scholarly journals CaM kinase II regulates cardiac hemoglobin expression through histone phosphorylation upon sympathetic activation

2019 ◽  
Vol 116 (44) ◽  
pp. 22282-22287
Author(s):  
Ali Reza Saadatmand ◽  
Viviana Sramek ◽  
Silvio Weber ◽  
Daniel Finke ◽  
Matthias Dewenter ◽  
...  
Keyword(s):  
Messenger Rna ◽  
Target Genes ◽  
Ventricular Myocardium ◽  
Cam Kinase Ii ◽  
Sympathetic Activation ◽  
Cam Kinase ◽  
In Vivo Experiments ◽  
Underlying Mechanisms

Sympathetic activation of β-adrenoreceptors (β-AR) represents a hallmark in the development of heart failure (HF). However, little is known about the underlying mechanisms of gene regulation. In human ventricular myocardium from patients with end-stage HF, we found high levels of phosphorylated histone 3 at serine-28 (H3S28p). H3S28p was increased by inhibition of the catecholamine-sensitive protein phosphatase 1 and decreased by β-blocker pretreatment. By a series of in vitro and in vivo experiments, we show that the β-AR downstream protein kinase CaM kinase II (CaMKII) directly binds and phosphorylates H3S28. Whereas, in CaMKII-deficient myocytes, acute catecholaminergic stimulation resulted in some degree of H3S28p, sustained catecholaminergic stimulation almost entirely failed to induce H3S28p. Genome-wide analysis of CaMKII-mediated H3S28p in response to chronic β-AR stress by chromatin immunoprecipitation followed by massive genomic sequencing led to the identification of CaMKII-dependent H3S28p target genes. Forty percent of differentially H3S28p-enriched genomic regions were associated with differential, mostly increased expression of the nearest genes, pointing to CaMKII-dependent H3S28p as an activating histone mark. Remarkably, the adult hemoglobin genes showed an H3S28p enrichment close to their transcriptional start or end sites, which was associated with increased messenger RNA and protein expression. In summary, we demonstrate that chronic β-AR activation leads to CaMKII-mediated H3S28p in cardiomyocytes. Thus, H3S28p-dependent changes may play an unexpected role for cardiac hemoglobin regulation in the context of sympathetic activation. These data also imply that CaMKII may be a yet unrecognized stress-responsive regulator of hematopoesis.

scholarly journals LncRNA DLX6-AS1 regulates osteosarcoma progression via the miR-200a-3p/GPM6P axis

STEMedicine ◽  
2022 ◽  
Vol 3 (1) ◽  
pp. e111
Author(s):  
Tianyi Wu ◽  
Chen Huang ◽  
Feng Wang
Keyword(s):  
Target Genes ◽  
Critical Role ◽  
Tumor Xenograft ◽  
In Vivo Experiments ◽  
Biochemical Assays ◽  
Cell Metastasis ◽  
Underlying Mechanisms ◽  
Si Rna

LncRNA DLX6-AS1 takes part in the progression of various cancers. However, it is not elaborated clearly in osteosarcoma (OS) development. Therefore, we aimed to explore the impacts and specific mechanisms of DLX6-AS1 on the progression of OS. We estimated the pattern of DLX6-AS1 expression in Ost tissues and cells via quantitative reverse transcription polymerase chain reaction. A number of biochemical assays were carried out to assess the effects of DLX6-AS1. Target genes were predicted by bioinformatics methods. Then we used the transfection of si-RNA, miRNA inhibitor, and miRNA mimics to explore the underlying mechanisms and built tumor xenograft models for the in vivo experiments. A higher expression of DLX6-AS1 was found in OS tissues and cell lines, while knockdown of DXL6-AS1 suppressed OS cell metastasis and proliferation in vitro and in vivo. Mechanistically, it was revealed that DXL6-AS1 sponged miR-200a-3p, thus positively regulating the downstream GPM6B. In summary, DLX6-AS1 knockdown would inhibit OS cell migration, cell invasion, and cell proliferation, in which the DXL6-AS1/ miR-200a-3p/ GPM6B axis played a critical role.

scholarly journals MACC1 Suppresses Cell Apoptosis in Hepatocellular Carcinoma by Targeting the HGF/c-MET/AKT Pathway

2015 ◽  
Vol 35 (3) ◽  
pp. 983-996 ◽  
Author(s):  
Yingmin Yao ◽  
Chanwei Dou ◽  
Zhongtang Lu ◽  
Xin Zheng ◽  
Qingguang Liu
Keyword(s):  
Cell Apoptosis ◽  
Target Genes ◽  
Nuclear Translocation ◽  
Annexin V ◽  
Akt Pathway ◽  
Caspase 9 ◽  
Dapi Staining ◽  
In Vivo Experiments

