Flowchart: Preparation: Muc1


Text Box: Tnfr                   




Text Box: Muc1

Breast cancer




Text Box: Egfr



Am J Physiol Lung Cell Mol Physiol. 2007 Jun 15; [Epub ahead of print]Click here to read Links

TNF-{alpha} induces MUC1 gene transcription in lung epithelial cells: Its signaling pathway and biological implication.

Koga T, Kuwahara I, Lillehoj EP, Lu W, Miyata T, Isohama Y, Kim KC.

Lovelace Respiratory Research Institute, Albuquerque, New Mexico, United States; Kumamoto University, Kumamoto, Japan.

The current study was conducted to elucidate the mechanism through which TNF-alpha stimulates expression of MUC1, a membrane-tethered mucin. A549 human lung adenocarcinoma cells treated with TNF-alpha exhibited significantly higher MUC1 protein levels in detergent lysates compared with cells treated with vehicle alone. Increased MUC1 protein levels were correlated with significantly higher levels of MUC1 mRNA in TNF-alpha-treated cells compared with controls. However, TNF-alpha did not alter MUC1 transcript stability, implying increased de novo transcription induced by the cytokine. TNF-alpha increased MUC1 gene promoter activity in A549 cells transfected with a promoter-luciferase reporter plasmid. Both U0126, an inhibitor of MEK1/2, and dominant negative ERK1 prevented TNF-alpha-induced MUC1 promoter activation, and anti-TNFR1 antibody blocked TNF-alpha-stimulated ERK1/2 activation. MUC1 promoter activation by TNF-alpha also was blocked by mithramycin A, an inhibitor of Sp1, as well as either deletion or mutation of a putative Sp1 binding site in the MUC1 promoter located between nucleotides -99 and -90. TNF-alpha-stimulated binding of Sp1 to the MUC1 promoter in intact cells was demonstrated by chromatin immunoprecipitation assay. We conclude that TNF-alpha induces MUC1 gene transcription through a TNFR1 --> MEK1/2 --> ERK1/2 --> Sp1 pathway. Key words: MUC1, TNF-alpha, signaling, Sp1, MAP kinase.

PMID: 17575006 [PubMed - as supplied by publisher]


Cancer Res. 2007 Jul 15;67(14):6591-8.Click here to read Links

Transforming Growth Factor {alpha} Dependent Cancer Progression Is Modulated by Muc1.

Pochampalli MR, Bitler BG, Schroeder JA.

Department of Molecular and Cellular Biology, Arizona Cancer Center, and Bio5 Institute, University of Arizona, Tucson, Arizona.

Transforming growth factor alpha (TGFalpha) is a potent inducer of cellular transformation, through its binding and activation of the epidermal growth factor receptor (EGFR). Previous studies in our laboratory showed that EGFR could also be affected by the glycoprotein MUC1, which inhibits ligand-stimulated degradation of EGFR in breast epithelial cell lines. To determine the effect of Muc1 expression on TGFalpha/EGFR-dependent breast transformation, we crossed the WAP-TGFalpha transgenic mouse model of breast cancer onto a Muc1-null background. We found that the loss of Muc1 expression dramatically affects mammary gland transformation and progression. Although 100% of WAP-TGFalpha/Muc1(+/+) mice form mammary gland tumors by 1 year, only 37% of WAP-TGFalpha/Muc1(-/-) form tumors by this time. This difference is also associated with a delay in onset, with a doubling of onset time observed in the WAP-TGFalpha/Muc1(-/-) compared with the WAP-TGFalpha/Muc1(+/+) mice. Analysis of signal transduction pathways revealed that activation of cyclin D1 expression is significantly suppressed in tumors derived from WAP-TGFalpha/Muc1(-/-) animals compared with those expressing Muc1. The loss of Muc1 expression also results in a significant inhibition in the formation of hyperplastic lesions during tumor progression. On the C57Bl/6 inbred background, pulmonary lesions were observed in 28 of 29 WAP-TGFalpha/Muc1(+/+) animals (including one metastatic pulmonary adenocarcinoma and multiple perivascular lymphomas), although none were detected in the WAP-TGFalpha/Muc1(-/-) animals. Together, these data indicate that Muc1 is an important modulator of TGFalpha-dependent tumor progression. [Cancer Res 2007;67(14):6591-8].

