Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pulmonary fibrosis is a progressive disorder characterized by the loss of alveolar architecture through epithelial and endothelial cell apoptosis and fibroblast proliferation. Recent studies showed that angiotensin-converting enzyme (ACE) activity is increased in fibrotic tissues, and ACE inhibitors administered in vivo ameliorate fibrosis, suggesting that ACE may play a critical role. However, the regulation of ACE expression is not well understood. In the present study, we demonstrate that bleomycin, a chemotherapeutic agent which induces pulmonary fibrosis in animals and humans, increases gene expression of ACE. Treatment of primary bovine pulmonary artery endothelial cells with 0.1 to 1.0 microg/ml bleomycin increased ACE enzymatic activity and ACE mRNA, as monitored by hippuryl-L-histidyl-L-leucine assay and competitive quantitative reverse transcriptase polymerase chain reaction (RT-PCR), respectively. Luciferase reporter constructs showed that upregulation of ACE transcription by bleomycin is mediated through element(s) in the 97-bp ACE promoter. Bleomycin activated p42/p44 mitogen-activated protein kinase (MAPK) and induced nuclear translocation and activation of the early growth response (Egr)-1 transcription factor, a factor previously shown to positively regulate ACE expression. The MAPK kinase1/2 (MEK1/2) inhibitor U0126 blocked MAPK and Egr-1 activation by bleomycin, suggesting that Egr-1 activation is MAPK dependent. These data provide the first evidence that bleomycin activates ACE gene expression through the MAPK pathway and Egr-1.
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PMID:Bleomycin upregulates gene expression of angiotensin-converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor. 1171 4

Aminoglycoside antibiotics (AGAs) are nephrotoxic, with most of the damage confined to the proximal tubule, but the mechanism for cellular toxicity is not clear. It has been previously shown that the extracellular-calcium sensing receptor (CaR) is expressed in intact rat proximal tubule and can be stimulated by the AGA neomycin. To investigate whether CaR could contribute to AGA-induced nephrotoxicity, the acute responses to various AGAs in the proximal tubule-derived opossum kidney (OK) cell line were examined. The presence in OK cells of CaR-related transcripts and protein was demonstrated by northern analyses, reverse transcriptase-PCR, immunocytochemistry, and immunoblotting. OK cells responded to elevated extracellular calcium (Ca(2+)(o)) and neomycin but also to gentamicin and tobramycin with an increase in cytosolic [Ca(2+)]. Ca(2+)(o), neomycin, and gentamicin also activated the extracellular signal-regulated kinases, ERK1 and ERK2. Neomycin-induced ERK activation was both dose- and time-dependent and was attenuated by inhibitors of phosphatidylinositol 3-kinase, phosphatidylinositol bisphosphate (PIP(2))-specific phospholipase C, and MEK1, but not of protein kinase C. Thus, proximal tubular OK cells express a CaR that mediates Ca(2+)(i) mobilization and PIP(2)-PLC-dependent ERK activation in response to AGAs and thus could play a role in AGA-induced nephrotoxicity.
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PMID:Aminoglycosides increase intracellular calcium levels and ERK activity in proximal tubular OK cells expressing the extracellular calcium-sensing receptor. 1203 77

Thrombin, a serine protease generated by the activation of the blood coagulation cascade following vessel injury, induces vascular endothelial growth factor-(VEGF) release. However, the molecular mechanism of thrombin-induced VEGF release is largely unknown. Anagonist of protease-activated receptor-i (PARI), SFLL-RNPNDKYEPF, mimicked thrombin-induced VEGF release in human vascular smooth muscle (HVSM) cells, as determined by enzyme-linked immunosorbent assay, reverse transcriptase-polymerase chain reaction, and Northern blotting. In contrast, the agonist of PAR3, TFR- GAP, did not affect VEGF release or expression. SFLL-RNPNDKYEPF, but not TFRGAP, up-regulated [Ca2-]i.Moreover, the calcium ionophone A23187 was found to trigger VEGF release in HVSM cells. Thrombin-inducedVEGF release was blocked by anti-thrombin, heparin, a synthetic thrombin receptor inhibitor E5510, the calcium chelator BAPTA, the protein kinase C inhibitor calphostin C, and the MEK1/2 inhibitor U0126. Thus, our data show that thrombin caused VEGF release via PARI activation in a manner dependent on [Ca2+]i and p44/42 downstream from the receptor activation.
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PMID:The agonist of the protease-activated receptor-1 (PAR) but not PAR3 mimics thrombin-induced vascular endothelial growth factor release in human smooth muscle cells. 1451 37