Background & Aims: To investigate the expression and prognostic value of MACC1 in patients with HCC and identify the mechanism by which MACC1 inhibits HCC cell apoptosis. Methods: MACC1 and p-AKT expression was studied using immunohistochemistry of both HCC tissues and adjacent liver tissues. qRT-PCR and western immunoblotting were used to examine the expression of target genes at the mRNA and protein levels, respectively. The MTT assay was used to assess cell viability, and cell apoptosis was determined by DAPI staining, Annexin V/PI staining and Caspase 3/7 assay. Nude mice were used to perform in vivo experiments. Results: The overexpression of MACC1 was found in HCC tissues and was correlated with poor postsurgical prognosis. There was a positive relationship between MACC1 and p-AKT expression in HCC tissues. In vitro experiments showed that MACC1 repressed HCC cell apoptosis and promoted cell growth. Knockdown of c-MET abolished the anti-apoptotic function of MACC1. Next, MACC1 was verified to activate PI3K/AKT signaling by sensitizing HGF/c-MET signaling in HCC. MACC1 overexpression enhanced the HGF-driven phosphorylation of BAD, Caspase 9 and FKHRL1 and inhibited their pro-apoptotic functions in HCC cells. Finally, MACC1 was shown to inhibit cell apoptosis and promote HCC growth in vivo. Conclusions: This investigation revealed that MACC1 overexpression predicted worse prognosis after liver resection, which was attributed to the repression of HCC cell apoptosis via a molecular mechanism in which MACC1 accelerated the activation of the HGF/c-MET/PI3K/AKT pathway and phosphorylated BAD, Caspase 9 and FKHRL1, ultimately preventing their nuclear translocation and their pro-apoptotic function.

scholarly journals Oncogenic hijacking of a developmental transcription factor evokes vulnerability toward oxidative stress in Ewing sarcoma

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Aruna Marchetto ◽  
Shunya Ohmura ◽  
Martin F. Orth ◽  
Maximilian M. L. Knott ◽  
Maria V. Colombo ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factor ◽  
Ewing Sarcoma ◽  
Target Genes ◽  
Transcriptome Profiling ◽  
In Vivo Experiments ◽  
Ets Family ◽  
Pdx Models

AbstractEwing sarcoma (EwS) is an aggressive childhood cancer likely originating from mesenchymal stem cells or osteo-chondrogenic progenitors. It is characterized by fusion oncoproteins involving EWSR1 and variable members of the ETS-family of transcription factors (in 85% FLI1). EWSR1-FLI1 can induce target genes by using GGAA-microsatellites as enhancers.Here, we show that EWSR1-FLI1 hijacks the developmental transcription factor SOX6 – a physiological driver of proliferation of osteo-chondrogenic progenitors – by binding to an intronic GGAA-microsatellite, which promotes EwS growth in vitro and in vivo. Through integration of transcriptome-profiling, published drug-screening data, and functional in vitro and in vivo experiments including 3D and PDX models, we discover that constitutively high SOX6 expression promotes elevated levels of oxidative stress that create a therapeutic vulnerability toward the oxidative stress-inducing drug Elesclomol.Collectively, our results exemplify how aberrant activation of a developmental transcription factor by a dominant oncogene can promote malignancy, but provide opportunities for targeted therapy.

Disruption of HOX-PBX Interaction; a Potential Therapeutic Target in Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2057-2057
Author(s):  
Jastinder Sohal ◽  
Ruji Begum ◽  
Neal Fischbach ◽  
Davinder Theti ◽  
Ruth Pettengell ◽  
...  
Keyword(s):  
Target Genes ◽  
Hox Genes ◽  
Family Members ◽  
Normal Function ◽  
Leukemic Cells ◽  
Blue Staining ◽  
Rt Pcr ◽  
In Vivo Experiments

