scholarly journals Tristetraprolin regulation of interleukin-22 production

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Lorena Härdle ◽  
Malte Bachmann ◽  
Franziska Bollmann ◽  
Andrea Pautz ◽  
Tobias Schmid ◽  
...  
Keyword(s):  
T Cells ◽  
Half Life ◽  
Hek293 Cells ◽  
Luciferase Reporter ◽  
Jurkat T Cells ◽  
Interleukin 22 ◽  
Novel Target ◽  
Post Transcriptional Regulation ◽  
Mrna Half Life

Abstract Interleukin (IL)-22 is a STAT3-activating cytokine displaying characteristic AU-rich elements (ARE) in the 3′-untranslated region (3′-UTR) of its mRNA. This architecture suggests gene regulation by modulation of mRNA stability. Since related cytokines undergo post-transcriptional regulation by ARE-binding tristetraprolin (TTP), the role of this destabilizing protein in IL-22 production was investigated. Herein, we demonstrate that TTP-deficient mice display augmented serum IL-22. Likewise, IL-22 mRNA was enhanced in TTP-deficient splenocytes and isolated primary T cells. A pivotal role for TTP is underscored by an extended IL-22 mRNA half-life detectable in TTP-deficient T cells. Luciferase-reporter assays performed in human Jurkat T cells proved the destabilizing potential of the human IL-22-3′-UTR. Furthermore, overexpression of TTP in HEK293 cells substantially decreased luciferase activity directed by the IL-22-3′-UTR. Transcript destabilization by TTP was nullified upon cellular activation by TPA/A23187, an effect dependent on MEK1/2 activity. Accordingly, IL-22 mRNA half-life as determined in TPA/A23187-stimulated Jurkat T cells decreased under the influence of the MEK1/2 inhibitor U0126. Altogether, data indicate that TTP directly controls IL-22 production, a process counteracted by MEK1/2. The TTP-dependent regulatory pathway described herein likely contributes to the role of IL-22 in inflammation and cancer and may evolve as novel target for pharmacological IL-22 modulation.

scholarly journals Regulation of thrombospondin-1 expression through AU-rich elements in the 3′UTR of the mRNA

2011 ◽  
Vol 16 (1) ◽  
Author(s):  
Asa Mcgray ◽  
Timothy Gingerich ◽  
James Petrik ◽  
Jonathan Lamarre
Keyword(s):  
Gene Expression ◽  
Mrna Stability ◽  
Half Life ◽  
Rna Binding ◽  
Binding Protein ◽  
Site Directed Mutagenesis ◽  
Hek293 Cells ◽  
Luciferase Reporter ◽  
Thrombospondin 1 ◽  
Mrna Half Life

AbstractThrombospondin-1 (TSP-1) is a matricellular protein that participates in numerous normal and pathological tissue processes and is rapidly modulated by different stimuli. The presence of 8 highly-conserved AU rich elements (AREs) within the 3′-untranslated region (3′UTR) of the TSP-1 mRNA suggests that post-transcriptional regulation is likely to represent one mechanism by which TSP-1 gene expression is regulated. We investigated the roles of these AREs, and proteins which bind to them, in the control of TSP-1 mRNA stability. The endogenous TSP-1 mRNA half-life is approximately 2.0 hours in HEK293 cells. Luciferase reporter mRNAs containing the TSP-1 3′UTR show a similar rate of decay, suggesting that the 3′UTR influences the decay rate. Site-directed mutagenesis of individual and adjacent AREs prolonged reporter mRNA halflife to between 2.2 and 4.4 hours. Mutation of all AREs increased mRNA half life to 8.8 hours, suggesting that all AREs have some effect, but that specific AREs may have key roles in stability regulation. A labeled RNA oligonucleotide derived from the most influential ARE was utilized to purify TSP-1 AREbinding proteins. The AU-binding protein AUF1 was shown to associate with this motif. These studies reveal that AREs in the 3′UTR control TSP-1 mRNA stability and that the RNA binding protein AUF1 participates in this control. These studies suggest that ARE-dependent control of TSP-1 mRNA stability may represent an important component in the control of TSP-1 gene expression.

