The repressor activity of Even-skipped is highly conserved, and is sufficient to activate engrailed and to regulate both the spacing and stability of parasegment boundaries

Development ◽  
2002 ◽  
Vol 129 (19) ◽  
pp. 4411-4421 ◽  
Author(s):  
Miki Fujioka ◽  
Galina L. Yusibova ◽  
Nipam H. Patel ◽  
Susan J. Brown ◽  
James B. Jaynes
Keyword(s):  
Target Genes ◽  
Chimeric Protein ◽  
Drosophila Embryo ◽  
Dependent Manner ◽  
Coding Region ◽  
Rescue System ◽  
The Stability ◽  
Dose Dependent ◽  
Blastoderm Stage ◽  
Repressor Activity

During segmentation of the Drosophila embryo, even skipped is required to activate engrailed stripes and to organize odd-numbered parasegments. A 16 kb transgene containing the even skipped coding region can rescue normal engrailed expression, as well as all other aspects of segmentation, in even skipped null mutants. To better understand its mechanism of action, we functionally dissected the Even-skipped protein in the context of this transgene. We found that Even-skipped utilizes two repressor domains to carry out its function. Each of these domains can function autonomously in embryos when fused with the Gal4 DNA-binding domain. A chimeric protein consisting only of the Engrailed repressor domain and the Even-skipped homeodomain, but not the homeodomain alone, was able to restore function, indicating that the repression of target genes is sufficient for even skipped function at the blastoderm stage, while the homeodomain is sufficient to recognize those target genes. When Drosophila Even skipped was replaced by its homologs from other species, including a mouse homolog, they could provide substantial function, indicating that these proteins can recognize similar target sites and also provide repressor activity. Using this rescue system, we show that broad, early even skipped stripes are sufficient for activation of both odd- and even-numbered engrailed stripes. Furthermore, these ‘unrefined’ stripes organize odd-numbered parasegments in a dose-dependent manner, while the refined, late stripes, which coincide cell-for-cell with parasegment boundaries, are required to ensure the stability of the boundaries.

scholarly journals Evaluation of Hepatotoxicity with Treatment Doses of Flucytosine and Amphotericin B for Invasive Fungal Infections

2016 ◽  
Vol 2016 ◽  
pp. 1-9 ◽  
Author(s):  
Alexandra Folk ◽  
Coralia Cotoraci ◽  
Cornel Balta ◽  
Maria Suciu ◽  
Hildegard Herman ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Target Genes ◽  
Fungal Infections ◽  
Combined Therapy ◽  
Hepatic Tissue ◽  
Dependent Manner ◽  
Liver Injuries ◽  
Dose Dependent Increase ◽  
Dose Dependent ◽  
Different Doses

Invasive fungal infection is a well-known cause of morbidity and mortality in immunocompromised patients. In this study we aimed to evaluate the hepatotoxicity induced by combined therapy of flucytosine and amphotericin B, at three different doses administered to mice for 14 days: 50 mg/kg flucytosine and 300 μg/kg amphotericin B; 100 mg/kg flucytosine and 600 μg/kg amphotericin B; 150 mg/kg flucytosine and 900 μg/kg amphotericin B. Liver injuries were evaluated by analysis of optic and electron microscopy samples, changes in TNF-α, IL-6, and NF-κB inflammation markers levels of expression, and evaluation of mRNA profiles. Histological and ultrastructural analysis revealed an increase in parenchymal and portal inflammation in mice and Kupffer cells activation. Combined antifungal treatment stimulated activation of an inflammatory pathway, demonstrated by a significant dose-dependent increase of TNF-αand IL-6 immunoreactivity, together with mRNA upregulation. Also, NF-κB was activated, as suggested by the high levels found in hepatic tissue and upregulation of target genes. Our results suggest that antifungal combined therapy exerts a synergistic inflammatory activation in a dose-dependent manner, through NF-κB pathway, which promotes an inflammatory cascade during inflammation. The use of combined antifungal therapy needs to be dose limiting due to the associated risk of liver injury, especially for those patients with hepatic dysfunction.