Abstract Most cases of acute myeloid leukemia (AML) are closely associated with gene rearrangements. Appraisal of these translocations and analysis of mouse models of leukemia has revealed that several members of the homeodomain containing family of transcription factors are implicated in the pathogenesis of leukemia. Overexpression of HOXA9 in murine models leads to the development of AML. This study focuses on the role of a subset of the HOX genes and their potential as a target for therapeutic intervention. We have designed a synthetic peptide, HXP4, that disrupts the interaction between HOX and PBX leading to growth inhibition of leukemic cells. An in vitro HOX-induced AML model of leukemia was utilised to determine the efficacy of HXP4 as a therapeutic agent. Using this immortalised cell line overexpressing HOXA9 (imHOXA9), we tested the efficacy of HXP4 in vitro. Cells were treated with HXP4 for four days and analysed. All results are expressed relative to untreated control cells. Following a 60μM dose of HXP4, no viable cells were detected as determined by trypan blue staining, suggesting that HXP4 was cytotoxic. Following treatment with a lower dose of 6μM HXP4, and re-suspension in drug-free medium for a further 6 days, cell regrowth was observed, suggesting a cytostatic effect. RT-PCR was performed to identify potential downstream targets of HXP4. Qualitative analysis showed other HOX family members to be unaffected by treatment with either HXP4 dose. A more detailed study was performed using quantitative RT-PCR with imHOXA9. Cells were treated with either 60μM HXP4, 3μM etoposide, or a combination of the two agents (H+Et) and harvested after 1, 2, and 4 hours. In general, no significant change in gene expression was observed in other HOX family members. However, HOXA1 was upregulated 3-fold when treated with HXP4, and HOXA2 was downregulated 2-fold in HXP4 and H+Et treated cells. The reasons for this are as yet unclear. HXP4 also downregulated N-RAS 3.5-fold at two hours. However, complete loss of N-RAS expression following H+Et treatment suggests that HXP4 may be more effective in combination with etoposide. CDC25 expression was slightly downregulated in HXP4-treated cells. The normal function of CDC25 is to activate CDC2 kinase in the nucleus, however in the absence of CDC25, CDC2 remains inactive leading to a delay in mitosis, supporting the proposed cytostatic mode of HXP4 action. For reasons as yet unclear, CD34 expression was upregulated 4-fold and 6-fold in HXP4 and H+Et treated cells respectively. These preliminary results suggest that HXP4 is a cytostatic agent at relatively low concentrations, with a reversible antiproliferative effect. Downstream genes regulated by disrupting the HOX-PBX interaction with HXP4 have been identified by RT-PCR, but microarray analysis will provide a more comprehensive screen for target genes. In vivo experiments are currently in progress. In conclusion blocking the interaction between HOX and PBX may represent a therapeutic strategy in leukemia treatment.

scholarly journals Open the LID: LXRα regulates ChREBPα transactivity in a target gene-specific manner through an agonist-modulated LBD-LID interaction

2019 ◽  
Author(s):  
Qiong Fan ◽  
Rikke C. Nørgaard ◽  
Ivar Grytten ◽  
Cecilie M. Ness ◽  
Christin Lucas ◽  
...  
Keyword(s):  
Target Genes ◽  
Glucose Sensing ◽  
Activation Domains ◽  
Specific Target ◽  
Glucose And Lipid Metabolism ◽  
Element Binding Protein ◽  
Underlying Mechanisms ◽  
Responsive Element

ABSTRACTThe cholesterol-sensing nuclear receptor liver X receptor (LXR) and the glucose-sensing transcription factor carbohydrate responsive element-binding protein (ChREBP) are central players in regulating glucose and lipid metabolism in liver. We have previously shown that LXR regulates ChREBP transcription and activity; however, the underlying mechanisms are unclear. In the current study, we demonstrate that LXRα and ChREBPα interact physically, and show a high co-occupancy at regulatory regions in the mouse genome. LXRα co-activates ChREBPα, and regulates ChREBP-specific target genes in vitro and in vivo. This co-activation is dependent on functional recognition elements for ChREBP, but not for LXR, indicating that ChREBPα recruits LXRα to chromatin in trans. The two factors interact via their key activation domains; ChREBPα’s low glucose inhibitory domain (LID) and the ligand-binding domain (LBD) of LXRα. While unliganded LXRα co-activates ChREBPα, ligand-bound LXRα surprisingly represses ChREBPα activity on ChREBP-specific target genes. Mechanistically, this is due to a destabilized LXRα:ChREBPα interaction, leading to reduced ChREBP-binding to chromatin and restricted activation of glycolytic and lipogenic target genes. This ligand-driven molecular switch highlights an unappreciated role of LXRα that was overlooked due to LXR lipogenesis-promoting function.

scholarly journals Down-regulated HSDL2 expression suppresses cell proliferation and promotes apoptosis in papillary thyroid carcinoma

2019 ◽  
Vol 39 (6) ◽  
Author(s):  
Jing Zeng ◽  
Xiao Ma ◽  
Jinjing Wang ◽  
Ran Liu ◽  
Yun Shao ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Papillary Thyroid Carcinoma ◽  
Thyroid Carcinoma ◽  
Target Genes ◽  
Mrna Level ◽  
Papillary Thyroid ◽  
Bioinformatics Analyses ◽  
In Vivo Experiments