Mechanical strain plays a crucial role in bone remodeling during growth and development and healing of bone besides systemic and local factors. One of the major factors involves in remodeling process is matrix metalloproteinases (MMPs) such as MMP-13 that has been shown to degrade the native interstitial collagens in several tissues. To study how mechanical strain affects extracellular matrix degradation by MMP-13, a biaxial strain was applied to MC3T3-E1 osteoblastic cells plated onto a collagen-coated flexible elastic membrane. The MMP-13 protein and mRNA expression were determined by Western blotting and reverse transcriptase-PCR, respectively. The zymographic activities of MMP-13 increased dramatically at 30 min, reached a peak by 2-fold at 1 h, and maintained up to 4 h. Moreover, the MMP-13 and c-fos mRNA expressed at 5 min, increased to 2.8- and 3-fold at 1 h, respectively, and gradually declined thereafter. Cycloheximide and actinomycin D did not inhibit the MMP-13 and c-fos mRNA expression, suggesting that such expression does not require de novo protein synthesis and not change their stabilities. To investigate which of the mitogen-activated protein kinase (MAPK) pathways involves in the expression of MMP-13, inhibitors such as PD98059, SB203580, and SP600125 were used. However, only PD98059 (an inhibitor of MEK1/2 activation) inhibited MMP-13 and c-fos gene expression; the result was further substantiated by transfecting with the dominant negative mutants of MEK1/2 (MEK K97R) and ERK2. Taken together, our results showed that mechanical strain induces the MMP-13 expression through MEK-ERK signaling pathway to regulate mechanical adaptation.
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PMID:Mechanical strain induces collagenase-3 (MMP-13) expression in MC3T3-E1 osteoblastic cells. 1504 66

Vasoactive intestinal peptide (VIP) upregulates the expression of vascular endothelial cell growth factor (VEGF(189), VEGF(165) and VEGF(121)) mRNAs in human prostate cancer LNCaP cells, as shown by reverse transcriptase-polymerase chain reaction (RT-PCR). Real-time RT-PCR indicated that the effect was maximal by 1-2 h and must be accounted for increased transcription since VIP decreased VEGF(165) mRNA stability. VIP stimulated VEGF(165) protein synthesis as measured by ELISA. VIP regulation of VEGF expression was mediated by VPAC(1) receptor and was cAMP/protein kinase A (PKA) dependent. Phosphoinositide 3-kinase (PI3-K) and mitogen-activated protein kinase MEK1/2 systems may also be involved as shown with specific kinase inhibitors. These actions together with the observation of VIP-induced neuroendocrine differentiation in LNCaP cells suggest a proangiogenic potential of VIP in prostate cancer.
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PMID:Vasoactive intestinal peptide increases vascular endothelial growth factor expression and neuroendocrine differentiation in human prostate cancer LNCaP cells. 1509 99

To understand how mitogenic signals are transduced into the trophoblasts in preimplantation embryos, the expression of mitogen-activated protein kinase (MAPK) pathway molecules was tested. We used immunocytochemical means and reverse transcriptase-polymerase chain reaction to test whether MAPK pathway molecule gene products exist at the protein and phosphoprotein level in the zygote and the RNA level in the egg and zygote. In addition, all antibodies detected the correct-sized major band in Westerns of placental cell lines representing the most prevalent cell type in preimplantation embryos. A majority of mRNA transcripts of MAPK pathway genes were detected in unfertilized eggs, and all were expressed in the zygote. We found that the MAPK pathway protein set consisting of the following gene products was present: FRS2 alpha, GRB2, GAB1, SOS1, Ha-ras, Raf1/RafB, MEK1,2,5, MAPK/ERK1,2, MAPK/ERK5, and RSK1,2,3 (see abbreviations). These proteins were detected in trophoblasts in embryonic day (E) 3.5 embryos when they could mediate mitogenic fibroblast growth factor signals from the embryo or colony stimulating factor-1 signals from the uterus. The phosphorylation state and position of the phosphoproteins in the cells suggested that they might function in mediating mitogenic signals. Interestingly, a subtle transition from maternal MAPK function to zygotic function was suggested by the localization for three MAPK pathway enzymes between E2.5 and E3.5, Raf1 phospho is largely cell membrane-localized at E2.5 and E3.5, and MEK1,2 phospho accumulates in the nucleus on E2.5 and E3.5. However, MAPK phospho shifts from nuclear accumulation at E2.5 to cytoplasmic accumulation at E3.5. This finding is similar to the cytoplasmic MAPK phospho localization reported in fibroblast growth factor signaling fields in postimplantation embryos (Corson et al. [2003] Development 130:4527-4537). This spatial and temporal expression study lays a foundation to plan and analyze perturbation studies aimed at understanding the role of the major mitogenic pathway in preimplantation mouse embryos.
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PMID:Entire mitogen activated protein kinase (MAPK) pathway is present in preimplantation mouse embryos. 1530 88

