FEBS JOURNAL, Issue 24 2000
Hwan Young Lee
A novel gene, matR, located upstream of matABC, transcribed in the opposite direction, and encoding a putative regulatory protein by sequence analysis was discovered from Rhizobium leguminosarum bv. trifolii. The matA, matB, and matC genes encode malonyl-CoA decarboxylase, malonyl-CoA synthetase, and a presumed malonate transporter, respectively. Together, these enzymes catalyze the uptake and conversion of malonate to acetyl-CoA. The deduced amino-acid sequence of matR showed sequence similarity with GntR from Bacillus subtilis in the N-terminal region encoding a helix-turn-helix domain. Electrophoretic mobility shift assay indicated that MatR bound to a fragment of DNA corresponding to the mat promoter region. The addition of malonate or methylmalonate increased the association of MatR and DNA fragment. DNase I footprinting assays identified a MatR binding site encompassing 66 nucleotides near the mat promoter. The mat operator region included an inverted repeat (TCTTGTA/TACACGA) centered ,46.5 relative to the transcription start site. Transcriptional assays, using the luciferase gene, revealed that MatR represses transcription from the mat promoter and malonate alleviates MatR-mediated repression effect on the expression of Pmat -luc+ reporter fusion. [source]

The Balance Between Concurrent Activation of ERs and PPARs Determines Daidzein-Induced Osteogenesis and Adipogenesis,

JOURNAL OF BONE AND MINERAL RESEARCH, Issue 5 2004
ZhiChao Dang PhD
Abstract The soy phytoestrogen daidzein has biphasic dose responses, but the underlying mechanisms are not yet clear. Transcriptional and biochemical data show that PPARs, in addition to ERs, are molecular targets of daidzein, which divergently regulates osteogenesis and adipogenesis. Dose responses are the result of a balance among PPARs and between ERs and PPARs. Introduction: Soy phytoestrogens have been used for the purposes of treatment and prevention of osteoporosis. Biphasic dose responses of daidzein, one of the main soy phytoestrogens, have long been recognized, but the underlying molecular mechanisms of action are not yet clear. Materials and Methods: Mouse bone marrow cells and mouse osteoprogenitor KS483 cells that concurrently differentiate into osteoblasts and adipocytes were cultured. Biochemical measurement of alkaline phosphatase (ALP) activity, RT-PCR, and gene reporter assays were used in this study. Results: Daidzein, one of the major soy phytoestrogens, had biphasic effects on osteogenesis and adipogenesis. Daidzein stimulated osteogenesis (ALP activity and nodule formation) and decreased adipogenesis (the number of adipocytes) at concentrations below 20 ,M, whereas it inhibited osteogenesis and stimulated adipogenesis at concentrations higher than 30 ,M. When estrogen receptors (ERs) were blocked by ICI182,780, daidzein-induced effects were not biphasic. A decrease in osteogenesis and an increase in adipogenesis were observed at the concentrations higher than 20 and 10 ,M, respectively. In addition to ERs, daidzein transactivated not only peroxisome proliferator-activate receptor , (PPAR,), but also PPAR, and PPAR, at micromolar concentrations. Activation of PPAR, had no direct effects on osteogenesis and adipogenesis. In contrast, activation of PPAR, stimulated osteogenesis but had no effects on adipogenesis, whereas PPAR, inhibited osteogenesis and stimulated adipogenesis. Transfection experiments show that an activation of PPAR, or PPAR, by daidzein downregulated its estrogenic transcriptional activity, whereas activation of PPAR, upregulated its estrogenic transcriptional activity. Activation of ER, or ER, by daidzein downregulated PPAR, transcriptional activity but had no influence on PPAR, or PPAR, transcriptional activity. Conclusions: Daidzein at micromolar concentrations concurrently activates different amounts of ERs and PPARs, and the balance of the divergent actions of ERs and PPARs determines daidzein-induced osteogenesis and adipogenesis. [source]

Transcriptional and post-transcriptional control of DNA methyltransferase 3B is regulated by phosphatidylinositol 3 kinase/Akt pathway in human hepatocellular carcinoma cell lines

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2010
Chuanzhong Mei
Abstract DNA methyltransferases (DNMTs) are essential for maintenance of aberrant methylation in cancer cells and play important roles in the development of cancers. Unregulated activation of PI3K/Akt pathway is a prominent feature of many human cancers including human hepatocellular carcinoma (HCC). In present study, we found that DNMT3B mRNA and protein levels were decreased in a dose- and time-dependent manner in HCC cell lines with LY294002 treatment. However, we detected that LY294002 treatment did not induce increase of the degradation of DNMT3B protein using protein decay assay. Moreover we found that Akt induced alteration of the expression of DNMT3B in cells transfected with myristylated variants of Akt2 or cells transfected with small interfering RNA respectively. Based on DNMT3B promoter dual-luciferase reporter assay, we found PI3K pathway regulates DNMT3B expression at transcriptional level. And DNMT3B mRNA decay analysis suggested that down-regulation of DNMT3B by LY294002 is also post-transcriptional control. Furthermore, we demonstrated that LY294002 down-regulated HuR expression in a time-dependent manner in BEL-7404. In summary, we have, for the first time, demonstrate that PI3K/Akt pathway regulates the expression of DNMT3B at transcriptional and post-transcriptional levels, which is particularly important to understand the effects of PI3K/Akt and DNMT3B on hepatocarcinogenesis. J. Cell. Biochem. 111: 158,167, 2010. © 2010 Wiley-Liss, Inc. [source]

Site-specific proteolysis of cyclooxygenase-2: A putative step in inflammatory prostaglandin E2 biosynthesis