Abstract Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer. Hydroxysteroid dehydrogenase like 2 (HSDL2) can regulate lipid metabolism and take part in cell proliferation. The purpose of the present study was to explore functional role of HSDL2 gene in PTC. The expression of HSDL2 protein in PTC tissues was estimated using immunohistochemistry analysis (IHC). HSDL2 mRNA level was detected through quantitative real-time polymerase chain reaction (qRT-PCR). Effects of HSDL2 gene on cell proliferation and apoptosis were assessed using the shRNA method for both in vitro and in vivo experiments. Potential target genes of HSDL2 were determined via bioinformatics analyses and Western blotting. HSDL2 was up-regulated in PTC tissues and cell lines compared with the controls (all P<0.05). Inhibiting HSDL expression could suppress PTC cell proliferation and cycle, and promote apoptosis in vitro. In vivo, the knockdown of HSDL2 gene could significantly suppress tumor growth (all P<0.05). Furthermore, AKT3, NFATc2 and PPP3CA genes might be potential targets of HSDL2 in PTC. HSDL2 expression was increased in PTC tissues and cells, which could promote tumor progression in vitro and in vivo.

Translational regulation of oskar mRNA occurs independent of the cap and poly(A) tail in Drosophila ovarian extracts

Development ◽  
1999 ◽  
Vol 126 (22) ◽  
pp. 4989-4996 ◽  
Author(s):  
Y.S. Lie ◽  
P.M. Macdonald
Keyword(s):  
Translational Regulation ◽  
Mrna Translation ◽  
Tail Length ◽  
Posterior Pole ◽  
In Vivo Experiments ◽  
Body Patterning ◽  
Underlying Mechanisms ◽  
Inhibition Studies

Translational regulation plays a prominent role in Drosophila body patterning. Progress in elucidating the underlying mechanisms has been limited by the lack of a homologous in vitro system that supports regulation. Here we show that extracts prepared from Drosophila tissues are competent for translation. Ovarian extracts, but not embryonic extracts, support the Bruno response element- and Bruno-dependent repression of oskar mRNA translation, which acts in vivo to prevent protein synthesis from transcripts not localized to the posterior pole of the oocyte. Consistent with suggestive evidence from in vivo experiments, regulation in vitro does not involve changes in poly(A) tail length. Moreover, inhibition studies strongly suggest that repression does not interfere with the process of 5′ cap recognition. Translational regulation mediated through the Bruno response elements is thus likely to occur via a novel mechanism.

scholarly journals miR-181a Ameliorates the Progression of Myasthenia Gravis by Regulating TRIM9

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Qiang Wang ◽  
Yunquan Liu ◽  
Shixiang Kuang ◽  
Ruozhao Li ◽  
Ning Weng ◽  
...  
Keyword(s):  
Myasthenia Gravis ◽  
Target Genes ◽  
Mononuclear Cells ◽  
Clinical Symptoms ◽  
Inflammatory Factors ◽  
Peripheral Blood Mononuclear ◽  
Inflammatory Microenvironment ◽  
In Vivo Experiments

Abnormally activated CD4+ T cells are considered to be an important factor in the pathogenesis of myasthenia gravis (MG). In the pathogenesis of MG, the imbalance of proinflammatory cytokines and immune cells maintains the imbalance of immune response and inflammatory microenvironment. Studies have shown that miRNA is involved in the pathogenesis of MG. In our experiment, we extracted peripheral blood mononuclear cells (PBMCs) from MG patients and detected the expression of miR-181a and TRIM9 in PBMCs by qRT-PCR. In vitro experiments were conducted to explore the regulatory mechanism of miR-181a on target genes and its influence on inflammatory factors related to MG disease. Experimental autoimmune myasthenia gravis (EAMG) model mice are established, and the effects of miR-181a on EAMG symptoms and inflammatory factors are explored through in vivo experiments. According to a total of 40 EAMG mice that were successfully modeled, all EAMG mice showed symptoms of muscle weakness; their diet was reduced; their weight gain was slow; and even weight loss occurred. In MG patients and EAMG mice, the expression of miR-181a was low and TRIM9 was highly expressed. Bioinformatics website and dual-luciferase report analysis of miR-181a had a targeting relationship with TRIM9, and miR-181a could target the expression of TRIM9. After upregulating miR-181a or interfering with TRIM9, serum miR-181a in EAMG mice was significantly upregulated; TRIM9 was significantly downregulated; its clinical symptoms were reduced; and the expression of inflammatory factors was reduced. The study finally learned that miR-181a can reduce the level of MG inflammatory factors by targeting the expression of TRIM9 and has the effect of improving the symptoms of MG.