First-in-human clinical, pharmacokinetic (PK), and pharmacodynamic (PD) phase I study of the first-in-class spliceosome inhibitor E7107 administered IV (bolus) on days 1, 8, and 15 every 28 days to patients with solid tumors

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 3508-3508
Author(s):  
F. A. Eskens ◽  
F. J. Ramos ◽  
H. Burger ◽  
M. J. de Jonge ◽  
J. Wanders ◽  
...  
Keyword(s):  
Target Genes ◽  
Expression Profiles ◽  
Systemic Exposure ◽  
Fold Increase ◽  
Pharmacokinetic Analysis ◽  
Base Line ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Grade 3 ◽  
Dose Dependent

3508 Background: E7107 is a potent first-in-class inhibitor of the spliceosome, most likely by interacting with spliceosome-associated protein-130 (SAP 130). Splicing removes intron sequences from pre-mRNA and exons are fused resulting in the formation of mature mRNA. Alternative splicing frequently encodes oncoproteins. E7107 interferes the maturation process of pre-mRNA to mRNA, with consequent changes in protein expression profiles. Methods: Objectives of this study were to explore (1) tolerability and safety profile, (2) PK, (3) PD effects on pre-mRNA splicing, and (4) anti tumor activity of E7107 administered as bolus infusion on days 1, 8, 15 of a 28-day schedule Results: 36 patients (21M/15F, median age 61yrs (45–79)) received E7107 doses of 0.6 mg/m2 (n=4), 0.9 mg/m2 (n=3), 1.3 mg/m2 (n=3), 2.0 mg/m2 (n=3), 3.0 mg/m2 (n=4), 4.5 mg/m2 (n=3) and 4.0 mg/m2 (n=16). At 4.5 mg/m2 two episodes of DLT (grade 3 and 4 diarrhea) and at 4.0 mg/m2 one episode of DLT (a combination of grade 3 nausea, vomiting and abdominal cramps) were observed. Other frequently occurring side effects were mainly gastrointestinal. The maximum tolerable dose (MTD) is 4.0 mg/m2. No complete or partial responses were observed. Pharmacokinetic analysis revealed large volume of distribution (Vss: 279 to 1369 L), high systemic clearance (CL: 111 to 253 L/hr), and moderate elimination half-life (t1/2: 5.3 to 15.1 hr). Systemic exposure on Days 1 and 15 (Cmax, AUC0-∞) increased in a dose-dependent manner. At the MTD, mRNA levels of selected target genes (TRAPPC4, SLC25A19, GTF2H1), monitored in PBMC's, showed a 15–25-fold decrease, whereas unspliced pre-mRNA levels of DNAJB1 and EIF4A1 showed a 10–25-fold increase. Notably, at days 1 and 15, modulations generally peaked at 2–6 hr after end of the infusion and almost completely recovered to base-line levels at 24–48 hr. Conclusions: The MTD for E7107 using this schedule is 4.0 mg/m2. PK is dose-dependent and reproducible within subjects. PD analysis revealed dose-dependent reversible inhibition of pre-mRNA processing of target genes, confirming proof-of-principle activity of E7107. [Table: see text]

scholarly journals Properdin: binding to C3b and stabilization of the C3b-dependent C3 convertase.

1975 ◽  
Vol 142 (4) ◽  
pp. 856-863 ◽  
Author(s):  
D T Fearon ◽  
K F Austen
Keyword(s):  
Divalent Cations ◽  
Inhibitory Effect ◽  
Alternative Complement Pathway ◽  
Dependent Manner ◽  
Complement Pathway ◽  
First Order ◽  
Alternative Complement ◽  
The Stability ◽  
Dose Dependent

A function of P in the alternative complement pathway is to prolong the first order decay of the hemolytic sites on EAC43B in a dose-dependent manner. As the number of initial convertase sites is not changed, even when activated properdin (P) increases the t1/2 10-fold or more, P acts to stabilize rather than to uncover additional sites. P binds to EAC43 to generate EAC43P in a reaction that proceeds slightly more rapidly at 15 degrees C than at 0 degrees C, but reaches the same plateau and does not require divalent cations. The presence of P on EAC43P not only stabilizes the convertase subsequently formed on that cell, but, alternatively, permits transfer to convertase sites on other cells with the stability of the recipient intermediate being dependent on the P available for transfer. The capacity of P to bind to C3b and stabilize C3B contrasts with the inhibitory effect of the C3b inactivator on formation of this amplification convertase.

scholarly journals Differential regulation of target genes by different alleles of the segmentation gene hunchback in Drosophila.

