scholarly journals P23 Acts as Functional RBP in the Macrophage Inflammation Response

2021 ◽  
Vol 8 ◽  
Author(s):  
Sebastian de Vries ◽  
Vladimir Benes ◽  
Isabel S. Naarmann-de Vries ◽  
Cornelia Rücklé ◽  
Katharina Zarnack ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Raw 264.7 ◽  
Macrophage Migration ◽  
Raw 264.7 Macrophages ◽  
Mrna Destabilization ◽  
Bacterial Lipopolysaccharides ◽  
And Migration ◽  
Cellular Immune ◽  
Phagocytotic Activity

Macrophages exert the primary cellular immune response. Pathogen components like bacterial lipopolysaccharides (LPS) stimulate macrophage migration, phagocytotic activity and cytokine expression. Previously, we identified the poly(A)+ RNA interactome of RAW 264.7 macrophages. Of the 402 RNA-binding proteins (RBPs), 32 were classified as unique in macrophages, including nineteen not reported to interact with nucleic acids before. Remarkably, P23 a HSP90 co-chaperone, also known as cytosolic prostaglandin E2 synthase (PTGES3), exhibited differential poly(A)+ RNA binding in untreated and LPS-induced macrophages. To identify mRNAs bound by P23 and to elucidate potential regulatory RBP functions in macrophages, we immunoprecipitated P23 from cytoplasmic extracts of cross-linked untreated and LPS-induced cells. RNAseq revealed that enrichment of 44 mRNAs was reduced in response to LPS. Kif15 mRNA, which encodes kinesin family member 15 (KIF15), a motor protein implicated in cytoskeletal reorganization and cell mobility was selected for further analysis. Noteworthy, phagocytic activity of LPS-induced macrophages was enhanced by P23 depletion. Specifically, in untreated RAW 264.7 macrophages, decreased P23 results in Kif15 mRNA destabilization, diminished KIF15 expression and accelerated macrophage migration. We show that the unexpected RBP function of P23 contributes to the regulation of macrophage phagocytotic activity and migration.

scholarly journals Polyamine Transport Protein PotD Protects Mice against Haemophilus parasuis and Elevates the Secretion of Pro-Inflammatory Cytokines of Macrophage via JNK–MAPK and NF–κB Signal Pathways through TLR4

Vaccines ◽  
2019 ◽  
Vol 7 (4) ◽  
pp. 216 ◽  
Author(s):  
Ke Dai ◽  
Xiaoyu Ma ◽  
Zhen Yang ◽  
Yung-Fu Chang ◽  
Sanjie Cao ◽  
...  
Keyword(s):  
Immune Responses ◽  
Inflammatory Cytokines ◽  
Transport System ◽  
Signal Pathways ◽  
Raw 264.7 ◽  
Raw 264.7 Macrophages ◽  
Cellular Immune Responses ◽  
Polyamine Transport ◽  
Cellular Immune ◽  
Pro Inflammatory Cytokines

The potD gene, belonging to the well-conserved ABC (ATP-binding cassette) transport system potABCD, encodes the bacterial substrate-binding subunit of the polyamine transport system. In this study, we found PotD in Haemophilus (Glaesserella) parasuis could actively stimulate both humoral immune and cellular immune responses and elevate lymphocyte proliferation, thus eliciting a Th1-type immune response in a murine immunity and infection model. Stimulation of Raw 264.7 macrophages with PotD validated that Toll-like receptor 4, rather than 2, participated in the positive transcription and expression of pro-inflammatory cytokines IL–1β, IL–6, and TNF–α using qPCR and ELISA. Blocking signal-regulated JNK–MAPK and RelA(p65) pathways significantly decreased PotD-induced pro-inflammatory cytokine production. Overall, we conclude that vaccination of PotD could induce both humoral and cellular immune responses and provide immunoprotection against H. parasuis challenge. The data also suggest that Glaesserella PotD is a novel pro-inflammatory mediator and induces TLR4-dependent pro-inflammatory activity in Raw 264.7 macrophages through JNK–MAPK and RelA(p65) pathways.

scholarly journals Phosphorylation of the ARE-binding protein DAZAP1 by ERK2 induces its dissociation from DAZ

2006 ◽  
Vol 399 (2) ◽  
pp. 265-273 ◽  
Author(s):  
Simon Morton ◽  
Huei-Ting Yang ◽  
Ntsane Moleleki ◽  
David G. Campbell ◽  
Philip Cohen ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Rna Binding ◽  
Binding Protein ◽  
Mitogen Activated Protein Kinase ◽  
Raw 264.7 ◽  
Raw 264.7 Macrophages ◽  
Hek 293 ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

