Glucocorticoids inhibit IL-1β-induced GM-CSF expression at multiple levels: roles for the ERK pathway and repression by MKP-1

2010 ◽  
Vol 427 (1) ◽  
pp. 113-124 ◽  
Author(s):  
Robert Newton ◽  
Elizabeth M. King ◽  
Wei Gong ◽  
Christopher F. Rider ◽  
Karl J. Staples ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Transcriptional Repression ◽  
Actinomycin D ◽  
De Novo ◽  
A549 Cells ◽  
Mitogen Activated Protein Kinase ◽  
Erk Pathway ◽  
Gm Csf ◽  
Erk Phosphorylation

In the present study, IL (interleukin)-1β increased GM-CSF (granulocyte/macrophage colony-stimulating factor) expression from pulmonary A549 cells and primary HBE (human bronchial epithelial) cells. These responses were repressed by the glucocorticoid dexamethasone, allowing the use of A549 cells as a relevant model. IL-1β induced GM-CSF release into the culture medium by 6 h and in cell lysates (cytosolic) at 2 h. These effects were profoundly inhibited by dexamethasone, yet IL-1β-induced GM-CSF mRNA and unspliced nRNA (nuclear RNA; a surrogate of transcription rate) were modestly inhibited by dexamethasone at times up to 2 h. Although this indicates an effect on protein synthesis, actinomycin D chase experiments also indicated post-transcriptional repression by dexamethasone. Dexamethasone-dependent mRNA repression increased with time and was prevented by translational blockade. In addition, dexamethasone and the dissociated steroid RU24858 repressed GM-CSF release in an actinomycin D-sensitive manner, thereby implicating glucocorticoid-induced gene expression. At 2 h, IL-1β-induced expression of GM-CSF protein, but not mRNA, was sensitive to the MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase] inhibitors PD098059 and U0126. Although this indicates a role for the MEK/ERK pathway in GM-CSF translation, PD098059 subsequently destabilized GM-CSF mRNA. Dexamethasone and RU24858 both reduced IL-1β-induced ERK phosphorylation and increased MKP-1 (MAPK phosphatase-1) expression. Inhibition of ERK phosphorylation was reproduced by MKP-1 overexpression and prevented by MKP-1-targeting siRNA (small interfering RNA). Since MKP-1 prevented GM-CSF expression by transcriptional, post-transcriptional and translational processes, we propose that glucocorticoids induce MKP-1 expression to reduce both MEK/ERK activation and GM-CSF protein synthesis. Thus de novo gene expression, particularly of MKP-1, is involved in the repressive effects of glucocorticoids.

scholarly journals Protein-synthesis-dependent induction of annexin I by glucocorticoid

1991 ◽  
Vol 275 (2) ◽  
pp. 313-319 ◽  
Author(s):  
W T Wong ◽  
S C Frost ◽  
H S Nick
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Time Course ◽  
Actinomycin D ◽  
De Novo ◽  
Lung Epithelial Cells ◽  
Annexin I ◽  
Lung Epithelial ◽  
Maximal Induction ◽  
I Gene

We demonstrate that annexin I/lipocortin I (lipo I) gene expression is regulated by dexamethasone (DEX) in mouse 3T3-L1 fibroblasts and LA-4 lung epithelial cells. We have characterized this induction further in 3T3-L1 fibroblasts. At 24 h after addition of DEX, the levels of lipo I mRNA and protein increased 5-fold and 1.5-fold respectively. Time-course experiments revealed that the induction was delayed by 2-4 h after DEX addition. Half-maximal induction of both lipo I mRNA and protein was achieved with 10 nM-DEX. Both actinomycin D and cycloheximide blocked the DEX effect on lipo I mRNA expression. These results indicate that the induction of lipo I by DEX has a transcriptional component and requires protein synthesis de novo.

scholarly journals Viral Gene Expression Patterns in Human Herpesvirus 6B-Infected T Cells

2002 ◽  
Vol 76 (15) ◽  
pp. 7578-7586 ◽  
Author(s):  
Bodil Øster ◽  
Per Höllsberg
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Protein Synthesis ◽  
De Novo ◽  
Expression Patterns ◽  
Human Herpesvirus ◽  
Viral Gene Expression ◽  
Polymerase Activity ◽  
Viral Gene ◽  
De Novo Protein Synthesis