Genetics ◽  
1994 ◽  
Vol 138 (1) ◽  
pp. 125-134 ◽  
Author(s):  
M Hülskamp ◽  
W Lukowitz ◽  
A Beermann ◽  
G Glaser ◽  
D Tautz
Keyword(s):  
Transcription Factor ◽  
Zinc Finger ◽  
Target Genes ◽  
Regulatory Gene ◽  
Differential Regulation ◽  
Coding Region ◽  
Segmentation Gene ◽  
Zinc Finger Domains ◽  
Morphogenetic Gradient ◽  
Blastoderm Stage

Abstract hunchback (hb) is a key regulatory gene in the early segmentation gene hierarchy of Drosophila. It codes for a transcription factor of the Cys2-His2 zinc finger type and shows two separate zinc finger domains in its coding region. hb forms a morphogenetic gradient in the middle of the embryo that is required for setting the spatial boundaries of several target genes. We have analyzed the molecular lesions found in the different hb alleles and have studied the differential effects of these alleles on a number of such target genes. We find that in mutants in which the HB protein lacks a functional second finger domain, the regulation of the target genes Krüppel (Kr) and knirps (kni) is differentially affected. While this domain is required for the correct regulation of Kr, it is not necessary for the repression of kni. Furthermore, mutations affecting this domain lead to a decreased protein stability. The integration of the expression pattern of target genes was found to be distorted in a second class of mutants between the two finger domains which lead to gain of function or neomorphic phenotypes. The effects of these mutations were studied in detail and it was found that they fall into two classes, the first one interfering with the function of the maternal hb product, the second leading to a delayed segmentation. The function of the latter class appears to be linked to the secondary expression of hb in the parasegment 4 (PS4) stripe at blastoderm stage.

scholarly journals Sp8 regulatory function in the limb bud ectoderm

2020 ◽  
Author(s):  
Rocío Pérez-Gómez ◽  
Marc Fernández-Guerrero ◽  
Víctor Campa ◽  
Juan F. Lopez-Gimenez ◽  
Alvaro Rada-Iglesias ◽  
...  
Keyword(s):  
Target Genes ◽  
Mouse Genetics ◽  
Limb Bud ◽  
Regulatory Function ◽  
Dependent Manner ◽  
Consensus Sequences ◽  
Genome Wide ◽  
Regulatory Functions ◽  
Dual Mechanism ◽  
Dose Dependent

ABSTRACTSp8 and Sp6 are two closely related Sp genes expressed in the limb ectoderm where they regulate proximo-distal and dorso-ventral patterning. Mouse genetics revealed that they act together in a dose-dependent manner but with Sp8 making a much greater contribution. Here, we combine ChIP-seq and RNA-seq genome-wide analyses to investigate the Sp8 regulatory network and mechanism of action. We find that Sp8 predominantly binds to putative distal enhancers to activate crucial limb patterning genes, including Fgf8, En1, Sp6 and Rspo2. Sp8 exerts its regulatory function by directly binding DNA at Sp consensus sequences or indirectly through Dlx5 interaction. Overall, our work underscores Sp8 master regulatory functions and supports a model in which it cooperates with other Dlx and Sp cofactors to regulate target genes. We believe that this model could help to properly understand the molecular basis of congenital malformations.Impact SentenceIn the limb ectoderm, Sp8 regulates master genes through a dual mechanism: directly binding DNA at Sp consensus sequences and indirectly engaging through Dlx5 interaction.

scholarly journals Regulation of transcription factor SP1 by β-catenin destruction complex modulates Wnt response

2018 ◽  
Author(s):  
Rafeeq Mir ◽  
Ankita Sharma ◽  
Saurabh J. Pradhan ◽  
Sanjeev Galande
Keyword(s):  
Transcription Factor ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Target Genes ◽  
Wnt Signaling Pathway ◽  
Feedback Mechanism ◽  
Regulation Of Transcription ◽  
Dependent Manner ◽  
Factor Specificity ◽  
The Stability