PMID: 17638868 [PubMed - in process]































Biochim Biophys Acta. 2007 Jul 6; [Epub ahead of print]

Molecular docking and spatial coarse graining simulations as tools to investigate substrate recognition, enhancer binding and conformational transitions in indoleamine-2,3-dioxygenase (IDO).

Macchiarulo A, Nuti R, Bellocchi D, Camaioni E, Pellicciari R.

Dipartimento di Chimica e Tecnologia del Farmaco, Università di Perugia, Via del Liceo 1, 06123 Perugia, Italy.

Indoleamine 2,3-dioxygenase (IDO) is an heme-containing enzyme involved in the regulation of important immunological responses and neurological processes. The enzyme catalyzes the oxidative cleavage of the pyrrole ring of the indole nucleus of tryptophan (Trp) to yield N-formylkynurenine, that is the initial and rate limiting step of the kynurenine pathway. Some indole derivatives have been reported to act as effectors of the enzyme by enhancing its catalytic activity. On the basis of the recent availability of the crystal structure of IDO, in this work we investigate substrate recognition and enhancer binding to IDO using molecular docking experiments. In addition, conformational transitions of IDO in response to substrate and enhancer binding are studied using coarse graining simulations with the program FIRST. The results enable us to identify (i) the binding site of enhancer modulators; (ii) the motion of an electrostatic gate that regulates the access of the substrate to the catalytic site of the enzyme; (iii) the movement of the anchoring region of a hairpin loop that may assist the shuttle of substrates/products to/from the catalytic site of IDO. These data, combined with available site-directed mutagenesis experiments, reveal that conformational transitions of IDO in response to substrate and enhancer binding are controlled by distinct combination of two conformational states (open and close) of the above structural motifs. On this basis, a molecular mechanism regarding substrate recognition and activity enhancement by indole derivatives is proposed.

PMID: 17644054 [PubMed - as supplied by publisher]

J Virol. 2007 Jul 11; [Epub ahead of print]Click here to read Links

Astrocyte indoleamine 2, 3 dioxygenase (IDO) is induced by the TLR3 ligand poly IC: mechanism of induction and role in anti-viral response.

Suh HS, Zhao ML, Rivieccio M, Choi S, Connolly E, Zhao Y, Takikawa O, Brosnan CF, Lee SC.

Department of Pathology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461 and National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Obu, Japan.

Indoleamine 2, 3-dioxygenase (IDO) is the first and rate-limiting enzyme in the kynurenine pathway of tryptophan catabolism and has been implicated in neurotoxicity and suppression of the antiviral T cell response in HIV encephalitis (HIVE). Here we show that the toll-like receptor 3 (TLR3) ligand polyinosinic-polycytidylic acid (PIC) induces the expression of IDO in human astrocytes. PIC was less potent than IFNgamma but more potent than IFNbeta in inducing IDO. PIC induction of IDO was in part mediated by IFNbeta but not IFNgamma, and both NF-kappaB and IRF3 were required. PIC also upregulated TLR3 thereby augmenting the primary (IFNbeta) and secondary (IDO and viperin) response genes upon subsequent stimulation with PIC. In HIVE, the transcripts for TLR3, IFNbeta, IDO and viperin were increased and IDO immunoreactivity was detected in reactive astrocytes as well as macrophages and microglia. PIC caused suppression of intracellular replication of VSVg-env HIV and human cytomegalovirus in a manner dependent on IRF3 and IDO. The involvement of IDO was demonstrated by partial but significant reversal of the PIC-mediated antiviral effect by IDO RNAi and/or tryptophan supplementation. Importantly, the cytokine IL-1 abolished IFNgamma-induced IDO enzyme activity in a nitric oxide-dependent manner without suppressing protein expression. Our results demonstrate that IDO is an innate antiviral protein induced by dsRNA and suggest a therapeutic utility for PIC in human viral infections. They also show that IDO activity can be dissociated from protein expression, indicating that the local CNS cytokine and nitric oxide environment determines IDO function.

PMID: 17626075 [PubMed - as supplied by publisher]






















Int Immunopharmacol. 2006 Jun;6(6):1034-8. Epub 2006 Feb 3.

Related Articles, Links

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IL-2 increased RANTES production and CD25 expression in cultured PBMCs only from antiretroviral treated HIV-1+ patients with detectable viral loads.

Lozano JM, Kindelan JM, Cabello A, Gonzalez R, Solana R, Pena J.

Department of Immunology, Hospital "Reina Sofia", University of Cordoba, Spain; Department of Virology, Pasteur Institute of Paris, France.