ABSTRACT Herpesvirus gene expression is divided into immediate-early (IE) or α genes, early (E) or β genes, and late (L) or γ genes on the basis of temporal expression and dependency on other gene products. By using real-time PCR, we have investigated the expression of 35 human herpesvirus 6B (HHV-6B) genes in T cells infected by strain PL-1. Kinetic analysis and dependency on de novo protein synthesis and viral DNA polymerase activity suggest that the HHV-6B genes segregate into six separate kinetic groups. The genes expressed early (groups I and II) and late (groups V and VI) corresponded well with IE and L genes, whereas the intermediate groups III and IV contained E and L genes. Although HHV-6B has characteristics similar to those of other roseoloviruses in its overall gene regulation, we detected three B-variant-specific IE genes. Moreover, genes that were independent of de novo protein synthesis clustered in an area of the viral genome that has the lowest identity to the HHV-6A variant. The organization of IE genes in an area of the genome that differs from that of HHV-6A underscores the distinct differences between HHV-6B and HHV-6A and may provide a basis for further molecular and immunological analyses to elucidate their different biological behaviors.

scholarly journals The Histone Chaperone Anti-Silencing Function 1 Is a Global Regulator of Transcription Independent of Passage through S Phase

2005 ◽  
Vol 25 (2) ◽  
pp. 652-660 ◽  
Author(s):  
Susan R. Zabaronick ◽  
Jessica K. Tyler
Keyword(s):  
Gene Expression ◽  
Transcriptional Regulation ◽  
Transcriptional Repression ◽  
De Novo ◽  
S Phase ◽  
Global Changes ◽  
Histone Chaperones ◽  
Direct Role ◽  
Broad Array ◽  
Assembly Factor

ABSTRACT We investigated the function of the histone H3/H4 chaperones anti-silencing function 1 (Asf1p) and chromatin assembly factor 1 (CAF-1) in global transcriptional regulation in budding yeast. Deletion of ASF1 or CAF-1 components led to global transcriptional misregulation, both activation and repression, of genes scattered throughout the 16 yeast chromosomes. To investigate direct effects on gene regulation, we developed an approach to destabilize Asf1p that results in its rapid degradation within minutes of transcriptional repression. Upon degradation of Asf1p, rapid global changes in gene expression occur without the requirement for passage through S phase or de novo protein synthesis. In particular, we demonstrate that the previously reported influence of Asf1p on histone gene expression is not a direct effect of loss of Asf1p. These data indicate that the histone chaperones CAF-1 and Asf1p regulate the gene expression of a broad array of genes in yeast and, in the case of Asf1p, this is likely to be due to a direct role in chromatin modulation during transcriptional regulation.

scholarly journals Ghrelin Increases Neuropeptide Y and Agouti-Related Peptide Gene Expression in the Arcuate Nucleus in Rat Hypothalamic Organotypic Cultures

Endocrinology ◽  
2006 ◽  
Vol 147 (11) ◽  
pp. 5102-5109 ◽  
Author(s):  
Motomitsu Goto ◽  
Hiroshi Arima ◽  
Minemori Watanabe ◽  
Masayuki Hayashi ◽  
Ryouichi Banno ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Neuropeptide Y ◽  
Mrna Expression ◽  
Gene Transcription ◽  
Arcuate Nucleus ◽  
De Novo ◽  
Organotypic Cultures ◽  
Related Peptide ◽  
De Novo Protein Synthesis

Ghrelin, which was identified from the rat stomach, is a potent stimulant for food intake. Several lines of evidence suggest that the orexigenic action of ghrelin is mediated via the neuropeptide Y (NPY) neurons in the arcuate nucleus, although the detailed mechanisms by which ghrelin stimulates NPY neurons are not clear. In this study, we examined the gene regulation of NPY and agouti-related peptide (AGRP), another orexigenic peptide synthesized in the NPY neurons, in the arcuate nucleus by ghrelin in hypothalamic organotypic cultures. Incubation of the hypothalamic explants with ghrelin significantly increased NPY and AGRP mRNA expression in the presence, but not absence, of dexamethasone. Glucocorticoids were also necessary for ghrelin action in vivo because an intracerebroventricular injection of ghrelin significantly increased NPY and AGRP mRNA expression in the arcuate nucleus only in sham-operated, but not in adrenalectomized rats. The stimulatory effects of ghrelin on gene expression were not blocked by a sodium channel blocker tetrodotoxin in the organotypic cultures. Ghrelin also increased NPY heteronuclear (hn) RNA expression, the first transcript that has been used as an indicator for gene transcription. The stimulatory effects of ghrelin on NPY gene expression were abolished in the presence of cycloheximide, which blocks translation, suggesting that de novo protein synthesis is required for ghrelin action. These data suggest that ghrelin stimulates NPY and AGRP gene expression independently of action potentials only in the presence of glucocorticoids. Furthermore, our data demonstrate stimulatory action of ghrelin on NPY gene transcription, which requires de novo protein synthesis.

