Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Whole-cell recordings were made from dorsomedial nucleus tractus solitarii neurons in thin coronal medullary slices of the rat, at the level of the area postrema. Monosynaptic excitatory postsynaptic currents (EPSCs) were evoked in the tractus solitarius by electrical stimulation in the presence of D-2-amino-5-phosphonopentanoic acid (AP5) and bicuculline. Currents were also evoked by pressure ejection of (S)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) in the presence of AP5, bicuculline, and tetrodotoxin or muscimol in the presence of 6,7-dinitroquinoxaline-2,3-dione and AP5. The metabotropic glutamate receptor (mGluR) agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylate [(1S,3R)-ACPD] reversibly depressed the EPSC and muscimol currents and reversibly potentiated AMPA currents. The effects of (1S,3R)-ACPD were blocked in the presence of a low concentration of the phosphoprotein phosphatase (PP)1 and PP2A inhibitor okadaic acid (OA) but not by a low concentration of the PP inhibitor calyculin A. The immunosuppressant agent FK506 failed to block (1S,3R)-ACPD effects on AMPA currents. However, (1S,3R)-ACPD applied in the presence of FK506 produced a reversible potentiation of muscimol currents. We previously demonstrated that the cell-permeant cGMP analog 8-Br-cGMP can mimic many of the effects of (1S,3R)-ACPD. OA antagonized the effects of 8-Br-cGMP in the present investigation. Finally, we previously demonstrated that brief tetanic stimulation results in the activation of a presynaptic mGluR autoreceptor and depression of subsequently evoked EPSCs. OA similarly blocked tetanus-induced depression of EPSCs. These findings suggest that mGluRs on tractus solitarius afferents and first-order nucleus tractus solitarii neurons may modulate glutamate release and AMPA and gamma-aminobutyric acid type A receptor activity via activation of one or more PPs, such as PP2A and/or calcineurin.
Mol Pharmacol 1994 Jun
PMID:Inhibition of phosphoprotein phosphatases blocks metabotropic glutamate receptor effects in the rat nucleus tractus solitarii. 751 97

Monoclonal antibodies are frequently used to identify protein isoforms. The present study documents certain artifacts in such applications and suggests methods for their testing. Two alpha subunit specific anti-calcineurin antibodies, VJ6 and VD3, reacted strongly with a brain isozyme, BPI, but not with the phosphatases from liver and spleen. Controlled proteolysis of BPI indicated that epitopes of both antibodies are localized to aminoterminal region, thus is suggesting that the lack of antibody reactivity could result from proteolytic artifacts. The occurrence of proteolysis during sample preparation can be monitored by including small quantity of BPI in the tissue extraction buffer. Rapid isolation procedure has been used to obtain liver and spleen isozymes reacting with VD3 but not VJ6 antibody.
Biochem Mol Biol Int 1994 Mar
PMID:Immunological approach for the identification of isozymes of CAM-stimulated phosphatase. 751 20

Calcineurin is a conserved Ca2+/calmodulin-dependent protein phosphatase that plays a critical role in Ca(2+)-mediated signaling in many cells. Yeast cells lacking functional calcineurin (cna1 cna2 or cnb1 mutants) display growth defects under specific environmental conditions, for example, in the presence of high concentrations of Na+, Li+, Mn2+, or OH- but are indistinguishable from wild-type cells under standard culture conditions. To characterize regulatory pathways that may overlap with calcineurin, we performed a synthetic lethal screen to identify mutants that require calcineurin on standard growth media. The characterization of one such mutant, cnd1-8, is presented. The CND1 gene was cloned, and sequence analysis predicts that it encodes a novel protein 1,876 amino acids in length with multiple membrane-spanning domains. CND1 is identical to the gene identified previously as FKS1, ETG1, and CWH53, cnd1 mutants are sensitive to FK506 and cyclosporin A and exhibit slow growth that is improved by the addition of osmotic stabilizing agents. This osmotic agent-remedial growth defect and microscopic evidence of spontaneous cell lysis in cnd1 cultures suggest that cell integrity is compromised in these mutants. Mutations in the genes for yeast protein kinase C (pkc1) and a MAP kinase (mpk1/slt2) disrupt a Ca(2+)-dependent signaling pathway required to maintain a normal cell wall and cell integrity. We show that pkc1 and mpk1/slt2 growth defects are more severe in the absence of calcineurin function and less severe in the presence of a constitutively active form of calcineurin. These observations suggest that calcineurin and protein kinase C perform independent but physiologically related functions in yeast cells. We show that several mutants that lack a functional vacuolar H(+)-ATPase (vma) require calcineurin for vegetative growth. We discuss possible roles for calcineurin in regulating intracellular ion homeostasis and in maintaining cell integrity.
Mol Cell Biol 1995 Aug
PMID:Calcineurin, the Ca2+/calmodulin-dependent protein phosphatase, is essential in yeast mutants with cell integrity defects and in mutants that lack a functional vacuolar H(+)-ATPase. 754 41