A protein in RAW 264.7 macrophages, which became phosphorylated in response to LPS (lipopolysaccharide), was identified as the RNA-binding protein called DAZAP1 [DAZ (deleted in azoospermia)-associated protein 1]. The phosphorylation of this protein was prevented by specific inhibition of MKK1 [MAPK (mitogen-activated protein kinase) kinase 1], indicating that it was phosphorylated via the classical MAPK cascade. Further experiments showed that DAZAP1 was phosphorylated stoichiometrically in vitro by ERK2 (extracellular-signal-regulated protein kinase 2) at two Thr-Pro sequences (Thr269 and Thr315), and that both sites became phosphorylated in HEK-293 (human embryonic kidney 293) cells in response to PMA or EGF (epidermal growth factor), or RAW 264.7 macrophages in response to LPS. Phosphorylation induced by each stimulus was prevented by two structurally distinct inhibitors of MKK1 (PD184352 and U0126), demonstrating that DAZAP1 is a physiological substrate for ERK1/ERK2. The mutation of Thr269 and Thr315 to aspartate or the phosphorylation of these residues caused DAZAP1 to dissociate from its binding partner DAZ. DAZ interacts with PABP [poly(A)-binding protein] and thereby stimulates the translation of mRNAs containing short poly(A) tails [Collier, Gorgoni, Loveridge, Cooke and Gray (2005) EMBO J. 24, 2656–2666]. In the present study we have shown that DAZ cannot bind simultaneously to DAZAP1 and PABP, and suggest that the phosphorylation-induced dissociation of DAZ and DAZAP1 may allow the former to stimulate translation by interacting with PABP.

scholarly journals Conceptual Advances in Control of Inflammation by the RNA-Binding Protein Tristetraprolin

2021 ◽  
Vol 12 ◽  
Author(s):  
Pavel Kovarik ◽  
Annika Bestehorn ◽  
Jeanne Fesselet
Keyword(s):  
Immune Responses ◽  
Mrna Stability ◽  
Binding Sites ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Degradation ◽  
Decay Rates ◽  
Mrna Destabilization

Regulated changes in mRNA stability are critical drivers of gene expression adaptations to immunological cues. mRNA stability is controlled mainly by RNA-binding proteins (RBPs) which can directly cleave mRNA but more often act as adaptors for the recruitment of the RNA-degradation machinery. One of the most prominent RBPs with regulatory roles in the immune system is tristetraprolin (TTP). TTP targets mainly inflammation-associated mRNAs for degradation and is indispensable for the resolution of inflammation as well as the maintenance of immune homeostasis. Recent advances in the transcriptome-wide knowledge of mRNA expression and decay rates together with TTP binding sites in the target mRNAs revealed important limitations in our understanding of molecular mechanisms of TTP action. Such orthogonal analyses lead to the discovery that TTP binding destabilizes some bound mRNAs but not others in the same cell. Moreover, comparisons of various immune cells indicated that an mRNA can be destabilized by TTP in one cell type while it remains stable in a different cell linage despite the presence of TTP. The action of TTP extends from mRNA destabilization to inhibition of translation in a subset of targets. This article will discuss these unexpected context-dependent functions and their implications for the regulation of immune responses. Attention will be also payed to new insights into the role of TTP in physiology and tissue homeostasis.

scholarly journals m6A Reader YTHDF2 Regulates LPS-Induced Inflammatory Response

2019 ◽  
Vol 20 (6) ◽  
pp. 1323 ◽  
Author(s):  
Ruiqing Yu ◽  
Qimeng Li ◽  
Zhihui Feng ◽  
Luhui Cai ◽  
Qiong Xu
Keyword(s):  
Inflammatory Response ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mapk Signaling ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Tnf Α ◽  
Mrna Transcripts ◽  
The Stability ◽  
Cell Stress Response

N6-methyladenosine (m6A) is an abundant mRNA modification that affects multiple biological processes, including those involved in the cell stress response and viral infection. YTH domain family 2 (YTHDF2) is an m6A-binding protein that affects the localization and stability of targeted mRNA. RNA-binding proteins (RBPs) can regulate the stability of inflammatory gene mRNA transcripts, thus participating in the regulation of inflammatory processes. As an RBP, the role of YTHDF2 in the LPS-induced inflammatory reaction has not been reported. To elucidate the function of YTHDF2 in the inflammatory response of macrophages, we first detected the expression level of YTHDF2 in RAW 264.7 cells, and found that it was upregulated after LPS stimulation. YTHDF2 knockdown significantly increased the LPS-induced IL-6, TNF-α, IL-1β, and IL-12 expression and the phosphorylation of p65, p38, and ERK1/2 in NF-κB and MAPK signaling. Moreover, the upregulated expression of TNF-α and IL-6 in cells with silenced YTHDF2 expression was downregulated by the NF-κB, p38, and ERK inhibitors. YTHDF2 depletion increased the expression and stability of MAP2K4 and MAP4K4 mRNAs. All of these results suggest that YTHDF2 knockdown increases mRNA expression levels of MAP2K4 and MAP4K4 via stabilizing the mRNA transcripts, which activate MAPK and NF-κB signaling pathways, which promote the expression of proinflammatory cytokines and aggravate the inflammatory response in LPS-stimulated RAW 264.7 cells.

