial production of nitric oxide is catalyzed by a nitric-oxide synthase (mtNOS). This enzyme has the same cofactor and substrate requirements as other constitutive nitric-oxide synthases. Its occurrence was demonstrated in various mitochondrial preparations from different organs and species using diverse approaches (oxidation of oxymyoglobin, electron paramagnetic resonance in conjunction with spin trap, radiolabeled L-arginine, immunohistochemistry, nitric-oxide electrode). MtNOS has been identified as the alpha isoform of nNOS, acylated at a Thr or Ser residue, and phosphorylated at the C-terminal end. Endogenous nitric oxide reversibly inhibits oxygen consumption and ATP synthesis by competitive inhibition of cytochrome oxidase. Nitric oxide is the first molecule that fulfills the requirement for a cytochrome oxidase activity modulator: it is a competitive inhibitor, produced endogenously at a fair rate near the target site, at concentrations high enough to exhibit an inhibitory effect on cytochrome oxidase. The role of the mitochondrial nitric oxide production is discussed in terms of the physiological (modulating oxygen gradients into tissues) and pathological (abrogation of oxygen gradient modification, apoptosis, protein nitrative/oxidative stress) implications.
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PMID:Mitochondrial nitric-oxide synthase: role in pathophysiology. 1471 Oct 5


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