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2007
Arturo Mancini
Abstract Cyclooxygenase-2 (COX-2) catalyzes the rate-limiting step in inflammatory prostanoid biosynthesis. Transcriptional, post-transcriptional, and post-translational covalent modifications have been defined as important levels of regulation for COX-2 gene expression. Here, we describe a novel regulatory mechanism in primary human cells involving regulated, sequence-specific proteolysis of COX-2 that correlates with its catalytic activity and ultimately, the biosynthesis of prostaglandin E2 (PGE2). Proinflammatory cytokines induced COX-2 expression and its proteolysis into stable immunoreactive fragments of 66, 42,44, 34,36, and 28 kDa. Increased COX-2 activity (PGE2 release) was observed coincident with the timing and degree of COX-2 proteolysis with correlation analysis confirming a linear relationship (R2,=,0.941). Inhibition of induced COX-2 activity with non-steroidal anti-inflammatory drugs (NSAIDs) and COX-2 selective inhibitors also abrogated cleavage. To determine if NSAID inhibition of proteolysis was related to drug-binding-induced conformational changes in COX-2, we assayed COX-inactive NSAID derivatives that fail to bind COX-2. Interestingly, these compounds suppressed COX-2 activity and cleavage in a correlated manner, thus suggesting that the observed NSAID-induced inhibition of COX-2 cleavage occurred through COX-independent mechanisms, presumably through the inhibition of proteases involved in COX-2 processing. Corroborating this observation, COX-2 cleavage and activity were mutually suppressed by calpain/cathepsin protease inhibitors. Our data suggest that the nascent intracellular form of COX-2 may undergo limited proteolysis to attain full catalytic capacity. J. Cell. Biochem. 101: 425,441, 2007. © 2006 Wiley-Liss, Inc. [source]

Transcriptional and proteolytic regulation of the insulin-like growth factor-I system of equine articular chondrocytes by recombinant equine interleukin-1,

JOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2006
Ryan M. Porter
Interleukin-1 (IL-1) and insulin-like growth factor-I (IGF-I), which have opposing effects on matrix metabolism within articular cartilage, are thought to play prominent roles in the pathogenesis of osteoarthritis. To better understand the link between these anabolic (IGF-I) and catabolic (IL-1) stimuli, we examined exogenous IL-1 regulation of the IGF-I signaling system of articular chondrocytes (ACs). Equine ACs from non-arthritic stifle joints were expanded in monolayer culture, encapsulated for 10 days in alginate beads, and stimulated as high-density monolayers with recombinant equine IL-1, (0, 1, 10 ng/ml) for 48 h. IL-1, enhanced expression of IGF-IR levels, as determined by both [125I]-IGF-I binding studies and Western blotting, while reducing the concentration of endogenous IGF-I detected in conditioned media by radioimmunoassay. Western ligand blotting revealed that chondrocytes primarily secreted IGF binding proteins (IGFBPs) with molecular weights of 28,30 and 32,34 kDa, which were identified as IGFBPs 5 and 2, respectively, and that IL-1, treatment diminished IGFBP-2, the prominent homolog in conditioned media. Northern blot analysis suggested IL-1, regulation of IGF-I and, to some extent, IGF-IR was mediated by transcription; however, the cytokine did not affect IGFBP-2 expression. To test for evidence of proteolysis by matrix metalloproteinases (MMPs), additional cultures were co-incubated with inhibitors for MMPs 2/9, 3, and 8. IGFBP-2 suppression was partially reversed by gelatinase (MMP-2/9) inhibition. In summary, these findings further delineate the role of IL-1 as a key regulator of the IGF-I system within articular cartilage, demonstrating that regulation occurs through both direct (transcriptional) and indirect (proteolytic) mechanisms. J. Cell. Physiol. 209: 542,550, 2006. © 2006 Wiley-Liss, Inc. [source]

Transcriptional and translational control of C/EBPs: The case for "deep" genetics to understand physiological function

BIOESSAYS, Issue 8 2010
Claus Nerlov
Abstract The complexity of organisms is not simply determined by the number of their genes, but to a large extent by how gene expression is controlled. In addition to transcriptional regulation, this involves several layers of post-transcriptional control, such as translational repression, microRNA-mediated mRNA degradation and translational inhibition, alternative splicing, and the regulated generation of functionally distinct gene products from a single mRNA through alternative use of translation initiation sites. Much progress has been made in describing the molecular basis for these gene regulatory mechanisms. However, it is now a major challenge to translate this knowledge into deeper understanding of the physiological processes, both normal and pathological, that they govern. Using the C/EBP family of transcription factors as an example, the present review describes recent genetic experiments addressing this general problem and discusses how the physiological importance of newly discovered regulatory mechanisms might be determined. [source]

dTrf2 is required for transcriptional and developmental responses to ecdysone during Drosophila metamorphosis

DEVELOPMENTAL DYNAMICS, Issue 11 2007
Arash Bashirullah
Abstract The TATA box-binding protein (TBP) related factor 2 (TRF2) has been well characterized at a biochemical level and in cultured cells. Relatively little, however, is known about how TRF2 functions in specific biological pathways during development. Here, we show that Drosophila TRF2 (dTRF2) plays an essential role in responses to the steroid hormone ecdysone during the onset of metamorphosis. Hypomorphic dTrf2 mutations lead to developmental arrest during prepupal and early pupal stages with defects in major ecdysone-triggered biological responses, including puparium formation, anterior spiracle eversion, gas bubble translocation, adult head eversion, and larval salivary gland cell death. The transcription of key ecdysone-regulated target genes is delayed and reduced in dTrf2 mutants. dTrf2 appears to be required for the proper timing and levels of ecdysone-regulated gene expression required for entry into metamorphosis. Developmental Dynamics 236:3173,3179, 2007. © 2007 Wiley-Liss, Inc. [source]

Expression of a dominant negative form of Daxx in vivo rescues motoneurons from Fas (CD95)-induced cell death

DEVELOPMENTAL NEUROBIOLOGY, Issue 2 2005
Cedric Raoul
Abstract Fas-induced death of motoneurons in vitro has been shown to involve two signaling cascades that act together to execute the death program: a Fas-Daxx-ASK-1-p38 kinase-nNOS branch, which controls transcriptional and post-translational events, and the second classical Fas-FADD-caspase-8 branch. To analyze the role of Daxx in the developmental motoneuron cell death, we studied Fas-dependent cell death in motoneurons from transgenic mice that overexpress a dominant-negative form of Daxx. Motoneurons purified from these transgenic mice are resistant to Fas-induced death. This protective effect is specific to Fas because ultraviolet irradiation-triggered death is not affected by the transgene. The Daxx and the FADD pathways work in parallel because only Daxx, but not FADD, is involved in the transcriptional control of neuronal nitric oxide synthase and nitric oxide production. Nevertheless, we do not observe involvement of Daxx in developmental motoneuronal cell death, as the pattern of naturally occurring programmed cell death in vivo is normal in transgenic mice overexpressing the dominant negative form of Daxx, suggesting that Daxx-independent pathways are used during development. © 2004 Wiley Periodicals, Inc. J Neurobiol, 2005 [source]