scholarly journals Thy-1 mRNA destabilization by norepinephrine a 3′ UTR cAMP responsive decay element and involves RNA binding proteins

2010 ◽  
Vol 24 (7) ◽  
pp. 1078-1088 ◽  
Author(s):  
Melissa D. LaJevic ◽  
Sujatha P. Koduvayur ◽  
Veronique Caffrey ◽  
Rhonna L. Cohen ◽  
Donald A. Chambers
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Destabilization

scholarly journals Effect of modified RNA-binding proteins HuR on biological behavior of bladder cancer T24 cell line

2019 ◽  
Author(s):  
Kewen Zheng ◽  
Yan Su ◽  
Xiaomin Han ◽  
Qiang Ma
Keyword(s):  
Bladder Cancer ◽  
Cell Viability ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Annexin V ◽  
Biological Behavior ◽  
Control Group ◽  
Related Factors ◽  
T24 Cells ◽  
And Migration

Background: In tumors, the role of human antigen R (HuR) involves regulating tumor cell proliferation, differentiation, apoptosis, angiogenesis, and lymphangiogenesis. Previous studies have revealed the expression of HuR can be detected in bladder cancer and related to biological behavior of malignancy. Methods: T24 cells were transfected by HuR overexpression vector and HuR knockdown vector. The cells were divided into control group, overExp-HuR group and cas9-HuR group. The cell viability after 48 h was detected by MTT, the apoptosis was detected by Annexin V-APC/7-AAD double staining, the cell migration was detected by Transwell assay, and the expression of HuR, cyclinD1 and apoptosis-related factors (Bcl-2) were detected by fluorescence quantitative PCR and Western blot. Results: Compared with the control group, the cell viability after 48 h in the overExp-HuR group increased significantly, and decreased in the cas9-HuR group (P<0.05). And the number of migrating cells increased in the overExp-HuR group, and decreased in the cas9-HuR group (P<0.05). The apoptosis rate of the overExp-HuR group decreased significantly, and increased significantly in the cas9-HuR group (P<0.05). The mRNA and protein expression of HuR, cyclinD1 and Bcl-2 of the overExp-HuR group increased, and decreased significantly in cas9-HuR group (P<0.05). Conclusion: HuR promote the proliferation and migration of T24 cells, and inhibit cell apoptosis. And the mechanism may be related to the expression of cyclin D and apoptosis-related proteins Bcl2.

scholarly journals U2AF2 Promote the Proliferation and Migration of Esophageal Adenocarcinoma Cells by Sustaining the Mrna Stability of SNORA21

2021 ◽  
Author(s):  
Gao Lijuan ◽  
Chen Yongshun ◽  
Li Bin
Keyword(s):  
Western Blot ◽  
Esophageal Adenocarcinoma ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Expression Level ◽  
Brdu Incorporation ◽  
Western World ◽  
Adenocarcinoma Cells ◽  
And Migration ◽  
Proliferation And Migration

Abstract Background Esophageal adenocarcinoma has gains predominance with squamous carcinoma moving to a second place in the western world. The 5-year survival rate of patients with esophageal cancer (EC) is still between 15–25%, although the treatment strategies for EC have been improved. RNA-binding proteins (RBPs) are key players in post-transcriptional events. Particularly, the U2 snRNP auxiliary (U2AF2), as a canonical RBP, has been reported to play an important role in the development, progression, and metastasis of several human cancers. However, the biological role of U2AF2 in the context of malignancies, including EC, has not yet been reported. This study is aimed to investigate the role and mechanism of U2AF2 in esophageal adenocarcinoma cell line. Methods The expression level of U2AF2 and SNORA21 was analysed by TCGA database. The expression level of U2AF2 and SNORA21 in esophageal adenocarcinoma cells was determined by Western Blot and RT-qPCR assay. The cell viability and proliferation was detected by CCK8 assay and BrdU incorporation assay. The apoptosis was examined by caspase3 immunocytofluorescense. The protein levels of Bax, Bcl-2 and E-cadherin were explored by Western Blot assay. The interaction between U2AF2 and SNORA21 was predicted by Starbase 3.0 and confirmed by RIP assay. The RNA regulated by U2AF2 was through RNA-seq analysis on the basis of downregulation of U2AF2. Results The expression of U2AF2 and SNORA21 was found to be upregulated and served as an unfavourable factor which negatively related with overall survival of esophageal adenocarcinoma patients. Silencing the expression of U2AF2 or SNORA21 could suppress the proliferation and migration of OE19 and OE33 cells and stimulate its apoptosis. Moreover, we identified that the U2AF2 could bind and interact with SNORA21 to sustain its stability. Furthermore, we show that overexpression of SNORA21 could rescue the proliferation inhibition phenotype caused by si-U2AF2. Conclusion Our study provides several lines of evidence supporting the intriguing concept that U2AF2 could promote the proliferation of OE19 and OE33 cells by directly targeting SNORA21, which might serve as a novel candidate biomarker and a new target for the treatment of esophageal adenocarcinoma.

