Genetic evidence for the transcriptional-activating function of Homothorax during adult fly development

Development ◽  
2001 ◽  
Vol 128 (18) ◽  
pp. 3405-3413 ◽  
Author(s):  
Adi Inbal ◽  
Naomi Halachmi ◽  
Charna Dibner ◽  
Dale Frank ◽  
Adi Salzberg
Keyword(s):  
Transcriptional Activity ◽  
Target Genes ◽  
Genetic Evidence ◽  
In Vivo Studies ◽  
Loss Of Function ◽  
Native Form ◽  
Downstream Target ◽  
Activating Function

Homothorax (HTH) is a homeobox-containing protein, which plays multiple roles in the development of the embryo and the adult fly. HTH binds to the homeotic cofactor Extradenticle (EXD) and translocates it to the nucleus. Its function within the nucleus is less clear. It was shown, mainly by in vitro studies, that HTH can bind DNA as a part of ternary HTH/EXD/HOX complexes, but little is known about the transcription regulating function of HTH-containing complexes in the context of the developing fly. Here we present genetic evidence, from in vivo studies, for the transcriptional-activating function of HTH. The HTH protein was forced to act as a transcriptional repressor by fusing it to the Engrailed (EN) repression domain, or as a transcriptional activator, by fusing it to the VP16 activation domain, without perturbing its ability to translocate EXD to the nucleus. Expression of the repressing form of HTH in otherwise wild-type imaginal discs phenocopied hth loss of function. Thus, the repressing form was working as an antimorph, suggesting that normally HTH is required to activate the transcription of downstream target genes. This conclusion was further supported by the observation that the activating form of HTH caused typical hth gain-of-function phenotypes and could rescue hth loss-of-function phenotypes. Similar results were obtained with XMeis3, the Xenopus homologue of HTH, extending the known functional similarity between the two proteins. Competition experiments demonstrated that the repressing forms of HTH or XMeis3 worked as true antimorphs competing with the transcriptional activity of the native form of HTH. We also describe the phenotypic consequences of HTH antimorph activity in derivatives of the wing, labial and genital discs. Some of the described phenotypes, for example, a proboscis-to-leg transformation, were not previously associated with alterations in HTH activity. Observing the ability of HTH antimorphs to interfere with different developmental pathways may direct us to new targets of HTH. The HTH antimorph described in this work presents a new means by which the transcriptional activity of the endogenous HTH protein can be blocked in an inducible fashion in any desired cells or tissues without interfering with nuclear localization of EXD.

n-Propyl gallate activates hypoxia-inducible factor 1 by modulating intracellular oxygen-sensing systems

2008 ◽  
Vol 411 (1) ◽  
pp. 97-105 ◽  
Author(s):  
Motohide Kimura ◽  
Satoshi Takabuchi ◽  
Tomoharu Tanaka ◽  
Miyahiko Murata ◽  
Kenichiro Nishi ◽  
...  
Keyword(s):  
Propyl Gallate ◽  
Transcriptional Activity ◽  
Target Genes ◽  
Cultured Cells ◽  
Oxygen Sensing ◽  
Hypoxia Inducible Factor ◽  
Downstream Target ◽  
Hypoxia Inducible Factor 1

HIF-1 (hypoxia-inducible factor 1) is a master regulator of cellular adaptive responses to hypoxia. The expression and transcriptional activity of the HIF-1α subunit is stringently controlled by intracellular oxygen tension through the action of prolyl and asparaginyl hydroxylases. In the present study we demonstrate that PG (n-propyl gallate) activates HIF-1 and expression of its downstream target genes under normoxic conditions in cultured cells and in mice. The stability and transcriptional activity of HIF-1α are increased by PG. PG treatment inhibits the interaction between HIF-1α and VHL (von Hippel–Lindau protein) and promotes the interaction between HIF-1α and p300, indicating that PG inhibits the activity of both prolyl and asparaginyl HIF-1α hydroxylases. We conclude that PG activates HIF-1 and enhances the resultant gene expression by directly affecting the intracellular oxygen sensing system in vitro and in vivo and that PG represents a lead compound for the development of a non-toxic activator of HIF-1.

scholarly journals MODL-16. ABEMACICLIB, A SELECTIVE CDK4/6 INHIBITOR, RESTRICTS GROWTH OF PEDIATRIC GLIAL-LINEAGE TUMORS IN VITRO AND IN VIVO