Development of the specialized AMPA receptors of auditory neurons

DEVELOPMENTAL NEUROBIOLOGY, Issue 3 2002
Steven G. Sugden
Abstract At maturity, the AMPA receptors of auditory neurons exhibit very rapid desensitization kinetics and high permeability to calcium, reflecting the predominance of GluR3 flop and GluR4 flop subunits and the paucity of GluR2. We used mRNA analysis and immunoblotting to contrast the development of AMPA receptor structure in the chick cochlear nucleus [nucleus magnocellularis (NM)] with that of the slowly desensitizing and calcium-impermeable AMPA receptors of brainstem motor neurons in the nucleus of the glossopharyngeal/vagal nerves. The relative abundance of transcripts for GluRs 1,4 changes substantially in auditory (but not motor) neurons after embryonic day (E)10, with large decreases in GluR2 and increases in GluR3 and GluR4. Relative to the motor neurons, NM neurons show a higher abundance of flop isoforms of GluRs 2,4 at E10, suggesting that auditory neurons are already biased toward expression of flop isoforms before the onset of synaptic function at E11. Immunoreactivities in NM show very distinct developmental patterns from E13 onward: GluR2 declines by >90%, GluR3 increases threefold, and GluR4 remains relatively constant. Our results show that there are a series of critical points during normal development, most occurring after the onset of function, when rapid changes in receptor structure (occurring via both transcriptional and post-transcriptional control mechanisms) produce the specialized AMPA receptor functions that enable auditory neurons to accurately encode acoustic information. © 2002 Wiley Periodicals, Inc. J Neurobiol 52: 189,202, 2002 [source]

Exendin-4 protects pancreatic beta cells from human islet amyloid polypeptide-induced cell damage: potential involvement of AKT and mitochondria biogenesis

DIABETES OBESITY & METABOLISM, Issue 9 2010
R. Fan
Aim: Glucagon-like peptide-1 (GLP-1) stimulates beta-cell proliferation and enhances beta-cell survival, whereas oligomerization of human islet amyloid polypeptide (hIAPP) may induce beta-cell apoptosis and reduce beta-cell mass. Type 2 diabetes is associated with increased expression of IAPP. As GLP-1-based therapy is currently developed as a novel antidiabetic therapy, we examined the potential protective action of the GLP-1 receptor agonist exendin-4 on hIAPP-induced beta-cell apoptosis. Methods: The study was performed in clonal insulinoma (INS-1E) cells. Both method of transcriptional and translational and sulphorhodamine B (SRB) assays were used to evaluate cell viability and cell mass. Western blot analysis was applied to detect protein expression. Transfection of constitutively active protein kinase B (PKB/AKT) was performed to examine the role of AKT. Mitochondrial biogenesis was quantified by mitogreen staining and RT-PCR. Results: First, we confirmed that hIAPP induced cell apoptosis and growth inhibition in INS-1E cells. These effects were partially protected by exendin-4 in association with partial recovery of the hIAPP-mediated AKT inhibition. Furthermore, AKT constitutive activation attenuated hIAPP-induced apoptosis, whereas PI3K/AKT inhibition abrogated exendin-4-mediated effects. These findings suggest that the antiapoptotic and proliferative effects of exendin-4 in hIAPP-treated INS-1E cells were partially mediated through AKT pathway. Moreover, hIAPP induced FOXO1 but inhibited pdx-1 nucleus translocation. These effects were restored by exendin-4. Finally, mitogreen staining and RT-PCR revealed enhanced mitochondrial biogenesis by exendin-4 treatment. Conclusions: Collectively, these results suggest that GLP-1 receptor agonist protects beta cells from hIAPP-induced cell death partially through the activation of AKT pathway and improved mitochondrial function. [source]

Quorum sensing: the power of cooperation in the world of Pseudomonas

ENVIRONMENTAL MICROBIOLOGY, Issue 4 2005
Mario Juhas
Summary Work over the past few years has provided evidence that quorum sensing is a generic regulatory mechanism that allows bacteria to launch a unified, coordinated response in a population density-dependent manner to accomplish tasks which would be difficult, if not impossible, to achieve for a single bacterial cell. Quorum sensing systems are widespread among pseudomonads and the one of the human opportunistic pathogen Pseudomonas aeruginosa belongs to the most extensively studied cell-to-cell communication systems. In this organism, quorum sensing is highly complex and is made up of two interlinked N- acyl homoserine lactone (AHL)-dependent regulatory circuits, which are further modulated by a non-AHL-related signal molecule and numerous regulators acting both at the transcriptional and post-transcriptional level. This genetic complexity may be one of the key elements responsible for the tremendous environmental versatility of P. aeruginosa. Work of the past few years showed that quorum sensing is essential for the expression of a battery of virulence factors as well as for biofilm formation in P. aeruginosa and thus represents an attractive target for the design of novel drugs for the treatment of P. aeruginosa infections. Furthermore, the cell-to-cell communication ability was also demonstrated in a number of additional pseudomonads. [source]

Increased temperature and protein oxidation lead to HSP72 mRNA and protein accumulation in the in vivo exercised rat heart

EXPERIMENTAL PHYSIOLOGY, Issue 1 2009
Jessica L. Staib
Expression of myocardial heat shock protein 72 (HSP72), mediated by its transcription factor, heat shock factor 1 (HSF1), increases following exercise. However, the upstream stimuli governing exercise-induced HSF1 activation and subsequent Hsp72 gene expression in the whole animal remain unclear. Exercise-induced increases in body temperature may promote myocardial radical production, leading to protein oxidation. Conceivably, myocardial protein oxidation during exercise may serve as an important signal to promote nuclear HSF1 migration and activation of Hsp72 expression. Therefore, these experiments tested the hypothesis that prevention of exercise-induced increases in body temperature attenuates cardiac protein oxidation, diminishes HSF1 activation and decreases HSP72 expression in vivo. To test this hypothesis, in vivo exercise-induced changes in body temperature were manipulated by exercising male rats in either cold (4°C) or warm ambient conditions (22°C). Warm exercise increased both body temperature (+3°C) and myocardial protein oxidation, whereas these changes were attenuated by cold exercise. Interestingly, exercise in both conditions did not significantly increase myocardial nuclear localized phosphorylated HSF1. Nonetheless, warm exercise elevated left-ventricular HSP72 mRNA by ninefold and increased myocardial HSP72 protein levels by threefold compared with cold-exercised animals. Collectively, these data indicate that elevated body temperature and myocardial protein oxidation promoted exercise-induced cardiac HSP72 mRNA expression and protein accumulation following in vivo exercise. However, these results suggest that exercise-induced myocardial HSP72 protein accumulation is not a result of nuclear-localized, phosphorylated HSF1, indicating that other transcriptional or post-transcriptional regulatory mechanisms are involved in exercise-induced HSP72 expression. [source]

