Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Wheat root tips express a 73 kDa cognate isoform and a 77 kDa heat-shock-induced isoform of peptidyl prolyl cis-trans isomerase (FK506 binding protein; FKBP) that is part of a chaperone complex with hsp90. The 73 kDa and 77 kDa FKBPs have very similar sequences, differing primarily in the N- and C-terminal 20 amino acids. In order to define the potential functional roles of these proteins, the 73 kDa and 77 kDa FKBPs were localized in root tips using antigen-affinity purified antibodies as a probe. The cognate 73 kDa FKBP is localized in the cytoplasm and appears enriched around the periphery of the early vacuole and vesicles exiting the trans-Golgi. Parallel assays with antibodies directed against tonoplast aquaporin and pyrophosphatase confirmed the association of FKBP with an early vacuole compartment. Sucrose gradient centrifugation analysis of root tip lysates also showed that 73 kDa FKBP is co-fractionated with tonoplast aquaporin and V-ATPase in a light compartment near the top of the gradient. Heat-shock treatment of root tips induces the accumulation of 77 kDa FKBP while the abundance of 73 kDa FKBP remains constant. Quantitative EM immunogold assays of the intracellular distribution of FKBP over an 8 h heat-shock time-course showed that FKBP is initially present in the cytoplasm, but is transported into the nucleus where it accumulates in the nucleoplasm and into specific subnuclear domains. The results of this study show that the intracellular distribution of the high Mr FKBPs in wheat root tips differs at normal and elevated temperatures, indicating different functional roles for the FKBP isoforms.
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PMID:Differential distribution of the cognate and heat-stress-induced isoforms of high Mr cis-trans prolyl peptidyl isomerase (FKBP) in the cytoplasm and nucleoplasm. 1458 27

Clinically important resistance of fungal pathogens to azole antifungal drugs is most frequently caused by the over-expression of energy-dependent drug efflux pumps. These pumps usually belong to either the ATP-binding cassette (ABC) family or the major facilitator superfamily (MFS) class of membrane transporter. Little is known about how these pumps work and there is urgent need to develop pump antagonists that circumvent resistance. The expression system is based on an S. cerevisiae AD1-8u- strain deleted in seven major ABC transporters which has reduced background and endogenous efflux activity. Plasmid pABC3 was engineered to allow functional hyper-expression of foreign proteins in this host. The main advantages of our system are its cloning efficiency: the use of homologous recombination to stably integrate single copy constructs into the host genome under the control of a highly active transcriptional regulator. The expression system has been used to clone and express genes encoding drug efflux pumps from several pathogenic fungi. Furthermore, functional over-expression of human P-glycoprotein was also demonstrated. The protein hyper-expression system will be useful for the screening of pump inhibitors and the study of membrane protein pumping mechanisms. This system has been used to screen chemicals for pump inhibitors. It was found that FK506 and milbemycins chemosensitized pump-expressing and fluconazole-resistant strains and inhibited pump ATPase activity.
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PMID:[An efficient system for functional hyper-expression of multidrug efflux pumps in Saccharomyces cerevisiae]. 1511 61

Recently, we established an in vitro model of apoptosis induced by exposure of neuroblastoma SH-SY5Y cells to thapsigargin, an endoplasmic reticular calcium-ATPase inhibitor, and demonstrated that FK506 (tacrolimus) protected against apoptosis. The purpose of this paper was to investigate a possible correlation between the protective effect of FK506 against apoptosis and the regulation of the serum inducible kinase (SNK) and fibroblast growth factor inducible kinase (FNK) genes-which are polo-like kinases expressed abundantly in the brain by FK506. Thapsigargin increased the mRNA level of SNK and FNK in SH-SY5Y cells. FK506 inhibited the increase in SNK mRNA but not FNK mRNA. Deletion analysis of the SNK promoter showed that the promoter site, which was regulated by thapsigargin and FK506 in a calcineurin-dependent manner, is a cAMP response element (CRE)/activating transcription factor (ATF)-like element located 84 base pairs (bp) proximal to the transcriptional initiation site. Although transcription of the SNK gene was also regulated by tunicamycin, etoposide, or staurosporine, FK506 did not show any effects on these regulations. We recently reported that FK506 did not protect against apoptosis induced by these agents. These results indicate that the induction of SNK mRNA by thapsigargin in SH-SY5Y cells is regulated by FK506 via an inhibition of calcineurin at the transcriptional stage, and the transcriptional regulation of the SNK gene by FK506 was well correlated with the protective effect of the compound against apoptosis. Thus, transcriptional regulation of the SNK gene may be a biological marker for analysis of apoptosis of SH-SY5Y cells.
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PMID:Protective effect of FK506 against apoptosis of SH-SY5Y cells correlates with regulation of the serum inducible kinase gene. 1585 11