The immunosuppressive drugs FK506 and cyclosporin A block T-lymphocyte proliferation by inhibiting calcineurin, a critical signaling molecule for activation. Multiple intracellular receptors (immunophilins) for these drugs that specifically bind either FK506 and rapamycin (FK506-binding proteins [FKBPs]) or cyclosporin A (cyclophilins) have been identified. We report the cloning and characterization of a new 51-kDa member of the FKBP family from murine T cells. The novel immunophilin, FKBP51, is distinct from the previously isolated and sequenced 52-kDa murine FKBP, demonstrating 53% identity overall. Importantly, Western blot (immunoblot) analysis showed that unlike all other FKBPs characterized to date, FKBP51 expression was largely restricted to T cells. Drug binding to recombinant FKBP51 was demonstrated by inhibition of peptidyl prolyl isomerase activity. As judged from peptidyl prolyl isomerase activity, FKBP51 had a slightly higher affinity for rapamycin than for FK520, an FK506 analog. FKBP51, when complexed with FK520, was capable of inhibiting calcineurin phosphatase activity in an in vitro assay system. Inhibition of calcineurin phosphatase activity has been implicated both in the mechanism of immunosuppression and in the observed toxic side effects of FK506 in nonlymphoid cells. Identification of a new FKBP that can mediate calcineurin inhibition and is restricted in its expression to T cells suggests that new immunosuppressive drugs may be identified that, by virtue of their specific interaction with FKBP51, would be targeted in their site of action.
Mol Cell Biol 1995 Aug
PMID:FKBP51, a novel T-cell-specific immunophilin capable of calcineurin inhibition. 754 43

Dynamic regulation of ion transport is essential for homeostasis as cells confront changes in their environment. The gene HAL3 encodes a novel component of this regulatory circuit in the yeast Saccharomyces cerevisiae. Overexpression of HAL3 improves growth of wild-type cells exposed to toxic concentrations of sodium and lithium and suppresses the salt sensitivity conferred by mutation of the calcium-dependent protein phosphatase calcineurin. Null mutants of HAL3 display salt sensitivity. The sequence of HAL3 gives little clue to its function. However, alterations in intracellular cation concentrations associated with changes in HAL3 expression suggest that HAL3 activity may directly increase cytoplasmic K+ and decrease Na+ and Li+. Cation efflux in S. cerevisiae is mediated by the P-type ATPase encoded by the ENA1/PMR24 gene, a putative plasma membrane Na+ pump whose expression is salt induced. Acting in concert with calcineurin, HAL3 is necessary for full activation of ENA1 expression. This functional complementarity is also reflected in the participation of both proteins in recovery from alpha-factor-induced growth arrest. Recently, HAL3 was isolated as a gene (named SIS2) which when overexpressed partially relieves loss of transcription of G1 cyclins in mutants lacking the protein phosphatase Sit4p. Therefore, HAL3 influences cell cycle control and ion homeostasis, acting in parallel to the protein phosphatases Sit4p and calcineurin.
Mol Cell Biol 1995 Oct
PMID:Regulation of cation transport in Saccharomyces cerevisiae by the salt tolerance gene HAL3. 756 98