A unique lipoylation system in the Archaea

FEBS JOURNAL, Issue 15 2009
Lipoylation in Thermoplasma acidophilum requires two proteins
Members of the 2-oxoacid dehydrogenase multienzyme complex family play a key role in the pathways of central metabolism. Post-translational lipoylation of the dihydrolipoyl acyltransferase component of these complexes is essential for their activity, the lipoyllysine moiety performing the transfer of substrates and intermediates between the different active sites within these multienzyme systems. We have previously shown that the thermophilic archaeon, Thermoplasma acidophilum, has a four-gene cluster encoding the components of such a complex, which, when recombinantly expressed in Escherichia coli, can be assembled into an active multienzyme in vitro. Crucially, the E. coli host carries out the required lipoylation of the archaeal dihydrolipoyl acyltransferase component. Because active 2-oxoacid dehydrogenase multienzyme complexes have never been detected in any archaeon, the question arises as to whether Archaea possess a functional lipoylation system. In this study, we report the cloning and heterologous expression of two genes from Tp. acidophilum whose protein products together show significant sequence identity with the single lipoate protein ligase enzyme of bacteria. We demonstrate that both recombinantly expressed Tp. acidophilum proteins are required for lipoylation of the acyltransferase, and that the two proteins associate together to carry out this post-translational modification. From the published DNA sequences, we suggest the presence of functional transcriptional and translational regulatory elements, and furthermore we present preliminary evidence that lipoylation occurs in vivo in Tp. acidophilum. This is the first report of the lipoylation machinery in the Archaea, which is unique in that the catalytic activity is dependent on two separate gene products. Structured digital abstract ,,MINT-7103712: E2lipD (uniprotkb:Q9HIA5), CTD (uniprotkb:Q9HKT2) and LplA (uniprotkb:Q9HKT1) physically interact (MI:0915) by molecular sieving (MI:0071) [source]

Expression and regulation of alkaline phosphatases in human breast cancer MCF-7 cells

FEBS JOURNAL, Issue 5 2000
Lai-Chen Tsai
The effect of retinoic acid and dexamethasone on alkaline phosphatase (AP) expression was investigated in human breast cancer MCF-7 cells. Cellular AP activity was induced significantly by retinoic acid or dexamethasone in a time-dependent and dose-dependent fashion. A marked synergistic induction of AP activity was observed when the cells were incubated with both agents simultaneously. Two AP isozymes, tissue-nonspecific (TNAP) and intestinal (IAP), were shown to be expressed in MCF-7 cells as confirmed by the differential rate of thermal inactivation of these isozymes and RT-PCR. Based on the two-isozyme thermal-inactivation model, the specific activities for TNAP and IAP in each sample were analyzed. TNAP activity was induced only by retinoic acid and IAP activity was induced only by dexamethasone. Whereas dexamethasone conferred no significant effect on TNAP activity, retinoic acid was shown to inhibit IAP activity by , 50%. Interestingly, TNAP was found to be the only isozyme activity superinduced when the cells were costimulated with retinoic acid and dexamethasone. Northern blot and RT-PCR analysis were then used to demonstrate that the steady-state TNAP mRNA level was also superinduced, which indicates that the superinduction is regulated at the transcriptional or post-transcriptional levels. In the presence of the glucocorticoid receptor antagonist RU486, the dexamethasone-mediated induction of IAP activity was blocked completely as expected. However, the ability of RU486 to antagonize the action of glucocorticoid was greatly compromised in dexamethasone-mediated superinduction of TNAP activity. Furthermore, in the presence of retinoic acid, RU486 behaved as an agonist, and conferred superinduction of TNAP gene expression in the same way as dexamethasone. Taken together, these observations suggest that the induction of IAP activity by dexamethasone and the superinduction of TNAP by dexamethasone were mediated through distinct regulatory pathways. In addition, retinoic acid plays an essential role in the superinduction of TNAP gene expression by enabling dexamethasone to exert its agonist activity, which otherwise has no effect. [source]

Early transcriptional response of Saccharomyces cerevisiae to stress imposed by the herbicide 2,4-dichlorophenoxyacetic acid

FEMS YEAST RESEARCH, Issue 2 2006
Miguel Cacho Teixeira
Abstract The global gene transcription pattern of the eukaryotic experimental model Saccharomyces cerevisiae in response to sudden aggression with the widely used herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) was analysed. Under acute stress, 14% of the yeast transcripts suffered a greater than twofold change. The yeastract database was used to predict the transcription factors mediating the response registered in this microarray analysis. Most of the up-regulated genes in response to 2,4-D are known targets of Msn2p, Msn4p, Yap1p, Pdr1p, Pdr3p, Stp1p, Stp2p and Rpn4p. The major regulator of ribosomal protein genes, Sfp1p, is known to control 60% of the down-regulated genes, in particular many involved in the transcriptional and translational machinery and in cell division. The yeast response to the herbicide includes the increased expression of genes involved in the oxidative stress response, the recovery or degradation of damaged proteins, cell wall remodelling and multiple drug resistance. Although the protective role of TPO1 and PDR5 genes was confirmed, the majority of the responsive genes encoding multidrug resistance do not confer resistance to 2,4-D. The increased expression of genes involved in alternative carbon and nitrogen source metabolism, fatty acid ,-oxidation and autophagy was also registered, suggesting that acute herbicide stress leads to nutrient limitation. [source]

Integrated genomic profiling identifies candidate genes implicated in glioma-genesis and a novel LEO1 - SLC12A1 fusion gene