In MCF-7 (estrogen receptor (ER)+) and in MDA-MB-231 (ER-) cells stably transfected with either estrogen receptor alpha (ERalpha) or beta (ERbeta) subtype (MDA-MB-231 stably transfected with the mouse ERalpha cDNA (MERA) and MDA-MB-231 stably transfected with the human ERbeta cDNA (HERB), respectively) N-term heat shock protein of 90kDa (hsp90) ligands (geldanamycin and radicicol) and C-term hsp90 ligands (novobiocin) decrease the basal and estradiol (E(2))-induced transcription activity of ER on an estrogen responsive element (ERE)-LUC reporter construct concomitantly with or 1h after E(2) treatment. All hsp90 ligands induced an E(2)- and MG132-inhibited decrease of both ER cell content. However, the kinetics of these degradations are slower than those induced by the selective estrogen receptor down-regulator RU 58668 (RU). This suggests that inhibition of the hsp90 ATPase activity targets both ERs to the 26S proteasome and that hsp90 interacts with both ER subtypes. Rapamycin (Rapa) and cyclosporin A (CsA), ligands of immunophilins FK506 binding protein (FKBP52) and cyclophilin of 40kDa (CYP40) interacting in separate ER-hsp90 complexes, both induced a proteasomal-mediated degradation of ERs but not of their cognate immunophilin. Moreover, they also decrease the E(2)-induced luciferase transcription but weaker than RU and hsp90 ligands. Fluorescence activated cell sorter (FACS) analysis revealed a blockade of cell progression by RU and 4-hydroxy-tamoxifen at the G(1) phase of the cell cycle and an induction of apoptosis in MCF-7 cells. Rapa and mainly CsA (but not FK506) and hsp90 ligands promote by their own apoptosis in MCF-7, in MERA, and in HERB cells and in MDA-MB-231 ER-null cells. These data suggest that (1) hsp90, as for all steroid receptors, acts as a molecular chaperone for ERbeta; (2) ER-ligands (except tamoxifen), hsp90- and immunophilin-ligands (except FK506) target the two ER subtypes to a proteasome-mediated proteolysis via different signalling pathways; (3) hsp90- and immunophilin-ligands Rapa and CsA, alone or in association with anti-estrogens such as RU, may constitute a potential therapeutic strategy for breast cancer treatment.
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PMID:Estrogen receptor alpha and beta subtype expression and transactivation capacity are differentially affected by receptor-, hsp90- and immunophilin-ligands in human breast cancer cells. 1586 52

Deficiency of delta-sarcoglycan (delta-SG), a component of the dystrophin-glycoprotein complex (DGC), causes skeletal muscular dystrophy and cardiomyopathy in BIO14.6 hamsters. Here, we studied the involvement of abnormal Ca2+ homeostasis in muscle degeneration and the protective effect of drugs against Ca2+ handling proteins in vivo as well as in vitro. First, we characterized the properties of cultured myotubes from muscles of normal and BIO14.6 hamsters (30-60 days old). While there were no apparent differences in the levels of expression of various Ca2+ handling proteins (L-type Ca2+ channel, ryanodine receptor, SR-Ca2+ ATPase, and Na+/Ca2+ exchanger), muscle-specific proteins (contractile actin and acetylcholine receptor), or DGC member proteins except SGs, BIO14.6 myotubes showed a high degree of susceptibility to mechanical stressors, such as cyclic stretching and hypo-osmotic stress as compared to normal myotubes, as evidenced by marked increases in creatine phosphokinase (CK) release and bleb formation. BIO14.6 myotubes showed abnormal Ca2+ homeostasis characterized by elevated cytosolic Ca2+ concentration, frequent Ca2+ oscillation, and increased 45Ca2+ uptake. These abnormal Ca2+ events and CK release were significantly prevented by Ca2+ handling drugs, tranilast, diltiazem, and FK506. The calpain inhibitor E64 prevented CK release, but not 45Ca2+ uptake. Some of these drugs (tranilast, diltiazem, and FK506) also exerted a significant protective effect for muscle degeneration in BIO14.6 hamsters and mdx mice in vivo. These observations suggest that elevated Ca2+ entry through sarcolemmal Ca2+ channels predominantly contributes to muscle degeneration and that the drugs tested here may have novel therapeutic potential against muscular dystrophy.
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PMID:Protective effects of Ca2+ handling drugs against abnormal Ca2+ homeostasis and cell damage in myopathic skeletal muscle cells. 1600 51