In order to better understand the possible beneficial effects of intermittent IL-2 treatment as complement of antiretroviral therapy in HIV-1+ patients, we have measured the levels of RANTES in the supernatants and the CD25 expression in cultured PBMCs obtained from HIV-1+ individuals in presence of IL-2. The results showed a significant increases in RANTES production and in the expression of CD25+ in the cultures with IL-2 of PBMC obtained from HIV-1+ patients with a detectable viral load in comparison with both, HIV-1+ patients with no detectable viral loads and with healthy individuals. These results suggest that therapeutic IL-2 administered in addition to highly active anti-retroviral therapy (HAART) may contribute to increase the effect of this therapy by rising both RANTES production and CD25 expression only in HIV-1+ patients with detectable viral loads.

PMID: 16644491 [PubMed - in process]






Science. 2005 Jun 3;308(5727):1477-80.Click here to read Links

The structure of interleukin-2 complexed with its alpha receptor.

Rickert M, Wang X, Boulanger MJ, Goriatcheva N, Garcia KC.

Departments of Microbiology and Immunology, and Structural Biology, Stanford University School of Medicine, 299 Campus Drive, Fairchild D319, Stanford, CA 94305-5124, USA.

Interleukin-2 (IL-2) is an immunoregulatory cytokine that binds sequentially to the alpha (IL-2Ralpha), beta (IL-2Rbeta), and common gamma chain (gammac) receptor subunits. Here we present the 2.8 angstrom crystal structure of a complex between human IL-2 and IL-2Ralpha, which interact in a docking mode distinct from that of other cytokine receptor complexes. IL-2Ralpha is composed of strand-swapped "sushi-like" domains, unlike the classical cytokine receptor fold. As a result of this domain swap, IL-2Ralpha uses a composite surface to dock into a groove on IL-2 that also serves as a binding site for antagonist drugs. With this complex, we now have representative structures for each class of hematopoietic cytokine receptor-docking modules.

PMID: 15933202 [PubMed - indexed for MEDLINE]
























The effect of therapeutic hypothermia on the expression of inflammatory response genes following moderate traumatic brain injury in the rat.

Truettner JS, Suzuki T, Dietrich WD.

Department of Neurological Surgery, The Neurotrauma Research Center, The Miami Project to Cure Paralysis, University of Miami School of Medicine, Miami, FL 33136, USA.

Traumatic brain injury (TBI) initiates a cascade of cellular and molecular responses including both pro- and anti-inflammatory. Although post-traumatic hypothermia has been shown to improve outcome in various models of brain injury, the underlying mechanisms responsible for these effects have not been clarified. In this study, inflammation cDNA arrays and semi-quantitative RT-PCR were used to detect genes that are differentially regulated after TBI. In addition, the effect of post-traumatic hypothermia on the expression of selective genes was also studied. Rats (n = 6-8 per group) underwent moderate fluid-percussion (F-P) brain injury with and without hypothermic treatment (33 degrees C/3 h). RNA from 3-h or 24-h survival was analyzed for the expression of IL1-beta, IL2, IL6, TGF-beta2, growth-regulated oncogene (GRO), migration inhibitory factor (MIF), and MCP (a transcription factor). The interleukins IL-1beta, IL-2, and IL-6 and TGF-beta and GRO were strongly upregulated early and transiently from 2- to 30-fold over sham at 3 h, with normalization by 24 h. In contrast, the expressions of MIF and MCP were both reduced by TBI compared to sham. Post-traumatic hypothermia had no significant effect on the acute expression of the majority of genes investigated. However, the expression of TGF-beta2 at 24 h was significantly reduced by temperature manipulation. The mechanism by which post-traumatic hypothermia is protective may not involve a general genetic response of the inflammatory genes. However, specific genes, including TGF-beta2, may be altered and effect cell death mechanisms after TBI. Hypothermia differentially regulates certain

genes and may target more delayed responses underlying the secondary damage following TBI.

PMID: 15922484 [PubMed - in process]









 erase chain reaction in unstimulated PBMC and in PBMC incubated for 4h in the presence of concanavalin A (ConA) or phorbol 12-myristate 13-acetate plus ionomycin (PMA/I). Compared to unstimulated cells, stimulation with ConA and PMA/I increased the IL2 mRNA transcription in average by 300- and 20-fold, respectively. Nevertheless, no significant differences in IL2 transcription between the genotypes could be detected. These findings were confirmed by band shift studies using different oligonucleotides containing variations of the potential binding motif, which showed no differences in the gel mobility after incubation with nuclear extract containing GATA-3. The obtained results argue against an impact of this polymorphism on the IL2 transcription and the genetic disease resistance in sheep.





