GENES, CHROMOSOMES AND CANCER, Issue 6 2010
Linda B. C. Bralten
We performed genotyping and exon-level expression profiling on 21 glioblastomas (GBMs) and 19 oligodendrogliomas (ODs) to identify genes involved in glioma initiation and/or progression. Low-copy number amplifications (2.5 < n < 7) and high-copy number amplifications (n > 7) were more frequently observed in GBMs; ODs generally have more heterozygous deletions per tumor. Four high-copy amplicons were identified in more than one sample and resulted in overexpression of the known oncogenes EGFR, MDM2, and CDK4. In the fourth amplicon, RBBP5, a member of the RB pathway, may act as a novel oncogene in GBMs. Not all hCNAs contain known genes, which may suggest that other transcriptional and/or regulatory elements are the target for amplification. Regions with most frequent allelic loss, both in ODs and GBMs, resulted in a reduced expression of known tumor suppressor genes. We identified a homozygous deletion spanning the Pragmin gene in one sample, but direct sequencing of all coding exons in 20 other glioma samples failed to detect additional genetic changes. Finally, we screened for fusion genes by identifying aberrant 5,-3, expression of genes that lie over regions of a copy number change. A fusion gene between exon 11 of LEO1 and exon 10 of SLC12A1 was identified. Our data show that integrated genomic profiling can identify genes involved in tumor initiation, and/or progression and can be used as an approach to identify novel fusion genes. © 2010 Wiley-Liss, Inc. [source]

Inhibition of Rho-dependent pathways by Clostridium botulinum C3 protein induces a proinflammatory profile in microglia

GLIA, Issue 11 2008
Anja Hoffmann
Abstract Successful regeneration in the central nervous system crucially depends on the adequate environment. Microglia as brain immune-competent cells importantly contribute to this task by producing pro- and anti-inflammatory mediators. Any environmental change transforms these cells towards an activated phenotype, leading to major morphological, transcriptional and functional alterations. Rho GTPases affect multiple cellular properties, including the cytoskeleton, and C3 proteins are widely used to study their involvement. Especially C3bot from Clostridium botulinum has been considered to promote neuronal regeneration by changing Rho activity. Yet C3bot may exert cellular influences through alternative mechanisms. To determine the role of Rho-dependent pathways in microglia we investigated the influence of C3bot on functional properties of cultivated primary mouse microglial cells. Nanomolar concentrations of C3bot transformed microglia towards an activated phenotype and triggered the release of nitric oxide and several proinflammatory cyto- and chemokines. These inductions were not mediated by the ROCK-kinase pathway, since its selective inhibitors Y27632 and H1152 had no effect. C3-induced and Rho-mediated NO release was instead found to be under the control of NF,B, as revealed by treatment with the NF,B inhibitor PDTC. Thus, C3bot induces a proinflammatory response in microglia resembling the classical proinflammatory phenotype elicited by bacterial LPS. The findings are relevant for the use of C3bot in regenerative approaches. © 2008 Wiley-Liss, Inc. [source]

Kupffer cell,derived interleukin 10 is responsible for impaired bacterial clearance in bile duct,ligated mice

HEPATOLOGY, Issue 2 2004
Tetsuya Abe
Extrahepatic cholestasis often evokes liver injury with hepatocyte apoptosis, aberrant cytokine production, and,most importantly,postoperative septic complications. To clarify the involvement of aberrant cytokine production and hepatocyte apoptosis in impaired resistance to bacterial infection in obstructive cholestasis, C57BL/6 mice or Fas-mutated lpr mice were inoculated intraperitoneally with 107 colony-forming units of Escherichia coli 5 days after bile duct ligation (BDL) or sham celiotomy. Cytokine levels in sera, liver, and immune cells were assessed via enzyme-linked immunosorbent assay or real-time reverse-transcriptase polymerase chain reaction. BDL mice showed delayed clearance of E. coli in peritoneal cavity, liver, and spleen. Significantly higher levels of serum interleukin (IL) 10 with lower levels of IL-12p40 were observed in BDL mice following E. coli infection. Interferon , production from liver lymphocytes in BDL mice was not increased after E. coli infection either at the transcriptional or protein level. Kupffer cells from BDL mice produced low levels of IL-12p40 and high levels of IL-10 in vitro in response to lipopolysaccharide derived from E. coli. In vivo administration of anti,IL-10 monoclonal antibody ameliorated the course of E. coli infection in BDL mice. Furthermore, BDL- lpr mice did not exhibit impairment in E. coli killing in association with little hepatic injury and a small amount of IL-10 production. In conclusion, increased IL-10 and reciprocally suppressed IL-12 production by Kupffer cells are responsible for deteriorated resistance to bacterial infection in BDL mice. Fas-mediated hepatocyte apoptosis in cholestasis may be involved in the predominant IL-10 production by Kupffer cells. (HEPATOLOGY 2004;40:414,423.) [source]

Synaptophysin protein and mRNA expression in the human hippocampal formation from birth to old age,

HIPPOCAMPUS, Issue 8 2006
Sharon L. Eastwood
Abstract In the human neocortex, progressive synaptogenesis in early postnatal life is followed by a decline in synaptic density, then stability from adolescence until middle age. No comparable data are available in the hippocampus. In this study, the integral synaptic vesicle protein synaptophysin, measured immunoautoradiographically, was used as an index of synaptic terminal abundance in the hippocampal formation of 37 subjects from 5 weeks to 86 yr old, divided into 4 age groups (10 infants, 15 adolescents/young adults, 6 adults, and 6 elderly). In all hippocampal subfields, synaptophysin was lowest in infancy, but did not differ significantly between the older age groups, except in dentate gyrus (DG) where the rise was delayed until adulthood. A similar developmental profile was found in the rat hippocampus. We also measured synaptophysin mRNA in the human subjects and found no age-related changes, except in parahippocampal gyrus wherein the mRNA declined from infancy to adolescence, and again in old age. The synaptophysin protein data demonstrate a significant presynaptic component to human postnatal hippocampal development. In so far as synaptophysin abundance reflects synaptic density, the findings support an increase in hippocampal and parahippocampal synapse formation during early childhood, but provide no evidence for adolescent synaptic pruning. The mRNA data indicate that the maturational increases in synaptophysin protein are either translational rather than transcriptional in origin, or else are secondary to mRNA increases in neurons, the cell bodies of which lie outside the hippocampal formation. Published 2006 Wiley-Liss, Inc. [source]