Recently, the academic interest in the yeast Torulaspora delbrueckii has increased notably due to its high resistance to several types of stress, including salt and osmotic imbalance. However, the molecular mechanisms underlying these unusual properties are poorly understood. In Saccharomyces cerevisiae, the high-salt response is mediated by calcineurin, a conserved Ca(2+)/calmodulin-modulated protein phosphatase that regulates the transcriptional factor Crz1p. Here, we cloned the T. delbrueckii TdCRZ1 gene, which encodes a putative zinc finger transcription factor homologue to Crz1p. Consistent with this, overexpression of TdCRZ1 enhanced the salt tolerance of S. cerevisiae wild-type cells and suppressed the sensitivity phenotype of cnb1Delta and crz1Delta mutants to monovalent and divalent cations. However, T. delbrueckii cells lacking TdCrz1p showed phenotypes distinct from those previously observed in S. cerevisiae crz1Delta mutants. Quite remarkably, Tdcrz1-null cells were insensitive to high Na(+) and were more Li(+) tolerant than wild-type cells. Clearly, TdCrz1p was not required for the salt-induced transcriptional activation of the TdENA1 gene, encoding a putative P-type ATPase homologue to the main S. cerevisiae Na(+) pump ENA1. Furthermore, T. delbrueckii cells were insensitive to the immunosuppressive agents FK506 and cyclosporine A, both in the presence and in the absence of NaCl. Signaling through the calcineurin/Crz1 pathway appeared to be essential only on high-Ca(2+)/Mn(2+) media. Hence, T. delbrueckii and S. cerevisiae differ in the regulatory circuits and mechanisms that drive the adaptive response to salt stress.
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PMID:Regulation of salt tolerance by Torulaspora delbrueckii calcineurin target Crz1p. 1652 2

Glial cell line-derived neurotrophic factor (GDNF) is an important neurotrophic factor that has therapeutic implications for neurodegenerative disorders. We previously showed that leucine-isoleucine (Leu-Ile), an analog of a dipeptide-like structure of FK506 (tacrolimus), induces GDNF expression both in vivo and in vitro. In this investigation, we sought to clarify the cellular mechanisms underlying the GDNF-inducing effect of this dipeptide. Leu-Ile transport was investigated using fluorescein isothiocyanate-Leu-Ile in cultured neurons, and the results showed the transmembrane mobility of this dipeptide. By liquid chromatography-mass spectrometry and quartz crystal microbalance assay, we identified heat shock cognate protein 70 as a protein binding specifically to Leu-Ile, and molecular modeling showed that the ATPase domain is the predicted binding site. Leu-Ile stimulated Akt phosphorylation, which was attenuated significantly by heat shock protein 90 (Hsp90) inhibitor geldanamycin (GA). Moreover, enhanced interaction between phosphorylated Akt and Hsp90 was detected by immunoprecipitation. Leu-Ile elicited an increase in cAMP response element binding protein (CREB) phosphorylation, which was inhibited by GA, indicating that CREB is a downstream target of Hsp90/Akt signaling. Leu-Ile elevated the levels of GDNF mRNA and protein expression, whereas inhibition of CREB blocked such effects. Leu-Ile promoted the binding activity of phosphorylated CREB with cAMP response element. These findings show that CREB plays a key role in transcriptional regulation of GDNF expression induced by Leu-Ile. In conclusion, Leu-Ile activates Hsp90/Akt/CREB signaling, which contributes to the upregulation of GDNF expression. It may represent a novel lead compound for the treatment of dopaminergic neurons or motoneuron diseases.
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PMID:An analog of a dipeptide-like structure of FK506 increases glial cell line-derived neurotrophic factor expression through cAMP response element-binding protein activated by heat shock protein 90/Akt signaling pathway. 1655 84