The role of cbl family of ubiquitin ligases in gastric cancer exosome-induced apoptosis of Jurkat T cells

2009 ◽  
pp. 1-8
Author(s):  
Jing-Lei Qu ◽  
Xiu-Juan Qu ◽  
Ming-Fang Zhao ◽  
Yue-E Teng ◽  
Ye Zhang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
T Cells ◽  
Ubiquitin Ligases ◽  
Jurkat T Cells ◽  
Induced Apoptosis

scholarly journals Role for a Glycan Phosphoinositol Anchor in Fcγ Receptor Synergy

1997 ◽  
Vol 139 (5) ◽  
pp. 1209-1217 ◽  
Author(s):  
Jennifer M. Green ◽  
Alan D. Schreiber ◽  
Eric J. Brown
Keyword(s):  
T Cells ◽  
Cell Types ◽  
Polymorphonuclear Neutrophils ◽  
Fcγ Receptor ◽  
Fcγ Receptors ◽  
Jurkat T Cells ◽  
Gpi Anchor ◽  
Transmembrane Anchor ◽  
Activation Motif

While many cell types express receptors for the Fc domain of IgG (FcγR), only primate polymorphonuclear neutrophils (PMN) express an FcγR linked to the membrane via a glycan phosphoinositol (GPI) anchor. Previous studies have demonstrated that this GPI-linked FcγR (FcγRIIIB) cooperates with the transmembrane FcγR (FcγRIIA) to mediate many of the functional effects of immune complex binding. To determine the role of the GPI anchor in Fcγ receptor synergy, we have developed a model system in Jurkat T cells, which lack endogenously expressed Fcγ receptors. Jurkat T cells were stably transfected with cDNA encoding FcγRIIA and/or FcγRIIIB. Cocrosslinking the two receptors produced a synergistic rise in intracytoplasmic calcium ([Ca2+]i) to levels not reached by stimulation of either FcγRIIA or FcγRIIIB alone. Synergy was achieved by prolonged entry of extracellular Ca2+. Cocrosslinking FcγRIIA with CD59 or CD48, two other GPI-linked proteins on Jurkat T cells also led to a synergistic [Ca2+]i rise, as did crosslinking CD59 with FcγRIIA on PMN, suggesting that interactions between the extracellular domains of the two Fcγ receptors are not required for synergy. Replacement of the GPI anchor of FcγRIIIB with a transmembrane anchor abolished synergy. In addition, tyrosine to phenylalanine substitutions in the immunoreceptor tyrosine-based activation motif (ITAM) of the FcγRIIA cytoplasmic tail abolished synergy. While the ITAM of FcγRIIA was required for the increase in [Ca2+]i, tyrosine phosphorylation of crosslinked FcγRIIA was diminished when cocrosslinked with FcγRIIIB. These data demonstrate that FcγRIIA association with GPI-linked proteins facilitates FcγR signal transduction and suggest that this may be a physiologically significant role for the unusual GPI-anchored FcγR of human PMN.

The KH-type splicing regulatory protein (KSRP) regulates type III interferon expression post-transcriptionally

2019 ◽  
Vol 476 (2) ◽  
pp. 333-352 ◽  
Author(s):  
Lisa Schmidtke ◽  
Katharina Schrick ◽  
Sabrina Saurin ◽  
Rudolf Käfer ◽  
Fabian Gather ◽  
...  
Keyword(s):  
Half Life ◽  
Rna Binding ◽  
Binding Protein ◽  
Regulatory Protein ◽  
Rna Binding Protein ◽  
Luciferase Reporter ◽  
Regulatory Function ◽  
Type Iii ◽  
Type Iii Ifn ◽  
Mrna Half Life

Abstract Type III interferons (IFNs) are the latest members of the IFN family. They play an important role in immune defense mechanisms, especially in antiviral responses at mucosal sites. Moreover, they control inflammatory reactions by modulating neutrophil and dendritic cell functions. Therefore, it is important to identify cellular mechanisms involved in the control of type III IFN expression. All IFN family members contain AU-rich elements (AREs) in the 3′-untranslated regions (3′-UTR) of their mRNAs that determine mRNA half-life and consequently the expressional level of these cytokines. mRNA stability is controlled by different proteins binding to these AREs leading to either stabilization or destabilization of the respective target mRNA. The KH-type splicing regulatory protein KSRP (also named KHSRP) is an important negative regulator of ARE-containing mRNAs. Here, we identify the interferon lambda 3 (IFNL3) mRNA as a new KSRP target by pull-down and immunoprecipitation experiments, as well as luciferase reporter gene assays. We characterize the KSRP-binding site in the IFNL3 3′-UTR and demonstrate that KSRP regulates the mRNA half-life of the IFNL3 transcript. In addition, we detect enhanced expression of IFNL3 mRNA in KSRP−/− mice, establishing a negative regulatory function of KSRP in type III IFN expression also in vivo. Besides KSRP the RNA-binding protein AUF1 (AU-rich element RNA-binding protein 1) also seems to be involved in the regulation of type III IFN mRNA expression.