UbcH10 expression in human lymphomas

HISTOPATHOLOGY, Issue 6 2009
Giancarlo Troncone
Aims:, The UbcH10 ubiquitin-conjugating enzyme plays a key role in regulating mitosis completion. We have previously reported that UbcH10 overexpression is associated with aggressive thyroid, ovarian and breast carcinomas. The aim of this study was to investigate UbcH10 expression in human lymphomas. Methods and results:, Cell lines and tissue samples of Hodgkin's lymphoma (HL) and of non-Hodgkin's lymphoma (NHL) were screened for UbcH10 expression at transcriptional and translational levels. UbcH10 expression was related to the grade of malignancy. In fact, it was low in indolent tumours and high in a variety of HL and NHL cell lines and in aggressive lymphomas. It was highest in Burkitt's lymphoma, as shown by quantitative real-time polymerase chain reaction and by tissue microarray immunohistochemistry. Flow cytometry of cell lines confirmed that UbcH10 expression is cell-cycle dependent, steadily increasing in S phase, peaking in G2/M phase and dramatically decreasing in G0/G1 phases. We also showed that UbcH10 plays a relevant role in lymphoid cell proliferation, since blocking of its synthesis by RNA interference inhibited cell growth. Conclusions:, Taken together, these results indicate that UbcH10 is a novel lymphoid proliferation marker encompassing the cell cycle window associated with exit from mitosis. Its overexpression in aggressive lymphomas suggests that UbcH10 could be a therapeutic target in this setting. [source]

Expression of 5-lipoxygenase (5-LOX) in T lymphocytes

IMMUNOLOGY, Issue 2 2007
Jeanne M. Cook-Moreau
Summary 5-lipoxygenase (5-LOX) is the key enzyme responsible for the synthesis of the biologically active leukotrienes. Its presence has been reported in cells of the myeloid lineage and B lymphocytes but has not been formally defined in T lymphocytes. In this study, we provide evidence for 5-LOX expression on both transcriptional and translational levels in highly purified peripheral blood T cells as well as in human T lymphoblastoid cell lines (MOLT4 and Jurkat). Messenger RNA (mRNA) of 5-LOX was amplified by conventional reverse transcription,polymerase chain reaction (RT-PCR; MOLT4 and Jurkat cells) and by in situ RT-PCR (T lymphocytes). 5-LOX protein expression was confirmed by Western blot and immunofluorescence studies. 5-LOX was present primarily in the cytoplasm with some nuclear localization and was translocated to the nuclear periphery after culture in a mitosis-supporting medium. Fluorescence-activated cell sorter analysis of different T-lymphocyte populations, including CD4, CD8, CD45RO, CD45RA, T helper type 2, and T-cell receptor-,, and -,, expressing cells, did not identify a differential distribution of the enzyme. Purified peripheral blood T lymphocytes were incapable of synthesizing leukotrienes in the absence of exogenous arachidonic acid. Jurkat cells produced leukotriene C4 and a small amount of leukotriene B4 in response to CD3,CD28 cross-linking. This synthesis was abolished by two inhibitors of leukotriene synthesis, MK-886 and AA-861. The presence of 5-LOX in T lymphocytes but the absence of endogenous lipoxygenase metabolite production compared to Jurkat cells may constitute a fundamental difference between resting peripheral lymphocytes and leukaemic cells. [source]

Relevance of a new rat model of osteoblastic metastases from prostate carcinoma for preclinical studies using zoledronic acid

INTERNATIONAL JOURNAL OF CANCER, Issue 4 2008
François Lamoureux
Abstract Animal models that mimic osteoblastic metastases associated with prostate carcinoma are required to improve the therapeutic options in humans. A new model was then developed and characterized in immunocompetent rats. The bisphosphonate zoledronic acid (ZOL) was tested to validate this model as a therapeutic application. Rat AT6-1 prostate tumor cells were characterized in vitro at the transcriptional (bone and epithelial markers) and functional (induction of mineralized nodules) levels. The bone lesions induced after their direct injection into the femur bone marrow were characterized by radiography, microscanner and histology analyses. ZOL effects were studied in vivo on bone lesion development and in vitro on AT6-1 cell proliferation, apoptosis and cell cycle analysis. Apart from epithelial markers, AT6-1 cells express an osteoblast phenotype as they express osteoblastic markers and are able to induce mineralized nodule formation in vitro. A disorganization of the trabecular bone at the growth zone level was observed in vivo after intraosseous AT6-1 cell injection as well as cortical erosion. The tumor itself is associated with bone formation as revealed by SEM analysis and polarized light microscopy. ZOL prevents the development of such osteoblastic lesions, related to a direct inhibitory effect on tumor cell proliferation independent of caspase 3 activation, but associated with cell cycle arrest. A new rat model of osteoblastic bone metastases was validated in immunocompetent rats and used to show the relevance of using ZOL in such lesions, as this compound shows bifunctional effects on both bone remodelling and tumor cell proliferation. © 2007 Wiley-Liss, Inc. [source]

Update on peptidylarginine deiminases and deimination in skin physiology and severe human diseases

INTERNATIONAL JOURNAL OF COSMETIC SCIENCE, Issue 3 2007
M.-C. Méchin
Synopsis Deimination (or citrullination) is a recently described post-translational modification, but its consequences are not yet well understood. It is catalysed by peptidylarginine deiminases (PADs). These enzymes transform arginyl residues involved in a peptidyl link into citrullyl residues in a calcium-dependent manner. Several PAD substrates have already been identified like filaggrin and keratins K1 and K10 in the epidermis, trichohyalin in hair follicles, but also ubiquitous proteins like histones. PADs act in a large panel of physiological functions as cellular differentiation or gene regulation. It has been suggested that deimination plays a role in many major diseases such as rheumatoid arthritis, multiple sclerosis, Alzheimer's disease and psoriasis. Five human genes (PADIs), encoding five highly conserved paralogous enzymes (PAD1-4 and 6), have been characterized. These genes are clustered in a single locus, at 1p35-36 in man. Only PAD1-3 are expressed in human epidermis. PADs seem to be controlled at transcriptional, translational and activity levels and they present particular substrate specificities. In this review, we shall discuss these main biochemical, genetic and functional aspects of PADs together with their pathophysiological implications. Résumé La désimination (ou citrullination) est une modification post-traductionnelle catalysée par les peptidyl-arginine désiminases (PADs), décrite depuis peu et dont les conséquences sont encore mal comprises. Ces enzymes transforment, de façon dépendante du calcium, les résidus arginyl engagés dans un lien peptidique en résidus citrullyl. Plusieurs substrats ont été identifiés: la filaggrine et les cytokératines K1 et K10 de l'épiderme, la trichohyaline dans le follicule pileux mais aussi des protéines ubiquistes comme les histones. Les PADs interviennent dans de nombreuses fonctions physiologiques telles que la différenciation cellulaire ou la régulation génique. La désimination pourrait jouer un rôle dans plusieurs maladies sévères et fréquentes comme la polyarthrite rhumatoïde, la sclérose en plaque, la maladie d'Alzheimer ou encore le psoriasis. Cinq gènes humains (PADIs) codant pour 5 enzymes paralogues conservées (PAD1-4 et 6) ont été caractérisés. Ils sont regroupés en un seul locus, en 1p35-36 chez l'homme. Seules les PAD1-3 sont exprimées dans l'épiderme humain. Les PADs semblent contrôlées aux niveaux transcriptionnel et traductionnel, ainsi qu'au niveau de leur activité. Elles présentent chacune leurs spécificités de substrats. Ces principaux aspects biochimiques, génétiques et fonctionnels des PADs tout comme leurs implications physiopathologiques seront discutés dans cette revue. [source]