1,3-beta-D-Glucan is a major structural polymer of yeast and fungal cell walls and is synthesized from UDP-glucose by the multisubunit enzyme 1,3-beta-D-glucan synthase. Previous work has shown that the FKS1 gene encodes a 215-kDa integral membrane protein (Fks1p) which mediates sensitivity to the echinocandin class of antifungal glucan synthase inhibitors and is a subunit of this enzyme. We have cloned and sequenced FKS2, a homolog of FKS1 encoding a 217-kDa integral membrane protein (Fks2p) which is 88% identical to Fks1p. The residual glucan synthase activity present in strains with deletions of fks1 is (i) immunodepleted by antibodies prepared against FKS2 peptides, demonstrating that Fks2p is also a component of the enzyme, and (ii) more sensitive to the echinocandin L-733,560, explaining the increased sensitivity of fks1 null mutants to this drug. Simultaneous disruption of FKS1 and FKS2 is lethal, suggesting that Fks1p and Fks2p are alternative subunits with essential overlapping function. Analysis of FKS1 and FKS2 expression reveals that transcription of FKS1 is regulated in the cell cycle and predominates during growth on glucose, while FKS2 is expressed in the absence of glucose. FKS2 is essential for sporulation, a process which occurs during nutritional starvation. FKS2 is induced by the addition of Ca2+ to the growth medium, and this induction is completely dependent on the Ca2+/calmodulin-dependent phosphoprotein phosphatase calcineurin. We have previously shown that growth of fks1 null mutants is highly sensitive to the calcineurin inhibitors FK506 and cyclosporin A. Expression of FKS2 from the heterologous ADH1 promoter results in FK506-resistant growth. Thus, the sensitivity of fks1 mutants to these drugs can be explained by the calcineurin-dependent transcription of FKS2. Moreover, FKS2 is also highly induced in response to pheromone in a calcineurin-dependent manner, suggesting that FKS2 may also play a role in the remodeling of the cell wall during the mating process.
Mol Cell Biol 1995 Oct
PMID:Differential expression and function of two homologous subunits of yeast 1,3-beta-D-glucan synthase. 756 18

Stimulation of the T-cell antigen receptor (TCR) induces activation of multiple tyrosine kinases, resulting in phosphorylation of numerous intracellular substrates. One substrate is p95vav, which is expressed exclusively in hematopoietic and trophoblast cells. It contains a number of structural motifs, including Src homology 2, Src homology 3, and pleckstrin homology domains and a putative guanine nucleotide exchange domain. The role of p95vav in TCR-mediated signaling processes is unclear. Here, we show that overexpression of p95vav alone in Jurkat T cells leads to activation of the nuclear factors, including NFAT, involved in interleukin-2 expression. Furthermore, p95vav synergizes with TCR stimulation in inducing NFAT- and interleukin-2-dependent transcription. In contrast, NFAT activation by a G-protein-coupled receptor is not modulated by p95vav overexpression, suggesting that the effect is specific to the TCR signaling pathways. Although removal of the first 67 amino acids of p95vav activates its transforming potential in NIH 3T3 cells, this region appears to be required for its function in T cells. We further demonstrate that the p95vav-induced NFAT activation is not mimicked by Ras activation, though its function is dependent upon Ras and Raf. Furthermore, the activating function of p95vav is blocked by FK506, suggesting that its activity also depends on calcineurin. To further dissect p95vav involvement in TCR signaling, we analyzed various Jurkat mutants deficient in TCR signaling function or TCR expression and showed that an intact TCR signaling pathway is required for p95vav to function. However, overexpression of p95vav does not appear to influence TCR-induced protein tyrosine phosphorylation or increases in cytoplasmic free calcium. Taken together, our data suggest that p95vav plays an important role at an yet unidentified proximal position in the TCR signaling cascade.
Mol Cell Biol 1995 Aug
PMID:A functional T-cell receptor signaling pathway is required for p95vav activity. 762 28

The immunosuppressants cyclosporin A (CsA) and FK506 appear to block T-cell function by inhibiting the calcium-regulated phosphatase calcineurin. While multiple distinct intracellular receptors for these drugs (cyclophilins and FKBPs, collectively immunophilins) have been characterized, the functionally active ones have not been discerned. We found that overexpression of cyclophilin A or B or FKBP12 increased T-cell sensitivity to CsA or FK506, respectively, demonstrating that they are able to mediate the inhibitory effects of their respective immunosuppressants in vivo. In contrast, cyclophilin C, FKBP13, and FKBP25 had no effect. Direct comparison of the Ki of each drug-immunophilin complex for calcineurin in vitro revealed that although calcineurin binding was clearly necessary, it was not sufficient to explain the in vivo activity of the immunophilin. Subcellular localization was shown also to play a role, since gene deletions of cyclophilins B and C which changed their intracellular locations altered their activities significantly. Cyclophilin B has been shown previously to be located within calcium-containing intracellular vesicles; its ability to mediate CsA inhibition implies that certain components of the signal transduction machinery are also spatially restricted within the cell.
Mol Cell Biol 1993 Aug
PMID:Identification of the immunophilins capable of mediating inhibition of signal transduction by cyclosporin A and FK506: roles of calcineurin binding and cellular location. 768 44