Adaptive response of the yeast Saccharomyces cerevisiae to environmental alkalinization results in remodeling of gene expression. A key target is the gene ENA1, encoding a Na(+)-ATPase, whose induction by alkaline pH has been shown to involve calcineurin and the Rim101/Nrg1 pathway. Previous functional analysis of the ENA1 promoter revealed a calcineurin-independent pH responsive region (ARR2, 83 nucleotides). We restrict here this response to a small (42 nucleotides) ARR2 5.-region, named MCIR (minimum calcineurin independent response), which contains a MIG element, able to bind Mig1,2 repressors. High pH-induced response driven from this region was largely abolished in snf1 cells and moderately reduced in a rim101 strain. Cells lacking Mig1 or Mig2 repressors had a near wild type response, but the double mutant presented a high level of expression upon alkaline stress. Deletion of NRG1 (but not of NRG2) resulted in increased expression. Induction from the MCIR region was marginal in a quadruple mutant lacking Nrg1,2 and Mig1,2 repressors. In vitro band shift experiments demonstrated binding of Nrg1 to the 5. end of the ARR2 region. Furthermore, we show that Nrg1 binds in vivo around the MCIR region under standard growth conditions, and that binding is largely abolished after high pH stress. Therefore, the calcineurin-independent response of the ENA1 gene is under the regulation of Rim101 (through Nrg1) and Snf1 (through Nrg1 and Mig2). Accordingly, induction by alkaline stress of the entire ENA1 promoter in a snf1 rim101 mutant in the presence of the calcineurin inhibitor FK506 is completely abolished. Thus, the transcriptional response to alkaline stress of the ENA1 gene integrates three different signaling pathways.
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PMID:The transcriptional response of the yeast Na(+)-ATPase ENA1 gene to alkaline stress involves three main signaling pathways. 1702 28

Inquiries into the neurochemical mechanisms of vestibular compensation, a model of lesion-induced neuronal plasticity, reveal the involvement of both voltage-gated Ca(2+) channels (VGCC) and intracellular Ca(2+) signaling. Indeed, our previous microarray analysis showed an up-regulation of some calcium signaling-related genes such as the alpha2 subunit of L-type calcium channels, calcineurin, and plasma membrane Ca(2+) ATPase 1 (PMCA1) in the ipsilateral vestibular nuclear complex (VNC) following unilateral vestibular deafferentation (UVD). To further elucidate the role of calcium signaling-related molecules in vestibular compensation, we used a quantitative real-time polymerase chain reaction (PCR) method to confirm the microarray results and investigated changes in expression of these molecules at various stages of compensation (6 h to 2 weeks after UVD). We also investigated the changes in gene expression during Bechterew's phenomenon and the effects of a calcineurin inhibitor on vestibular compensation. Real-time PCR showed that genes for the alpha2 subunit of VGCC, PMCA2, and calcineurin were transiently up-regulated 6 h after UVD in ipsilateral VNC. A subsequent UVD, which induced Bechterew's phenomenon, reproduced a complete mirror image of the changes in gene expressions of PMCA2 and calcineurin seen in the initial UVD, while the alpha2 subunit of VGCC gene had a trend to increase in VNC ipsilateral to the second lesion. Pre-treatment by FK506, a calcineurin inhibitor, decelerated the vestibular compensation in a dose-dependent manner. Although it is still uncertain whether these changes in gene expression are causally related to the molecular mechanisms of vestibular compensation, this observation suggests that after increasing the Ca(2+) influx into the ipsilateral VNC neurons via up-regulated VGCC, calcineurin may be involved in their synaptic plasticity. Conversely, an up-regulation of PMCA2, a brain-specific Ca(2+) pump, would increase an efflux of Ca(2+) from those neurons and perhaps prevent cell damage following UVD.
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PMID:Unilateral vestibular deafferentation-induced changes in calcium signaling-related molecules in the rat vestibular nuclear complex. 1727 94

The past decade has witnessed multiple advances in our understanding of magnesium (Mg(2+)) homeostasis. The discovery that mutations in claudin-16/paracellin-1 or claudin-19 are responsible for familial hypomagnesemia with hypercalciuria and nephrocalcinosis provided insight into the molecular mechanisms governing paracellular transport of Mg(2+). Our understanding of the transcellular movement of Mg(2+) was similarly enhanced by the realization that defects in transient receptor potential melastatin 6 (TRPM6) cause hypomagnesemia with secondary hypocalcemia. This channel regulates the apical entry of Mg(2+) into epithelia. In so doing, TRPM6 alters whole-body Mg(2+) homeostasis by controlling urinary excretion. Consequently, investigation into the regulation of TRPM6 has increased. Acid-base status, 17beta estradiol, and the immunosuppressive agents FK506 and cyclosporine affect plasma Mg(2+) levels by altering TRPM6 expression. A mutation in epithelial growth factor is responsible for isolated autosomal recessive hypomagnesemia, and epithelial growth factor activates TRPM6. A defect in the gamma-subunit of the Na,K-ATPase causes isolated dominant hypomagnesemia by altering TRPM6 activity through a decrease in the driving force for apical Mg(2+) influx. We anticipate that the next decade will provide further detail into the control of the gatekeeper TRPM6 and, therefore, overall whole-body Mg(2+) balance.
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PMID:Molecular determinants of magnesium homeostasis: insights from human disease. 1856 69


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