The adenine nucleotide translocase type 1 (ANT1): A new factor in mitochondrial disease

IUBMB LIFE, Issue 9 2005
J. Daniel Sharer
Abstract Mitochondrial disorders of oxidative phosphorylation (OXPHOS) comprise a growing list of potentially lethal diseases caused by mutations in either mitochondrial (mtDNA) or nuclear DNA (nDNA). Two such conditions, autosomal dominant progressive external ophthalmoplegia (adPEO) and Senger's Syndrome, are associated with dysfunction of the heart and muscle-specific isoform of the adenine nucleotide translocase (ANT1), a nDNA gene product that facilitates transport of ATP and ADP across the inner mitochondrial membrane. AdPEO is a mtDNA deletion disorder broadly characterized by pathology involving the eyes, skeletal muscle, and central nervous system. In addition to ANT1, mutations in at least two other nuclear genes, twinkle and POLG, have been shown to cause mtDNA destabilization associated with adPEO. Senger's syndrome is an autosomal recessive condition characterized by congenital heart defects, abnormalities of skeletal muscle mitochondria, cataracts, and elevated circulatory levels of lactic acid. This syndrome is associated with severe depletion of ANT1, which may be the result of an as yet unidentified ANT1-specific transcriptional or translational processing error. ANT1 has also been associated with a third condition, autosomal dominant facioscapulohumeral muscular dystrophy (FSHD), an adult onset disorder characterized by variable muscle weakness in the face, feet, shoulders, and hips. FSHD patients possess specific DNA deletions on chromosome 4, which appear to cause derepression of several nearby genes, including ANT1. Early development of FSHD may involve mitochondrial dysfunction and increased oxidative stress, possibly associated with overexpression of ANT1. IUBMB Life, 57: 607-614, 2005 [source]

Transcriptional and post-transcriptional control of DNA methyltransferase 3B is regulated by phosphatidylinositol 3 kinase/Akt pathway in human hepatocellular carcinoma cell lines

Defining the transcriptome of accelerated and replicatively senescent keratinocytes reveals links to differentiation, interferon signaling, and Notch related pathways,

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2006
Ranjan J. Perera
Abstract Epidermal keratinocytes (KCs) undergo highly orchestrated morphological and molecular changes during transition from proliferative compartment into growth arrested early and late differentiation layers, prior to dying in outermost cornified layers of normal skin. Creation of stratum corneum is vital to barrier function protecting against infection. Transcriptional events in KCs regulating complex processes of differentiation and host defense required to maintain constant epidermal thickness and resistance to infection in either young or aged skin are largely unknown. Furthermore, as terminal differentiation is characterized by irreversible loss of replicative potential culminating in dead layers at the skin surface, this process may be viewed as a form of senescence. However, a complete transcriptional profile of senescent (SN) human KCs has not been previously defined to permit delineation of molecular boundaries involving differentiation and senescence. To fill this void, we utilized global transcriptional analysis of KCs maintained in vitro as either cultures of proliferating (PR) cells, early and late confluent (LC) (accelerated senescence) cultures, or KCs undergoing replicative senescence. Global gene expression profiling revealed early confluent (EC) KCs were somewhat similar to PR KCs, while prominent differences were evident when compared to LC KCs; which were also distinct from replicatively SN KCs. While confluent KCs have in common several genes regulating differentiation with replicatively SN KCs, the latter cells expressed elevated levels of genes involved in interferon signaling and inflammatory pathways. These results provide new insights into cell autonomous transcriptional-based programs operative within KCs contributing to replicative senescence, with partial sharing of genes involved in differentiation. In addition, regulation of KC senescence may involve participation of interferon signaling pathways derived from the important role of KCs in protecting skin from infection. Integrating all of the transcriptional data revealed a key role for Notch receptor mediated signaling in the confluency induced differentiation phenotype using this model system. J. Cell. Biochem. 98: 394,408, 2006. © 2006 Wiley-Liss, Inc. [source]

Transcriptional regulation of collagenase-3 by interleukin-1 alpha in osteoblasts

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2003
Samuel Varghese
Abstract Interleukin-1 (IL-1), is an autocrine/paracrine agent of the skeletal tissue and it regulates bone remodeling. Collagenase-3 or matrix metalloproteinase (MMP)-13 is expressed in osteoblasts and its expression is modulated by several cytokines including IL-1,. Because the molecular mechanism of increased synthesis of collagenase-3 in bone cells by IL-1, is not known, we investigated if collagenase-3 expression by IL-1, in osteoblasts is mediated by transcriptional or post-transcriptional mechanisms. Exposure of rat osteoblastic cultures (Ob cells) to IL-1, at concentrations higher than 0.5 nM increased the synthesis of collagenase-3 mRNA up to eightfold and the secretion of immunoreactive protein up to 21-fold. The effects of IL-1, on collagenase-3 were time- and dose-dependent. Although prostaglandins stimulate collagenase-3 expression, stimulation of collagenase-3 in Ob cells by IL-1, was not mediated through increased biosynthesis of prostaglandins. The half-life of collagenase-3 mRNA from control and IL-1,-treated Ob cells was similar suggesting that the stabilization of collagenase-3 mRNA did not contribute to the increase in collagenase-3. However, IL-1, stimulated the rate of transcription of the collagenase-3 gene by twofold to fourfold indicating regulation of collagenase-3 expression in Ob cells at the transcriptional level. Stimulation of collagenase-3 by IL-1, in osteoblasts may in part mediate the effects of IL-1, in bone metabolism. © 2003 Wiley-Liss, Inc. [source]