scholarly journals MiR-125b regulates inflammation in bovine mammary epithelial cells by targeting the NKIRAS2 gene

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Zhuo-Ma Luoreng ◽  
Da-Wei Wei ◽  
Xing-Ping Wang
Keyword(s):  
T Cells ◽  
Epithelial Cells ◽  
Dairy Cows ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Luciferase Reporter ◽  
Regulation Mechanism ◽  
Bovine Mammary Epithelial Cells ◽  
Tnf Α

AbstractMastitis is a complex inflammatory disease caused by pathogenic infection of mammary tissue in dairy cows. The molecular mechanism behind its occurrence, development, and regulation consists of a multi-gene network including microRNA (miRNA). Until now, there is no report on the role of miR-125b in regulating mastitis in dairy cows. This study found that miR-125b expression is significantly decreased in lipopolysaccharide (LPS)-induced MAC-T bovine mammary epithelial cells. Also, its expression is negatively correlated with the expression of NF-κB inhibitor interacting Ras-like 2 (NKIRAS2) gene. MiR-125b target genes were identified using a double luciferase reporter gene assay, which showed that miR-125b can bind to the 3′ untranslated region (3′ UTR) of the NKIRAS2, but not the 3′UTR of the TNF-α induced protein 3 (TNFAIP3). In addition, miR-125b overexpression and silencing were used to investigate the role of miR-125b on inflammation in LPS-induced MAC-T. The results demonstrate that a reduction in miR-125b expression in LPS-induced MAC-T cells increases NKIRAS2 expression, which then reduces NF-κB activity, leading to low expression of the inflammatory factors IL-6 and TNF-α. Ultimately, this reduces the inflammatory response in MAC-T cells. These results indicate that miR-125b is a pro-inflammatory regulator and that its silencing can alleviate bovine mastitis. These findings lay a foundation for elucidating the molecular regulation mechanism of cow mastitis.

Mixed tailing by TENT4A and TENT4B shields mRNA from rapid deadenylation

Science ◽  
2018 ◽  
Vol 361 (6403) ◽  
pp. 701-704 ◽  
Author(s):  
Jaechul Lim ◽  
Dongwan Kim ◽  
Young-suk Lee ◽  
Minju Ha ◽  
Mihye Lee ◽  
...  
Keyword(s):  
Gene Regulation ◽  
Half Life ◽  
Messenger Rna ◽  
Mrna Translation ◽  
Posttranscriptional Gene Regulation ◽  
And Function ◽  
Mrna Half Life ◽  
In Cells

RNA tails play integral roles in the regulation of messenger RNA (mRNA) translation and decay. Guanylation of the poly(A) tail was discovered recently, yet the enzymology and function remain obscure. Here we identify TENT4A (PAPD7) and TENT4B (PAPD5) as the enzymes responsible for mRNA guanylation. Purified TENT4 proteins generate a mixed poly(A) tail with intermittent non-adenosine residues, the most common of which is guanosine. A single guanosine residue is sufficient to impede the deadenylase CCR4-NOT complex, which trims the tail and exposes guanosine at the 3′ end. Consistently, depletion of TENT4A and TENT4B leads to a decrease in mRNA half-life and abundance in cells. Thus, TENT4A and TENT4B produce a mixed tail that shields mRNA from rapid deadenylation. Our study unveils the role of mixed tailing and expands the complexity of posttranscriptional gene regulation.

scholarly journals The protective role of miR-223 in sepsis-induced mortality

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Dan Liu ◽  
Zhiding Wang ◽  
Huijuan Wang ◽  
Feifei Ren ◽  
Yanqin Li ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Proliferation ◽  
T Cells ◽  
Protective Factor ◽  
Linear Regression Analysis ◽  
Multiple Linear Regression Analysis ◽  
Negative Control ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Jurkat T Cells