Increased expression of transforming growth factor (TGF)-beta(1) and tumor necrosis factor (TNF)-alpha are thought to play important roles in the development of pulmonary fibrosis. We recently reported that TNF-alpha upregulates TGF-beta(1) expression in primary mouse lung fibroblasts (MLFs), a key cell population in fibrogenesis. In the present study, we have investigated signal transduction pathways involved in TNF-alpha upregulation of TGF-beta(1) in both primary MLFs and the Swiss 3T3 fibroblast cell line. Treatment of fibroblasts with TNF-alpha resulted in a significant increase in TGF-beta(1) protein as measured by ELISA. The increase in protein was preceded by a 200-400% increase in TGF-beta(1) mRNA detected by quantitative, real-time, reverse transcriptase-polymerase chain reaction. Western blot analysis showed that TNF-alpha activated the extracellular signal-regulated kinase (ERK), and inhibitors of the ERK-specific mitogen-activated protein kinase pathway (PD98059 or U0126) blocked TNF-alpha induction of TGF-beta(1) mRNA and protein. mRNA stability experiments showed that TNF-alpha increased the half-life of TGF-beta(1) mRNA to more than 24 h compared with approximately 15 h in unstimulated cells. Expression of constitutively active MEK1 that selectively phosphorylates ERK was sufficient for TGF-beta(1) mRNA stabilization in Swiss 3T3 fibroblasts. These results indicate that TNF-alpha activates the ERK-specific mitogen-activated protein kinase pathway leading to increased TGF-beta(1) production in fibroblasts, primarily via a post-transcriptional mechanism that involves stabilization of the TGF-beta(1) transcript.
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PMID:Tumor necrosis factor-alpha induces transforming growth factor-beta1 expression in lung fibroblasts through the extracellular signal-regulated kinase pathway. 1565 32

Kaposi's sarcoma-associated herpesvirus (KSHV) in vitro target cell infection is characterized by the expression of the latency-associated genes ORF 73 (LANA-1), ORF 72, and K13 and by the transient expression of a very limited number of lytic genes such as lytic cycle switch gene ORF 50 (RTA) and the immediate early (IE) lytic K5, K8, and v-IRF2 genes. During the early stages of infection, several overlapping multistep complex events precede the initiation of viral gene expression. KSHV envelope glycoprotein gB induces the FAK-Src-PI3K-RhoGTPase (where FAK is focal adhesion kinase) signaling pathway. As early as 5 min postinfection (p.i.), KSHV induced the extracellular signal-regulated kinase 1 and 2 (ERK1/2) via the PI3K-PKCzeta-MEK pathway. In addition, KSHV modulated the transcription of several host genes of primary human dermal microvascular endothelial cells (HMVEC-d) and fibroblast (HFF) cells by 2 h and 4 h p.i. Neutralization of virus entry and infection by PI-3K and other cellular tyrosine kinase inhibitors suggested a critical role for signaling molecules in KSHV infection of target cells. Here we investigated the induction of ERK1/2 by KSHV and KSHV envelope glycoproteins gB and gpK8.1A and the role of induced ERK in viral and host gene expression. Early during infection, significant ERK1/2 induction was observed even with low multiplicity of infection of live and UV-inactivated KSHV in serum-starved cells as well as in the presence of serum. Entry of UV-inactivated virus and the absence of viral gene expression suggested that ERK1/2 induction is mediated by the initial signal cascade induced by KSHV binding and entry. Purified soluble gpK8.1A induced the MEK1/2 dependent ERK1/2 but not ERK5 and p38 mitogen-activated protein kinase (MAPK) in HMVEC-d and HFF. Moderate ERK induction with soluble gB was seen only in HMVEC-d. Preincubation of gpK8.1A with heparin or anti-gpK8.1A antibodies inhibited the ERK induction. U0126, a selective inhibitor for MEK/ERK blocked the gpK8.1A- and KSHV-induced ERK activation. ERK1/2 inhibition did not block viral DNA internalization and had no significant effect on nuclear delivery of KSHV DNA during de novo infection. Analyses of viral gene expression by quantitative real-time reverse transcriptase PCR revealed that pretreatment of cells with U0126 for 1 h and during the 2-h infection with KSHV significantly inhibited the expression of ORF 73, ORF 50 (RTA), and the IE-K8 and v-IRF2 genes. However, the expression of lytic IE-K5 gene was not affected significantly. Expression of ORF 73 in BCBL-1 cells was also significantly inhibited by preincubation with U0126. Inhibition of ERK1/2 also inhibited the transcription of some of the vital host genes such as DUSP5 (dual specificity phosphatase 5), ICAM-1 (intercellular adhesion molecule 1), heparin binding epidermal growth factor, and vascular endothelial growth factor that were up-regulated early during KSHV infection. Several MAPK-regulated host transcription factors such as c-Jun, STAT1alpha, MEF2, c-Myc, ATF-2 and c-Fos were induced early during infection, and ERK inhibition significantly blocked the c-Fos, c-Jun, c-Myc, and STAT1alpha activation in the infected cells. AP1 transcription factors binding to the RTA promoter in electrophoretic mobility shift assays were readily detected in the infected cell nuclear extracts which were significantly reduced by ERK inhibition. Together, these results suggest that very early during de novo infection, KSHV induces the ERK1/2 to modulate the initiation of viral gene expression and host cell genes, which further supports our hypothesis that beside the conduit for viral DNA delivery into the cytoplasm, KSHV interactions with host cell receptor(s) create an appropriate intracellular environment facilitating infection.
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PMID:ERK1/2 and MEK1/2 induced by Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) early during infection of target cells are essential for expression of viral genes and for establishment of infection. 1605 24