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii414-iii414
Author(s):  
Muh-Lii Liang ◽  
Tsung-Han Hsieh ◽  
Tai-Tong Wong
Keyword(s):  
Dna Repair ◽  
Therapeutic Strategy ◽  
Target Genes ◽  
Rna Seq ◽  
Downstream Target ◽  
Rb Phosphorylation ◽  
Glial Lineage

Abstract BACKGROUND Glial-lineage tumors constitute a heterogeneous group of neoplasms, comprising gliomas, oligodendrogliomas, and ependymomas, which account for 40%–50% of all pediatric central nervous system tumors. Advances in modern neuro-oncological therapeutics are aimed at improving neoadjuvant chemotherapy and deferring radiotherapy because radiation exposure may cause long-term side effects on the developing brain in young children. Despite aggressive treatment, more than half the high-grade gliomas (pHGGs) and one-third of ependymomas exhibit recurrence within 2 years of initial treatment. METHODS By using integrated bioinformatics and through experimental validation, we found that at least one gene among CCND1, CDK4, and CDK6 was overexpressed in pHGGs and ependymomas. RESULTS The use of abemaciclib, a highly selective CDK4/6 inhibitor, effectively inhibited cell proliferation and reduced the expression of cell cycle–related and DNA repair–related gene expression, which was determined through RNA-seq analysis. The efficiency of abemaciclib was validated in vitro in pHGGs and ependymoma cells and in vivo by using subcutaneously implanted ependymoma cells from patient-derived xenograft (PDX) in mouse models. Abemaciclib demonstrated the suppression of RB phosphorylation, downstream target genes of E2F, G2M checkpoint, and DNA repair, resulting in tumor suppression. CONCLUSION Abemaciclib showed encouraging results in preclinical pediatric glial-lineage tumors models and represented a potential therapeutic strategy for treating challenging tumors in children.

scholarly journals Mediator complex subunit Med19 binds directly GATA transcription factors and is required with Med1 for GATA-driven gene regulation in vivo

2020 ◽  
Vol 295 (39) ◽  
pp. 13617-13629
Author(s):  
Clément Immarigeon ◽  
Sandra Bernat-Fabre ◽  
Emmanuelle Guillou ◽  
Alexis Verger ◽  
Elodie Prince ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Target Genes ◽  
Direct Interaction ◽  
Mediator Complex ◽  
Loss Of Function ◽  
Transcriptional Responses ◽  
Rna Pol Ii ◽  
Evolutionarily Conserved

The evolutionarily conserved multiprotein Mediator complex (MED) serves as an interface between DNA-bound transcription factors (TFs) and the RNA Pol II machinery. It has been proposed that each TF interacts with a dedicated MED subunit to induce specific transcriptional responses. But are these binary partnerships sufficient to mediate TF functions? We have previously established that the Med1 Mediator subunit serves as a cofactor of GATA TFs in Drosophila, as shown in mammals. Here, we observe mutant phenotype similarities between another subunit, Med19, and the Drosophila GATA TF Pannier (Pnr), suggesting functional interaction. We further show that Med19 physically interacts with the Drosophila GATA TFs, Pnr and Serpent (Srp), in vivo and in vitro through their conserved C-zinc finger domains. Moreover, Med19 loss of function experiments in vivo or in cellulo indicate that it is required for Pnr- and Srp-dependent gene expression, suggesting general GATA cofactor functions. Interestingly, Med19 but not Med1 is critical for the regulation of all tested GATA target genes, implying shared or differential use of MED subunits by GATAs depending on the target gene. Lastly, we show a direct interaction between Med19 and Med1 by GST pulldown experiments indicating privileged contacts between these two subunits of the MED middle module. Together, these findings identify Med19/Med1 as a composite GATA TF interface and suggest that binary MED subunit–TF partnerships are probably oversimplified models. We propose several mechanisms to account for the transcriptional regulation of GATA-targeted genes.