Abstract Lymphocyte apoptosis appears to play an important role in immunodysfunction in sepsis. We investigated the role of miR-223 in cell proliferation and apoptosis to identify potential target downstream proteins in sepsis. We recruited 143 patients with sepsis and 44 healthy controls from the Chinese PLA General Hospital. Flow cytometry was used to sort monocytes, lymphocytes, and neutrophils from fresh peripheral blood. A miR-223 mimic and inhibitor were used for transient transfection of Jurkat T cells. Quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) was used to assess expression of the miRNAs in cells. Western blot analysis was performed to measure protein expression. We evaluated the cell cycle and apoptosis by using flow cytometry (FCM) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). Expression of miR-223 was significantly higher in the survivor group than in the nonsurvivor group. Multiple linear regression analysis revealed that SOFA scores correlated negatively with miR-223 and monocyte counts, with β coefficients (95% CI) of − 0.048 (− 0.077, − 0.019) and − 47.707 (− 83.871, − 11.543), respectively. miR-223 expression also correlated negatively with the percentage of apoptosis in lymphocytes. The rate of apoptosis in the miR-223 mimic group was significantly lower than that of the negative control, with an adverse outcome observed in the miR-223 inhibitor group. We also found that miR-223 enhanced the proliferation of Jurkat T cells and that inhibiting miR-223 had an inhibitory effect on the G1/S transition. We conclude that miR-223 can serve as a protective factor in sepsis by reducing apoptosis and enhancing cell proliferation in lymphocytes by interacting with FOXO1. Potential downstream molecules are HSP60, HSP70, and HTRA.

scholarly journals LINC01006 facilitates cell proliferation, migration and invasion in prostate cancer through targeting miR-34a-5p to up-regulate DAAM1

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Enhui Ma ◽  
Qianqian Wang ◽  
Jinhua Li ◽  
Xinqi Zhang ◽  
Zhenjia Guo ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Prostate Gland ◽  
Inhibitory Effect ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Protein Coding ◽  
Novel Target

Abstract Background Prostate cancer (PCa) is a kind of malignancy occurring in the prostate gland. Substantial researches have proved the major role of long noncoding RNAs (lncRNAs) in PCa. However, the role of long intergenic non-protein coding RNA 1006 (LINC01006) in PCa has not been investigated yet. Methods RT-qPCR was used to examine the expression levels of LINC01006 and its downstream targets. The function of LINC01006 in PCa was tested by in vitro and in vivo assays. With application of RNA pull down, RNA immunoprecipitation (RIP) and luciferase reporter assays, the interaction among LINC01006, miR-34a-5p and disheveled associated activator of morphogenesis 1 (DAAM1) were verified. Results LINC01006 expression presented high in PCa cell lines. LINC01006 silencing suppressed cell proliferative, migratory, invasive capacities while accelerated apoptotic rate. Besides, LINC01006 knockdown also suppressed tumor growth and metastasis in vivo. Furthermore, miR-34a-5p, a tumor suppressor in PCa, was sponged by LINC01006. Moreover, DAAM1 was targeted by miR-34a-5p and promoted PCa progression. More intriguingly, rescue assays suggested that the inhibitory effect of LINC01006 knockdown on PCa development was offset by DAAM1 overexpression. Conclusions LINC01006 promoted PCa progression by sponging miR-34a-5p to up-regulate DAAM1, providing a novel target for PCa therapy.

Transcriptional and post-transcriptional regulation of prolactin during the turkey reproductive cycle

1997 ◽  
Vol 18 (3) ◽  
pp. 223-231 ◽  
Author(s):  
Z Tong ◽  
G R Pitts ◽  
D N Foster ◽  
M E El Halawani
Keyword(s):  
Mrna Stability ◽  
Half Life ◽  
Northern Blot Analysis ◽  
Transcription Inhibitor ◽  
Steady State Levels ◽  
Incubating Birds ◽  
Plasma Prl ◽  
Photoinduced Changes ◽  
Post Transcriptional Regulation ◽  
Mrna Half Life