Transcriptional and posttranscriptional regulation of CSF-1 gene expression in human monocytes

1988 ◽  
Vol 8 (9) ◽  
pp. 3951-3954
Author(s):  
J Horiguchi ◽  
E Sariban ◽  
D Kufe
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Gene Transcription ◽  
Posttranscriptional Regulation ◽  
Drug Exposure ◽  
Actinomycin D ◽  
Protein Synthesis Inhibitor ◽  
Mrna Levels ◽  
Human Monocytes ◽  
Synthesis Inhibitor

Regulation of CSF-1 gene expression was investigated in human monocytes. CSF-1 transcripts were at low or undetectable levels in resting monocytes. However, in monocytes treated with 12-O-tetradecanoylphorbol-13-acetate (TPA), CSF-1 mRNA was increased by 3 h and reached maximal levels by 12 h of drug exposure. When nuclear run-on assays were used, CSF-1 gene transcription was also at low or undetectable levels in resting monocytes but was activated after TPA exposure. TPA-treated monocytes exposed to actinomycin D further demonstrated that the half-life of the CSF-1 mRNA is 0.9 h. The results also demonstrated that the protein synthesis inhibitor, cycloheximide (CHX), increases CSF-1 mRNA levels in both resting and TPA-treated monocytes. These effects of CHX occurred in the absence of detectable increases in CSF-1 gene transcription. Moreover, treatment of monocytes with CHX and actinomycin D demonstrated that inhibition of protein synthesis is associated with stabilization of the CSF-1 transcript. Taken together, these findings indicated that CSF-1 gene expression is controlled at both transcriptional and posttranscriptional levels in human monocytes.

Exosomal MicroRNA-328 from Bone Marrow Mesenchymal Stem Cells (BMSCs) Alleviates Acute Lung Injury Through Mitogen-Activated Protein Kinase/Extracellular Signal-Regulated Kinase (MAPK/ERK) Pathway

2022 ◽  
Vol 12 (2) ◽  
pp. 358-364
Author(s):  
Wei Zhang ◽  
Fang Liu ◽  
Caixia Zhang
Keyword(s):  
Cell Proliferation ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Cell Damage ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Mitogen Activated Protein Kinase ◽  
Erk Pathway ◽  
Mitogen Activated Protein ◽  
Induced Apoptosis

To elucidate the communication between exosomes (exo) derived from BMSCs and injured lung cells. BMSC-exo was isolated and characterized. Lung epithelial cells A549 were incubated with BMSC-exo, and treated by LPS to induce cell damage. CCK-8 assay was carried out to test cell proliferation, flow cytometry was adopted to analyze cell apoptosis, and RT-qPCR as well as Western blot analysis were selected to assess expression of apoptosis- and anti-apoptosis related proteins. Functional experiment was performed to identify the role of microRNA (miRNA)-328 in lung injury. LPS treatment significantly inhibited the viability of A549 cells, induced apoptosis of A549 cells by increasing Bax and casepase-3 levels and reducing Bcl-2 expression, whilst declined expression of miR-328 and suppressed the phosphorylation activation of the MAPK/ERK pathway. Meanwhile, the amount of IL-6, IL-1β and TNF-α were elevated in injured cells, but, the presence of BMSC-exo eliminated the elevation of the contents. Importantly, treatment with BMSC-exo increased miR-328 expression, activated MAPK MAPK/ERK pathway, inhibited apoptosis, and enhanced cell proliferation. However, the effect of BMSC-exo was attenuated when the cells were silenced for miR-328 expression. Collectively, BMSC-exo enriched miR-328 could relieve acute lung injury through MAPK/ERK pathway.

p38 MAPK activation by TGF-β1 increases MLC phosphorylation and endothelial monolayer permeability

2002 ◽  
Vol 282 (1) ◽  
pp. L146-L154 ◽  
Author(s):  
Peter L. Goldberg ◽  
Darren E. MacNaughton ◽  
Richard T. Clements ◽  
Fred L. Minnear ◽  
Peter A. Vincent
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
P38 Mapk ◽  
Transforming Growth Factor ◽  
Control Level ◽  
Mitogen Activated Protein Kinase ◽  
Endothelial Monolayer ◽  
Mapk Activation ◽  
Monolayer Permeability ◽  
Tgf Β1