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is rarely mutated in pancreatic cancers, but its regulation by transforming growth factor (TGF)-beta might mediate growth suppression and other oncogenic actions. Here, we examined the role of TGFbeta and the effects of oncogenic K-RAS/ERK upon PTEN expression in the absence of SMAD4. We utilized two SMAD4-null pancreatic cell lines, CAPAN-1 (K-RAS mutant) and BxPc-3 (WT-K-RAS), both of which express TGFbeta surface receptors. Cells were treated with TGFbeta1 and separated into cytosolic/nuclear fractions for western blotting with phospho-SMAD2, SMAD 2, 4 phospho-ATP-dependent tyrosine kinases (Akt), Akt and PTEN antibodies. PTEN mRNA levels were assessed by reverse transcriptase-polymerase chain reaction. The MEK1 inhibitor, PD98059, was used to block the downstream action of oncogenic K-RAS/ERK, as was a dominant-negative (DN) K-RAS construct. TGFbeta increased phospho-SMAD2 in both cytosolic and nuclear fractions. PD98059 treatment further increased phospho-SMAD2 in the nucleus of both pancreatic cell lines, and DN-K-RAS further improved SMAD translocation in K-RAS mutant CAPAN cells. TGFbeta treatment significantly suppressed PTEN protein levels concomitant with activation of Akt by 48 h through transcriptional reduction of PTEN mRNA that was evident by 6 h. TGFbeta-induced PTEN suppression was reversed by PD98059 and DN-K-RAS compared with treatments without TGFbeta. TGFbeta-induced PTEN expression was inversely related to cellular proliferation. Thus, oncogenic K-RAS/ERK in pancreatic adenocarcinoma facilitates TGFbeta-induced transcriptional down-regulation of the tumor suppressor PTEN in a SMAD4-independent manner and could constitute a signaling switch mechanism from growth suppression to growth promotion in pancreatic cancers.
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PMID:RAS/ERK modulates TGFbeta-regulated PTEN expression in human pancreatic adenocarcinoma cells. 1763 24

Antimicrobial peptides (AMPs) are important components of our first line of defense. Induction of AMPs such as LL-37 of the cathelicidin family might provide a novel approach in treating bacterial infections. In this study we identified 4-phenylbutyrate (PBA) as a novel inducer of AMP expression and investigated affected regulatory pathways. We treated various cell lines with PBA and assessed mRNA expression by real-time reverse transcriptase PCR (RT-PCR). Cathelicidin AMP (CAMP) gene expression was found to be upregulated in all four cell lines tested. Additionally, we found that the beta-defensin 1 gene was upregulated in the lung epithelial cell line VA10 while being downregulated in the monocytic cell line U937. Further we found that PBA induced CAMP gene expression synergistically with 1,25-dihydroxyvitamin D(3) at both protein and mRNA levels. The general mechanism of induction of CAMP gene expression by PBA was found to be dependent on protein synthesis. Results from quantitative chromatin immunoprecipitation experiments challenge the common view that histone deacetylase inhibitors directly increase CAMP gene expression. Furthermore, we have demonstrated that inhibition of the mitogen-activated protein kinases MEK1/2 and c-Jun N-terminal kinase attenuate PBA-induced CAMP gene expression. Similarly, alpha-methylhydrocinnamate (ST7), an analogue of PBA, increases CAMP gene expression. Our findings contribute to understanding of the regulation of AMP expression and suggest that PBA and/or ST7 is a promising drug candidate for treatment of microbial infections by strengthening the epithelial antimicrobial barriers.
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PMID:Phenylbutyrate induces antimicrobial peptide expression. 1977 Feb 73


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