ABSTRACT The present study examined turkey prolactin (PRL) transcription and PRL mRNA stability during different reproductive stages. Nuclear run-on transcription assays were performed using isolated nuclei from pituitaries of turkeys at different reproductive stages. Meanwhile, cytoplasmic PRL mRNA and plasma PRL were measured by slot blot and RIA respectively. The PRL transcription, pituitary cytoplasmic PRL mRNA abundance and plasma PRL levels increased after photostimulation and peaked at the incubating stage (P<0·05). A decrease in PRL transcription, pituitary cytoplasmic PRL mRNA and plasma PRL (P<0·05) was observed during the transition from incubation to photorefractoriness. Nest-deprivation reduced circulating PRL (P<0·05), whereas pituitary cytoplasmic PRL mRNA and PRL transcription were not significantly altered from those in incubating birds (P>0·05). The half-life of PRL mRNA was determined in pituitaries of non-photostimulated, laying, incubating and photorefractory hens. Primary pituitary cell cultures were treated with the transcription inhibitor actinomycin-D and the decay of the pre-existing PRL mRNA was quantified using Northern blot analysis. The PRL mRNA half-life was 1·5- and 1·4-fold greater in incubating and laying birds respectively than in non-photostimulated turkeys (P<0·05). The half-life of PRL mRNA in photorefractory and incubating hens was similar in spite of great differences in pituitary PRL mRNA steady-state levels and PRL transcription. Our data suggest that photoinduced changes in pituitary PRL mRNA and plasma PRL are due to changes in both PRL transcription and PRL mRNA stability. Nest-deprivation inhibits the PRL releasing mechanism(s) independently of PRL transcription in turkeys.

scholarly journals Costimulation by B7-1 and LFA-3 targets distinct nuclear factors that bind to the interleukin-2 promoter: B7-1 negatively regulates LFA-3-induced NF-AT DNA binding.

1997 ◽  
Vol 17 (3) ◽  
pp. 1314-1323 ◽  
Author(s):  
E Parra ◽  
M Varga ◽  
G Hedlund ◽  
T Kalland ◽  
M Dohlsten
Keyword(s):  
T Cells ◽  
Dna Binding ◽  
Transcriptional Activity ◽  
Interleukin 2 ◽  
Luciferase Reporter ◽  
Jurkat T Cells ◽  
Response Elements ◽  
Nuclear Factors ◽  
Hla Dr ◽  
Nf Kappab

We have characterized the regulation of nuclear factors involved in transcriptional control of the interleukin-2 (IL-2) promoter-enhancer activity in Jurkat T cells stimulated with superantigen presented on HLA-DR transfectants combined with the ligands LFA-3 (CD58) and B7-1 (CD80). Gel shift analyses showed that NF-AT was strongly induced in LFA-3-costimulated Jurkat T cells, suggesting that NF-AT is a key target nuclear factor for the CD2-LFA-3 pathway. Studies using HLA-DR-B7-1-LFA-3 triple transfectants showed that the LFA-3-induced NF-AT DNA binding activity was negatively regulated by B7-1 costimulation. In contrast, induction of a CD28 response complex containing only c-Rel proteins was seen after B7-1 costimulation. Both LFA-3 costimulation and B7-1 costimulation induced the AP-1 and NF-kappaB nuclear factors. Distinct compositions of the NF-AT complexes were seen in B7-1- and LFA-3-costimulated cells. LFA-3 induced primarily Jun-D, Fra-1, and Fra-2, while B7-1 induced June-D-Fos complexes. In contrast, AP-1 and NF-kappaB complexes induced in B7-1- and LFA-3-costimulated T cells showed similar contents. Transient transfection of Jurkat T cells with a construct encoding the IL-2 enhancer-promoter region (position -500 to +60) linked to a luciferase reporter gene revealed that B7-1 costimulation was required to induce strong transcriptional activity. Combined B7-1-LFA-3 costimulation resulted in a synergistic increase in IL-2 transcriptional activity. Multimers of the AP-1, NF-AT, NF-kappaB, and CD28 response elements showed distinct kinetics and activity after LFA-3 and B7-1 costimulation and revealed that B7-1 and LFA-3 converge to superinduce transcriptional activity of the AP-1, NF-AT, and CD28 response elements. Transcriptional studies with an IL-2 enhancer-promoter carrying a mutation in the CD28 response element site revealed that the activity was reduced by 80% after B7-1 and B7-1-LFA-3 costimulation whereas the transcriptional activity induced by LFA-3 was unaffected. Our data strongly suggest a selectivity in induction of nuclear factors by the CD2-LFA-3 and CD28-B7-1 pathways. This selectivity may contribute to regulation of the levels of IL-2 induced by LFA-3 and B7-1 costimulation and favor autocrine and paracrine T-cell responses, respectively.

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