ICAM-1 gene expression in endothelial cells: Effects on the inhibition of STAT3 phosphorylation.
Resveratrol suppresses IL-6-induced
Wung BS, Hsu MC, Wu CC, Hsieh CW.

Department of Applied Microbiology, National Chiayi University, Chiayi, Taiwan.

Resveratrol, a polyphenolic phytoaxelin present in red wine, has been suggested to protect against atherosclerosis and cardiovascular disease because of its antioxidant effects. Intercellular adhesion molecule (ICAM-1), induced by cytokines, has been hypothesized to play a role in the early events during atherosclerosis. In this study we tested the effects of resveratrol upon both IL-6-induced ICAM-1 gene expression and its underlying signaling pathways in endothelial cells (ECs). Resveratrol was found to inhibit both TNFalpha- and IL-6-induced ICAM-1 gene expression at the promoter, transcriptional and protein levels. Resveratrol also abrogates the tyr705 phosphorylation of STAT3 in IL-6-treated ECs, in a dose- and time-dependent manner. Although quercetin had similar effects, resveratrol showed higher inhibitory properties following 2-4 h pretreatments. Resveratrol has been shown to induce the activity of endothelial nitric oxide synthase (eNOS) and increase NO production. Consistent with this, the treatment of ECs with a NO donor (SNAP) reduces IL-6-induced STAT3 phosphorylation. Conversely, exposure of ECs to a NOS inhibitor reversed the effects of resveratrol upon IL-6-induced STAT3 phosphorylation. Furthermore, ECs transfected with constitutively active Rac1 (RacV12) showed increases in ICAM-1 promoter activity, intracellular reactive oxygen species (ROS) levels and STAT3 phosphorylation, and these increases were attenuated by resveratrol treatment. In summary, we demonstrate for the first time that resveratrol inhibits IL-6-induced ICAM-1 gene expression, in part, by interfering with Rac-mediated pathways via the attenuation of STAT3 phosphorylation. This study therefore provides important new insights that may contribute to the proposed beneficial effects of resveratrol in endothelial responses to cytokines during inflammation.

PMID: 16150460 [PubMed - as supplied by publisher]


















Bradykinin mimics ischemic preconditioning by generating reactive oxygen species (ROS). To identify intermediate steps leading to ROS generation, rabbit cardiomyocytes were incubated in reduced MitoTracker Red that becomes fluorescent after exposure to ROS. Fluorescence intensity in treated cells was expressed as % of that in paired untreated cells. Bradykinin (500nM) caused 51+/-16% increase in ROS generation (p<0.001). Co-incubation with either bradykinin B2 receptor blocker HOE140 (5 micro M) or free radical scavenger N-(2-mercaptopropionyl) glycine (1mM) prevented this increase confirming the response was receptor-mediated and ROS were actually being measured. Closing mitochondrial KATP (mKATP)channels with 5-hydroxydecanoate (5HD, 1mM) prevented increased ROS generation. Bradykinin-induced ROS generation was blocked by L-NAME (200 micro M) implicating nitric oxide as an intermediate. Blockade of guanylyl cyclase with ODQ (10 micro M) aborted bradykinin-induced ROS generation, but not that from diazoxide, a direct opener of mKATP channels. The PKG blocker 8-Br-cGMPS (25 micro M) eliminated bradykinin's effect. Conversely, direct activation of PKG with 8-pCPT-cGMP (100 micro M) increased ROS generation (39+/-15%, p<0.004) similar to bradykinin. This increase was blocked by 5HD. Finally, the nitric oxide donor SNAP (1 micro M)increased ROS by 34+/-6%. This increase was also blocked by 5HD. In intact rabbit hearts bradykinin (400nM) decreased infarction from 30.5+/-3.0% of the risk zone in control hearts to 11.9+/-1.4% (p<0.01). This protection was aborted by either L-NAME (200 micro M) or ODQ (2 micro M)(35.4+/-5.7% and 30.4+/-3.0% infarction, resp., pNS vs control). Hence, bradykinin preconditions through receptor-mediated production of nitric oxide which activates guanylyl cyclase. The resulting cGMP activates PKG that opens mKATP. Subsequent release of ROS triggers cardioprotection.