scholarly journals Effect of modified RNA-binding proteins HuR on biological behavior of bladder cancer T24 cell line

2019 ◽  
Author(s):  
Kewen Zheng ◽  
Yan Su ◽  
Xiaomin Han ◽  
Qiang Ma
Keyword(s):  
Bladder Cancer ◽  
Cell Viability ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Annexin V ◽  
Biological Behavior ◽  
Control Group ◽  
Related Factors ◽  
T24 Cells ◽  
And Migration

Background: In tumors, the role of human antigen R (HuR) involves regulating tumor cell proliferation, differentiation, apoptosis, angiogenesis, and lymphangiogenesis. Previous studies have revealed the expression of HuR can be detected in bladder cancer and related to biological behavior of malignancy. Methods: T24 cells were transfected by HuR overexpression vector and HuR knockdown vector. The cells were divided into control group, overExp-HuR group and cas9-HuR group. The cell viability after 48 h was detected by MTT, the apoptosis was detected by Annexin V-APC/7-AAD double staining, the cell migration was detected by Transwell assay, and the expression of HuR, cyclinD1 and apoptosis-related factors (Bcl-2) were detected by fluorescence quantitative PCR and Western blot. Results: Compared with the control group, the cell viability after 48 h in the overExp-HuR group increased significantly, and decreased in the cas9-HuR group (P<0.05). And the number of migrating cells increased in the overExp-HuR group, and decreased in the cas9-HuR group (P<0.05). The apoptosis rate of the overExp-HuR group decreased significantly, and increased significantly in the cas9-HuR group (P<0.05). The mRNA and protein expression of HuR, cyclinD1 and Bcl-2 of the overExp-HuR group increased, and decreased significantly in cas9-HuR group (P<0.05). Conclusion: HuR promote the proliferation and migration of T24 cells, and inhibit cell apoptosis. And the mechanism may be related to the expression of cyclin D and apoptosis-related proteins Bcl2.

scholarly journals Knockdown of Musashi RNA Binding Proteins Decreases Radioresistance but Enhances Cell Motility and Invasion in Triple-Negative Breast Cancer

2020 ◽  
Vol 21 (6) ◽  
pp. 2169
Author(s):  
Fabian M. Troschel ◽  
Annemarie Minte ◽  
Yahia Mahmoud Ismail ◽  
Amr Kamal ◽  
Mahmoud Salah Abdullah ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Invasion ◽  
Cell Cycle Progression ◽  
Triple Negative ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Notch Pathway ◽  
Cycle Progression ◽  
Invasion And Migration ◽  
And Migration

The therapeutic potential of Musashi (MSI) RNA-binding proteins, important stemness-associated gene expression regulators, remains insufficiently understood in breast cancer. This study identifies the interplay between MSI protein expression, stem cell characteristics, radioresistance, cell invasiveness and migration. MSI-1, MSI-2 and Notch pathway elements were investigated via quantitative polymerase chain reaction (qPCR) in 19 triple-negative breast cancer samples. Measurements were repeated in MDA-MB-231 cells after MSI-1 and -2 siRNA-mediated double knockdown, with further experiments performed after MSI silencing. Flow cytometry helped quantify expression of CD44 and leukemia inhibitory factor receptor (LIFR), changes in apoptosis and cell cycle progression. Proliferation and irradiation-induced effects were assessed using colony formation assays. Radiation-related proteins were investigated via Western blots. Finally, cell invasion assays and digital holographic microscopy for cell migration were performed. MSI proteins showed strong correlations with Notch pathway elements. MSI knockdown resulted in reduction of stem cell marker expression, cell cycle progression and proliferation, while increasing apoptosis. Cells were radiosensitized as radioresistance-conferring proteins were downregulated. However, MSI-silencing-mediated LIFR downregulation resulted in enhanced cell invasion and migration. We conclude that, while MSI knockdown results in several therapeutically desirable consequences, enhanced invasion and migration need to be counteracted before knockdown advantages can be fully exploited.

Sign in / Sign up

Export Citation Format

Share Document