MiR-30a-5p Downregulation Induces Neuronal Autophagy by Activating AMPK After 2856MHz Microwave Radiation

2021 ◽  
Author(s):  
Yanhui Hao ◽  
Wenchao Li ◽  
Hui Wang ◽  
Jing Zhang ◽  
Haoyu Wang ◽  
...  
Keyword(s):  
Microwave Radiation ◽  
Hippocampal Neurons ◽  
Target Genes ◽  
Neuronal Damage ◽  
Luciferase Reporter ◽  
Potential Health ◽  
Downstream Target ◽  
Neuronal Autophagy

Abstract Background With the development of science and technology, microwaves are being widely used. More and more attention has been paid to the potential health hazards of microwave exposure. The regulation of miR-30a-5p (miR-30a) on autophagy is involved in the pathophysiological process of many diseases. Our previous study found that 30 mW/cm2 microwave radiation could reduce miR-30a expression and activate neuronal autophagy in rat hippocampus. However, the roles played by miR-30a in microwave-induced neuronal autophagy and related mechanisms remain largely unexplored. Results In the present study, we established neuronal damage models by exposing rat hippocampal neurons and rat adrenal pheochromocytoma (PC12) cell-derived neuron-like cells to 30 mW/cm2 microwave, which resulted in miR-30a downregulation and autophagy activation in vivo and in vitro. Bioinformatics analysis was conducted, and Beclin1, Prkaa2, Irs1, Pik3r2, Rras2, Ddit4, Gabarapl2 and autophagy-related gene 12 (Atg12) were identified as potential downstream target genes of miR-30a involved in regulating autophagy. Based on our previous findings that microwave radiation can cause a neuronal energy metabolism disorder, Prkaa2, encoding adenosine 5’-monophosphate-activated protein kinase α2 (AMPKα2, an important catalytic subunit of energy sensor AMPK), was selected for further analysis. Dual-luciferase reporter assay results showed that Prkaa2 is a downstream target gene of miR-30a. Microwave radiation increased the expression and phosphorylation (Thr172) of AMPKα both in vivo and in vitro. Moreover, the transduction of cells with miR-30a mimics suppressed AMPKα2 expression, inhibited AMPKα (Thr172) phosphorylation and reduced autophagy flux in neuron-like cells. Importantly, miR-30a mimics abolished microwave-activated autophagy and inhibited microwave-induced AMPKα (Thr172) phosphorylation. Conclusions AMPKα2 was a newly founded downstream gene of miR-30a involved in autophagy regulation, and miR-30a downregulation after microwave radiation could promote neuronal autophagy by increasing AMPKα2 expression and activating AMPK signaling.

scholarly journals NFAT2-HDAC1 signaling contributes to the malignant phenotype of glioblastoma

2019 ◽  
Vol 22 (1) ◽  
pp. 46-57 ◽  
Author(s):  
Yifu Song ◽  
Yang Jiang ◽  
Dongxia Tao ◽  
Zixun Wang ◽  
Run Wang ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Transcriptional Activity ◽  
The Cancer Genome Atlas ◽  
Clinical Samples ◽  
Nuclear Factor ◽  
Malignant Phenotype ◽  
Histone Deacetylase 1 ◽  
Downstream Target

Abstract Background Deregulation of the nuclear factor of activated T cell (NFAT) pathway has been reported in several human cancers. Particularly, NFAT2 is involved in the malignant transformation of tumor cells and is identified as an oncogene. However, the role of NFAT2 in glioblastoma (GBM) is largely unknown. Methods The expression and prognostic value of NFAT2 were examined in the databases of the Repository of Molecular Brain Neoplasia Data and The Cancer Genome Atlas (TCGA) and clinical samples. The functional effects of silencing or overexpression of NFAT2 were evaluated in glioma stem cell (GSC) viability, invasion, and self-renewal in vitro and in tumorigenicity in vivo. The downstream target of NFAT2 was investigated. Results High NFAT2 expression was significantly associated with mesenchymal (MES) subtype and recurrent GBM and predicted poor survival. NFAT2 silencing inhibited the invasion and clonogenicity of MES GSC-enriched spheres in vitro and in vivo. NFAT2 overexpression promoted tumor growth and MES differentiation of GSCs. A TCGA database search showed that histone deacetylase 1 (HDAC1) expression was significantly correlated with that of NFAT2. NFAT2 regulates the transcriptional activity of HDAC1. Rescue of HDAC1 in NFAT2-knockdown GSCs partially restored tumor growth and MES phenotype. Loss of NFAT2 and HDAC1 expression resulted in hyperacetylation of nuclear factor-kappaB (NF-κB), which inhibits NF-κB–dependent transcriptional activity. Conclusion Our findings suggest that the NFAT2-HDAC1 pathway might play an important role in the maintenance of the malignant phenotype and promote MES transition in GSCs, which provide potential molecular targets for the treatment of GBMs.

scholarly journals Modulating PKCα Activity to Target Wnt/β-Catenin Signaling in Colon Cancer