Characterization of the upstream mouse Cbfa1/Runx2 promoter,

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2001
Z. S. Xiao
Abstract Cbfa1 (or Runx2/AML-3/PEPB2,) is a transcriptional activator of osteoblastic differentiation. To investigate the regulation of Cbfa1 expression, we isolated and characterized a portion of the 5,-flanking region of the Cbfa1 gene containing its "bone-related" or P1 promoter and exon 1. We identified additional coding sequence in exon 1 and splice donor sites that potentially give rise to a novel Cbfa1 isoform containing an 18 amino acid insert. In addition, primer extension mapping identified in the Cbfa1 promoter a minor mRNA start site located ,0.8 kb 5, upstream of the ATG encoding the MASN/p57 isoform and ,0.4 kb upstream of the previously reported start site. A luciferase reporter construct containing 1.4 kb of the mouse Cbfa1 promoter was analyzed in Ros 17/2.8 and MC3T3-E1 osteoblast cell lines that express high levels of Cbfa1 transcripts. The activity of this construct was also examined in non-osteoblastic Cos-7 and NIH3T3 cells that do not express Cbfa1 and mesenchymal-derived cell lines, including CH3T101/2, C2C12, and L929 cells, that express low levels of mature Cbfa1 transcripts. The 1.4 kb 5, flanking sequence of the Cbfa1 gene directed high levels of transcriptional activity in Ros 17/2.8 and MC3T3-E1 osteoblasts compared to non-osteoblasts Cos-7 cells, but this construct also exhibited high levels of expression in C310T1/2, L929, and C2C12 cells as well as NIH3T3 cells. In addition, Cbfa1 mRNA expression, but not the activity of the Cbfa1 promoter, was upregulated in a dose-dependent manner in pluripotent mesenchymal C2C12 by bone morphogenetic protein-2 (BMP-2). These data indicate that Cbfa1 is expressed in osteogenic as well as non-osteogenic cells and that the regulation of Cbfa1 expression is complex, possibly involving both transcriptional and post-transcriptional mechanisms. Additional studies are needed to further characterize important regulatory elements and to identify additional regions of the promoter and/or post-transcriptional events responsible for the cell-type restricted regulation of Cbfa1 expression. J. Cell. Biochem. 82: 647,659, 2001. © 2001 Wiley-Liss, Inc. [source]

Restoration of DNA-binding and growth-suppressive activity of mutant forms of p53 via a PCAF-mediated acetylation pathway,

JOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2010
Ricardo E. Perez
Tumor-derived mutant forms of p53 compromise its DNA binding, transcriptional, and growth regulatory activity in a manner that is dependent upon the cell-type and the type of mutation. Given the high frequency of p53 mutations in human tumors, reactivation of the p53 pathway has been widely proposed as beneficial for cancer therapy. In support of this possibility p53 mutants possess a certain degree of conformational flexibility that allows for re-induction of function by a number of structurally different artificial compounds or by short peptides. This raises the question of whether physiological pathways for p53 mutant reactivation also exist and can be exploited therapeutically. The activity of wild-type p53 is modulated by various acetyl-transferases and deacetylases, but whether acetylation influences signaling by p53 mutant is still unknown. Here, we show that the PCAF acetyl-transferase is down-regulated in tumors harboring p53 mutants, where its re-expression leads to p53 acetylation and to cell death. Furthermore, acetylation restores the DNA-binding ability of p53 mutants in vitro and expression of PCAF, or treatment with deacetylase inhibitors, promotes their binding to p53-regulated promoters and transcriptional activity in vivo. These data suggest that PCAF-mediated acetylation rescues activity of at least a set of p53 mutations. Therefore, we propose that dis-regulation of PCAF activity is a pre-requisite for p53 mutant loss of function and for the oncogenic potential acquired by neoplastic cells expressing these proteins. Our findings offer a new rationale for therapeutic targeting of PCAF activity in tumors harboring oncogenic versions of p53. J. Cell. Physiol. 225: 394,405, 2010. © 2010 Wiley-Liss, Inc. [source]

Transactivation of Src, PDGF receptor, and Akt is involved in IL-1,-induced ICAM-1 expression in A549 cells

JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007
Chih-Chung Lin
In previous study, interleukin-1, (IL-1,) has been shown to induce ICAM-1 expression through MAPKs and NF-,B in A549 cells. In addition to these pathways, transactivation of non-receptor tyrosine kinase (Src), PDGF receptors (PDGFRs), and phosphatidylinositol 3-kinase (PI3K)/Akt has been implicated in the expression of inflammatory genes. Here, we further investigated whether these different mechanisms participating in IL-1,-induced ICAM-1 expression in A549 cells. We initially observed that IL-1,-induced ICAM-1 promoter activity was attenuated by the inhibitors of Src (PP1), PDGFR (AG1296), PI3-K (LY294002 and wortmannin), and Akt (SH-5), revealed by reporter gene assay, Western blotting, and RT-PCR analyses. The involvement of Src and PI3-K/Akt in IL-1,-induced ICAM-1 expression was significantly attenuated by transfection of A549 cells with dominant negative plasmids of Src, p85 and Akt, respectively. Src, PDGFR, and PI3K/Akt mediated the effects of IL-1, because pretreatment with PP1, AG1296, and wortmannin also abrogated IL-1,-stimulated Src, PDGFR, and Akt phosphorylation, respectively. Moreover, pretreatment with p300 inhibitor (curcumin) also blocked ICAM-1 expression. We further confirmed that p300 was associated with ICAM-1 promoter which was dynamically linked to histone H4 acetylation stimulated by IL-1,, determined by chromatin immunoprecipitation assay. Association of p300 and histone-H4 to ICAM-1 promoter was inhibited by LY294002. Up-regulation of ICAM-1 enhanced the adhesion of neutrophils onto A549 cell monolayer exposed to IL-1,, which was inhibited by PP1, AG1296, LY294002, wortmannin, and helenalin. These results suggested that Akt phosphorylation mediated through transactivation of Src/PDGFR promotes the transcriptional p300 activity and eventually leads to ICAM-1 expression induced by IL-1,. J. Cell. Physiol. 211: 771,780, 2007. © 2007 Wiley-Liss, Inc. [source]