scholarly journals Platelet extracellular vesicles enhance the proangiogenic potential of adipose-derived stem cells in vivo and in vitro

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yanan Tang ◽  
Jiayan Li ◽  
Weiyi Wang ◽  
Bingyi Chen ◽  
Jinxing Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Extracellular Vesicles ◽  
Capillary Density ◽  
Tube Formation ◽  
In Vivo Experiments ◽  
Tissue Degeneration ◽  
Underlying Mechanisms ◽  
Migration Capacity

Abstract Background Adipose-derived mesenchymal stem cells (ADSC)-based therapy is an outstanding treatment strategy for ischaemic disease. However, the therapeutic efficacy of this strategy is not ideal due to the poor paracrine function and low survival rate of ADSCs in target regions. Platelet extracellular vesicles (PEVs) are nanoparticles derived from activated platelets that can participate in communication between cells. Accumulating evidence indicates that PEVs can regulate the biological functions of several cell lines. In the present study, we aimed to investigate whether PEVs can modulate the proangiogenic potential of ADSCs in vitro and in vivo. Methods PEVs were identified using scanning electron microscope (SEM), flow cytometry (FCM) and nanoparticle tracking analysis (NTA). The CCK8 assay was performed to detect proliferation of cells. Transwell and wound healing assays were performed to verify migration capacity of cells. AnnexinV-FITC/PI apoptosis kit and live/dead assay were performed to assess ADSCs apoptosis under Cocl2-induced hypoxia condition. The underlying mechanisms by which PEVs affected ADSCs were explored using real time-PCR(RT-PCR) and Western blot. In addition, matrigel plug assays were conducted and mouse hindlimb ischaemic models were established to investigate the proangiogenic potential of PEV-treated ADSCs in vivo. Results We demonstrated that ADSC could internalize PEVs, which lead to a series of biological reactions. In vitro, dose-dependent effects of PEVs on ADSC proliferation, migration and antiapoptotic capacity were observed. Western blotting results suggested that multiple proteins such as ERK, AKT, FAK, Src and PLCγ1 kinase may contribute to these changes. Furthermore, PEVs induced upregulation of several growth factors expression in ADSCs and amplified the proliferation, migration and tube formation of HUVECs induced by ADSC conditioned medium (CM). In in vivo experiments, compared with control ADSCs, the injection of PEV-treated ADSCs resulted in more vascularization in matrigel plugs, attenuated tissue degeneration and increased blood flow and capillary density in ischaemic hindlimb tissues. Conclusion Our data demonstrated that PEVs could enhance the proangiogenic potential of ADSCs in mouse hindlimb ischaemia. The major mechanisms of this effect included the promotion of ADSC proliferation, migration, anti-apoptosis ability and paracrine secretion.

scholarly journals Human TRA2A determines influenza A virus host adaptation by regulating viral mRNA splicing

2020 ◽  
Vol 6 (25) ◽  
pp. eaaz5764 ◽  
Author(s):  
Yinxing Zhu ◽  
Ruifang Wang ◽  
Luyao Yu ◽  
Huimin Sun ◽  
Shan Tian ◽  
...  
Keyword(s):  
Avian Influenza ◽  
Messenger Rna ◽  
Influenza A ◽  
Mammalian Species ◽  
Mrna Splicing ◽  
Influenza A Viruses ◽  
Splicing Silencer ◽  
Underlying Mechanisms

Several avian influenza A viruses (IAVs) have adapted to mammalian species, including humans. To date, the mechanisms enabling these host shifts remain incompletely understood. Here, we show that a host factor, human TRA2A (huTRA2A), inhibits avian IAV replication, but benefits human IAV replication by altered regulation of viral messenger RNA (mRNA) splicing. huTRA2A depresses mRNA splicing by binding to the intronic splicing silencer motif in the M mRNA of representative avian YS/H5N1 or in the NS mRNA of representative human PR8/H1N1 virus, leading to completely opposite effects on replication of the human and avian viruses in vitro and in vivo. We also confirm that the M-334 site and NS-234/236 sites are critical for TRA2A binding, mRNA splicing, viral replication, and pathogenicity. Our results reveal the underlying mechanisms of adaptation of avian influenza virus to human hosts, and suggest rational strategies to protect public health.

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