To gain a better understanding of the relationship between the female rat reproductive system and preproenkephalin (PPE) expressing neurons under physiological conditions, we examined changes in PPE mRNA levels in the mediobasal hypothalamus during the rat estrous cycle by means of northern blotting and in situ hybridization histochemistry (ISHH). In the Northern blot studies, we found that PPE mRNA levels in the mediobasal hypothalamus were significantly increased by noon of proestrus compared to those in the morning and stayed high until diestrus day 1, and returned toward low levels on diestrous day 2. In contrast, measured as controls, glyceraldehyde-3-phosphate-dehydrogenase mRNA levels were significantly higher on proestrus regardless of time of day compared to diestrus day 2, and levels of calcineurin mRNA on proestrous and estrous were significantly lower than diestrous day 1 and day 2. ISHH studies revealed that these changes in PPE mRNA levels were specific in the ventromedial hypothalamic nucleus pars ventrolateralis (VMHVL), since we could not see any significant changes in signal in other parts including ventromedial hypothalamic nucleus pars dorsomedialis and arcuate hypothalamic nucleus. In the VMHVL, PPE mRNA levels in the afternoon of proestrous were significantly higher than those in the afternoon of diestrous day 2 whereas no significant change in PPE mRNA was observed in the caudate-putamen. The present study provides additional information relevant to possible implications of PPE gene expression in female reproductive systems, since changes in PPE mRNA levels may be associated with estrogen as well as progesterone or other hormonal concentrations during the estrous cycle.(ABSTRACT TRUNCATED AT 250 WORDS)
Brain Res Mol Brain Res 1995 Jan
PMID:Changes in preproenkephalin messenger RNA level in the rat ventromedial hypothalamus during the estrous cycle. 770 66

Tau is a neuron-specific, microtubule-associated protein that forms paired helical filaments (PHFs) of Alzheimer's disease when aberrantly phosphorylated. We have attempted to elucidate the protein kinases and phosphatases that regulate tau phosphorylation. Incubation of rat, human, and rhesus monkey temporal neocortex slices with the phosphatase inhibitor okadaic acid induced epitopes of tau similar to those found in PHFs. Okadaic acid (1-20 microM) induced variant forms of tau at 60-68 kDa, which were recognized by the monoclonal antibodies Alz-50 (in humans only) and 5E2 and two polyclonal antipeptide antisera, OK-1 and OK-2. The phosphorylation-sensitive monoclonal antibody Tau-1 failed to recognize the slowest mobility forms of tau after okadaic acid treatment. FK-520 (1-10 microM), a potent inhibitor of calcineurin activity, was tested in brain slices and found not to alter tau mobility. However, combinations of FK-520 (5 microM) and okadaic acid (100 nM) caused tau mobility shifts similar to those seen after 10 microM okadaic acid treatment; similar results were seen using the calcineurin-selective inhibitor cypermethrin. Treatment of human slices with 10 microM okadaic acid decreased both protein phosphatase 2A and calcineurin activity; FK-520 inhibited only protein phosphatase 2B activity. A proposed tau-directed kinase, 42-kDa mitogen-activated protein kinase (p42mapk), was activated by okadaic acid (> 100 nM) but not FK-520 (5 microM). Nerve growth factor (100 ng/ml) activated p42mapk, particularly when used in combination with 100 nM okadaic acid; changes in tau mobility were seen when this kinase was activated. Forskolin (2 microM) antagonized the effects of nerve growth factor on both p42mapk activity and tau phosphorylation; forskolin alone had little effect on PHF-like tau formation induced by phosphatase inhibitors. These results outline complex interactions between tau-directed protein kinases and protein phosphatases and suggest potential sites for therapeutic intervention.
Mol Pharmacol 1995 Apr
PMID:Tau phosphorylation in brain slices: pharmacological evidence for convergent effects of protein phosphatases on tau and mitogen-activated protein kinase. 772 35


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