Cancers ◽  
2019 ◽  
Vol 11 (5) ◽  
pp. 693 ◽  
Author(s):  
Sébastien Dupasquier ◽  
Philippe Blache ◽  
Laurence Picque Lasorsa ◽  
Han Zhao ◽  
Jean-Daniel Abraham ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Transcriptional Activity ◽  
Target Genes ◽  
Tumor Development ◽  
Orphan Receptor ◽  
Nuclear Accumulation ◽  
Kinase C ◽  
Extracellular Signal

Inactivating mutations of the tumor suppressor Adenomatosis Polyposis Coli (APC), which are found in familial adenomatosis polyposis and in 80% of sporadic colorectal cancers (CRC), result in constitutive activation of the Wnt/β-catenin pathway and tumor development in the intestine. These mutations disconnect the Wnt/β-catenin pathway from its Wnt extracellular signal by inactivating the APC/GSK3-β/axin destruction complex of β-catenin. This results in sustained nuclear accumulation of β-catenin, followed by β-catenin-dependent co-transcriptional activation of Wnt/β-catenin target genes. Thus, mechanisms acting downstream of APC, such as those controlling β-catenin stability and/or co-transcriptional activity, are attractive targets for CRC treatment. Protein Kinase C-α (PKCα) phosphorylates the orphan receptor RORα that then inhibits β-catenin co-transcriptional activity. PKCα also phosphorylates β-catenin, leading to its degradation by the proteasome. Here, using both in vitro (DLD-1 cells) and in vivo (C57BL/6J mice) PKCα knock-in models, we investigated whether enhancing PKCα function could be beneficial in CRC treatment. We found that PKCα is infrequently mutated in CRC samples, and that inducing PKCα function is not deleterious for the normal intestinal epithelium. Conversely, di-terpene ester-induced PKCα activity triggers CRC cell death. Together, these data indicate that PKCα is a relevant drug target for CRC treatment.

scholarly journals The cerebral cavernous malformation proteins CCM2L and CCM2 prevent the activation of the MAP kinase MEKK3

2015 ◽  
Vol 112 (46) ◽  
pp. 14284-14289 ◽  
Author(s):  
Xavier Cullere ◽  
Eva Plovie ◽  
Paul M. Bennett ◽  
Calum A. MacRae ◽  
Tanya N. Mayadas
Keyword(s):  
Endothelial Cells ◽  
Cavernous Malformation ◽  
Body Axis ◽  
Cerebral Cavernous Malformations ◽  
Loss Of Function ◽  
Cavernous Malformations ◽  
Downstream Target ◽  
Density Flow

Three genes, CCM1, CCM2, and CCM3, interact genetically and biochemically and are mutated in cerebral cavernous malformations (CCM). A recently described member of this CCM family of proteins, CCM2-like (CCM2L), has high homology to CCM2. Here we show that its relative expression in different tissues differs from that of CCM2 and, unlike CCM2, the expression of CCM2L in endothelial cells is regulated by density, flow, and statins. In vitro, both CCM2L and CCM2 bind MEKK3 in a complex with CCM1. Both CCM2L and CCM2 interfere with MEKK3 activation and its ability to phosphorylate MEK5, a downstream target. The in vivo relevance of this regulation was investigated in zebrafish. A knockdown of ccm2l and ccm2 in zebrafish leads to a more severe “big heart” and circulation defects compared with loss of function of ccm2 alone, and also leads to substantial body axis abnormalities. Silencing of mekk3 rescues the big heart and body axis phenotype, suggesting cross-talk between the CCM proteins and MEKK3 in vivo. In endothelial cells, CCM2 deletion leads to activation of ERK5 and a transcriptional program that are downstream of MEKK3. These findings suggest that CCM2L and CCM2 cooperate to regulate the activity of MEKK3.

pVHL acts as a downstream target of E2F1 to suppress E2F1 activity

2013 ◽  
Vol 457 (1) ◽  
pp. 185-195 ◽  
Author(s):  
Wei Ji ◽  
Jing Wang ◽  
Wei Zhang ◽  
Xing Liu ◽  
Gang Ouyang ◽  
...  
Keyword(s):  
Binding Site ◽  
Transcriptional Activity ◽  
Downstream Target

pVHL is a downstream target of E2F1, which harbours an E2F1-binding site in its promoter. Moreover, pVHL binds to E2F1 in vitro and in vivo, resulting in inhibition of E2F1 transcriptional activity.