Transforming growth factor (TGF)-β1 increases endothelial monolayer permeability and myosin light chain phosphorylation (MLC-P) beginning 1–2 h posttreatment, suggesting that changes in gene expression may be required for these responses. The role of extracellular signal-regulated kinase (ERK) 1/2 and p38 mitogen-activated protein kinase (p38 MAPK) was investigated because both kinases have been implicated in regulating gene expression after TGF-β1. ERK1/2 phosphorylation increased threefold above the control level, and the increase was temporally associated with the increase in MLC-P. Inhibition of ERK1/2 phosphorylation with the MAPK kinase inhibitor U-0126 did not prevent the increase in either monolayer permeability or MLC-P. p38 MAPK phosphorylation increased fourfold above the control level, but unlike ERK1/2, this increase peaked 30 min and 1 h post-TGF-β1 treatment. Inhibition of p38 MAPK activity with SB-203580 prevented the increases in both monolayer permeability and MLC-P. Treatment of the monolayers with cycloheximide in conjunction with TGF-β1-inhibited MLC-P, showing a requirement for protein synthesis. These studies demonstrate that p38 MAPK activation and subsequent protein synthesis are part of the signal transduction pathway leading to the TGF-β1-induced increases in monolayer permeability and MLC-P.

Studies on the nature of the seromucoid and haptoglobin responses to experimental inflammation

1968 ◽  
Vol 46 (5) ◽  
pp. 477-481 ◽  
Author(s):  
M. Maung ◽  
D. G. Baker ◽  
R. K. Murray
Keyword(s):  
Protein Synthesis ◽  
Actinomycin D ◽  
De Novo ◽  
De Novo Synthesis ◽  
Experimental Inflammation ◽  
Haptoglobin Level

The effects of the administration of actinomycin D, ethionine, and puromycin on the elevations of the total seromucoid fraction and of one of its components (haptoglobin) occurring during experimental inflammation have been studied. All three inhibitors of protein synthesis abolished the elevation of haptoglobin level. Ethionine and puromycin also completely suppressed the elevation of total seromucoid level, whereas actinomycin D only partially suppressed it. The seromucoid and haptoglobin levels in control animals injected with only the inhibitors of protein synthesis were not in general significantly different from those of the animals injected with turpentine and these agents. The results are consistent with the concept that the elevation of various plasma glycoproteins occurring during inflammation is principally due to de novo synthesis of these proteins rather than release of preformed proteins from tissue pools.

scholarly journals Cell-shape-dependent modulation of p52(PAI-1) gene expression involves a secondary response pathway

1995 ◽  
Vol 306 (2) ◽  
pp. 497-504 ◽  
Author(s):  
P J Higgins ◽  
L Staiano-Coico ◽  
M P Ryan
Keyword(s):  
Protein Synthesis ◽  
Transcriptional Activation ◽  
Actinomycin D ◽  
De Novo ◽  
Rat Kidney ◽  
Secondary Response ◽  
Protein Levels ◽  
Run Off ◽  
Dependent Pathway ◽  
Pai 1

Expression of the rat p52(PAI-1) gene is positively regulated by agents that influence cellular microfilament organization and/or cell-to-substrate adhesion [e.g. cytochalasin D (CD) and sodium n-butyrate (NaB)] [Higgins, Chaudhari and Ryan (1991) Biochem. J. 273, 651-658; Higgins, Ryan and Providence (1994) J. Cell. Physiol. 159, 187-195]. As shape-responsive genes may be subject to inducer-specific controls, the biochemical mechanisms underlying the shape-dependent pathway of p52(PAI-1) gene regulation were examined in v-ras-transformed rat kidney (KNRK) cells. NaB and/or CD effectively stimulated p52(PAI-1) run-off transcription and augmented de novo p52(PAI-1) mRNA and protein synthesis in KNRK cells; induction at both the mRNA and protein levels was inhibited by actinomycin D. Pretreatment with cycloheximide (CX) markedly attenuated NaB- and/or CD-stimulated p52(PAI-1) expression. CX alone, however, induced low levels of p52(PAI-1) mRNA; increased p52(PAI-1) protein synthesis was evident after release of KNRK cells from CX blockade. Such CX-mediated induction was also sensitive to actinomycin D. Full stimulation of p52(PAI-1) expression in KNRK cells in response to the shape modulators NaB and/or CD involves transcriptional activation of the p52(PAI-1) gene, requires de novo RNA synthesis and occurs through a secondary-response (i.e. protein-synthesis-dependent) pathway.

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