scholarly journals A novel treatment strategy for lapatinib resistance in a subset of HER2-amplified gastric cancer

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Gang Ning ◽  
Qihui Zhu ◽  
Wonyoung Kang ◽  
Hamin Lee ◽  
Leigh Maher ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Lines ◽  
Target Genes ◽  
Kinase Inhibitor ◽  
Pharmacological Study ◽  
Mapk Signaling ◽  
Her2 Overexpression ◽  
Loss Of Function

Abstract Background Gastric cancer (GC) is one of the leading causes of cancer-related deaths worldwide. Human epidermal growth factor receptor 2 (HER2) amplification occurs in approximately 13–23% of all GC cases and patients with HER2 overexpression exhibit a poor prognosis. Lapatinib, a dual EGFR/HER2 tyrosine kinase inhibitor, is an effective agent to treat HER2-amplified breast cancer but it failed in gastric cancer (GC) clinical trials. However, the molecular mechanism of lapatinib resistance in HER2-amplified GC is not well studied. Methods We employed an unbiased, genome-scale screening with pooled CRISPR library on HER2-amplified GC cell lines to identify genes that are associated with resistance to lapatinib. To validate the candidate genes, we applied in vitro and in vivo pharmacological tests to confirm the function of the target genes. Results We found that loss of function of CSK or PTEN conferred lapatinib resistance in HER2-amplified GC cell lines NCI-N87 and OE19, respectively. Moreover, PI3K and MAPK signaling was significantly increased in CSK or PTEN null cells. Furthermore, in vitro and in vivo pharmacological study has shown that lapatinib resistance by the loss of function of CSK or PTEN, could be overcome by lapatinib combined with the PI3K inhibitor copanlisib and MEK inhibitor trametinib. Conclusions Our study suggests that loss-of-function mutations of CSK and PTEN cause lapatinib resistance by re-activating MAPK and PI3K pathways, and further proved these two pathways are druggable targets. Inhibiting the two pathways synergistically are effective to overcome lapatinib resistance in HER2-amplified GC. This study provides insights for understanding the resistant mechanism of HER2 targeted therapy and novel strategies that may ultimately overcome resistance or limited efficacy of lapatinib treatment for subset of HER2 amplified GC.

scholarly journals TMED3/RPS15A Axis promotes the development and progression of osteosarcoma

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Wei Xu ◽  
Yifan Li ◽  
Xiaojian Ye ◽  
Yunhan Ji ◽  
Yu Chen ◽  
...  
Keyword(s):  
Biological Function ◽  
Profile Analysis ◽  
Tumor Model ◽  
Underlying Mechanism ◽  
Molecular Therapy ◽  
Loss Of Function ◽  
Osteosarcoma Cells ◽  
Downstream Target

Abstract Background Osteosarcoma is a primary malignant tumor that mainly affects children and young adults. Transmembrane emp24 trafficking protein 3 (TMED3) may be involved in the regulation of malignant cancer behaviors. However, the role of TMED3 in osteosarcoma remains mysterious. In this study, the potential biological function and underlying mechanism of TMED3 in progression of osteosarcoma was elaborated. Methods The expression of TMED3 in osteosarcoma was analyzed by immunohistochemical staining. The biological function of TMED3 in osteosarcoma was determined through loss-of-function assays in vitro. The effect of TMED3 downregulation on osteosarcoma was further explored by xenograft tumor model. The molecular mechanism of the regulation of TMED3 on osteosarcoma was determined by gene expression profile analysis. Results The expression of TMED3 in osteosarcoma tissues was significantly greater than that in matched adjacent normal tissues. Knockdown of TMED3 inhibited the progression of osteosarcoma by suppressing proliferation, impeding migration and enhancing apoptosis in vitro. We further validated that knockdown of TMED3 inhibited osteosarcoma generation in vivo. Additionally, ribosomal protein S15A (RPS15A) was determined as a potential downstream target for TMED3 involved in the progression of osteosarcoma. Further investigations elucidated that the simultaneous knockdown of RPS15A and TMED3 intensified the inhibitory effects on osteosarcoma cells. Importantly, knockdown of RPS15A alleviated the promotion effects of TMED3 overexpression in osteosarcoma cells. Conclusions In summary, these findings emphasized the importance of TMED3/RPS15A axis in promoting tumor progression, which may be a promising candidate for molecular therapy of osteosarcoma.

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