AbstractThe ubiquitous transcription factor Specificity protein 1 (SP1) is heavily modified post-translationally. These modifications are critical for switching its functions and modulation of its transcriptional activity, DNA-binding and stability. However the mechanism governing the stability of SP1 by cellular signaling pathways is not well understood. Here, we provide biochemical and functional evidences that SP1 is an integral part of the Wnt signaling pathway. We identified a phosphodegron motif in SP1 that is specific to mammals. In absence of Wnt signaling, GSK3β kinase mediated phosphorylation and β-TrCP E3 ubiquitin ligase mediated ubiquitination is required to induce SP1 degradation. When Wnt signaling is on, SP1 is stabilized in β-catenin-dependent manner. SP1 directly interacts with β-catenin and Wnt signaling induces the stabilization of SP1 by impeding its interaction with β-TrCP and AXIN1, components of the destruction complex. Wnt signaling suppresses ubiquitination and subsequent proteosomal degradation of SP1. Furthermore, SP1 regulates Wnt-dependent stability of β-catenin and their mutual stabilization is critical for target gene expression, suggesting a feedback mechanism. Upon stabilization SP1 and β-catenin co-occupy the promoters of TCFL2/β-catenin target genes. Collectively, this study uncovers a direct link between SP1 and β-catenin in Wnt signaling pathway.

Epigenetic Repression of the Adaptor Molecule LAT2 by the Leukemic Fusion Protein AML1/ETO.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 987-987
Author(s):  
Jesus Duque-Afonso ◽  
Tobias Berg ◽  
Olaf Heidenreich ◽  
Michael Luebbert
Keyword(s):  
Fusion Protein ◽  
Molecular Mechanisms ◽  
Hdac Inhibitors ◽  
Histone H3 ◽  
Chimeric Protein ◽  
Dependent Manner ◽  
U937 Cells ◽  
Conditional Expression ◽  
Dose Dependent

Abstract The chromosomal translocation (8;21) fuses the hematopoietic transcription factor AML1 (RUNX1) with ETO, resulting in the leukemia-specific chimeric protein AML1/ETO. This fusion protein represses transcription by recruiting a nuclear co-repressor complex containing HDACs and DNMT1 to its target promoters. Previously, we have identified a novel in vivo AML1/ETO target gene, LAT2 (NTAL/LAB/WBSCR5), which is involved in FcεR I, c-Kit, B cell- and T cell receptor signalling. Notably, LAT2 is strongly repressed in AML1/ETO positive cells including primary AML blasts, which was confirmed by others in several large AML cohorts. We have now addressed the molecular mechanisms of AML1/ETO-mediated LAT2 repression. AML1/ETO was induced by Ponasterone A in an ecdysone-inducible system in U937 cells (9/14/18 cell line). To deplete AML1/ETO in t(8;21)-positive cells, we electroporated Kasumi-1 cells with AML1/ETO siRNA. To interfere with epigenetic modifications more directly, cells were treated with the DNMT inhibitor decitabine (DAC) and 4 different HDAC inhibitors. LAT2 expression was determined by Northern Blot, qRT-PCR and Western Blot. HDAC occupation and the histone status of the LAT2 promoter was examined by chromatin immunoprecipitation (ChIP). LAT2 mRNA was downregulated already after 4 hours of conditional expression of AML1/ETO in 9/14/18 cells, and constitutively repressed in the AML1/ETO-positive Kasumi-1 and SKNO-1 cells. siRNA-mediated AML1/ETO depletion caused a 9-fold upregulation of LAT2 in Kasumi-1 cells, suggesting a possible direct mechanism of repression. To address this question, we performed ChIP assays for the LAT2 promoter after AML1/ETO induction in 9/14/18 cells. AML1/ETO inhibited acetylation of histone H3, H3K9 and H4, but did not affect trimethylation of H3K4. These changes were associated with the recruitment of HDAC2, but not HDAC1 and HDAC3, to the LAT2 promoter. The HDAC inhibitors MS-275, SAHA, TSA and valproic acid induced LAT2 mRNA in a dose-dependent manner in AML1/ETO-expressing Kasumi-1, with MS-275 being the most efficient inhibitor. MS-275 induced LAT2 expression also in t(8;21)-positive SKNO-1, but not in AML1/ETO-negative HL60 and U937 cells. LAT2 mRNA was also upregulated in a dose-dependent manner after DAC treatment in Kasumi-1 cells. The combination of DAC and MS-275 had a synergistic effect on inhibition of cell growth, acetylation of histones H3 and H4, and re-expression of LAT2 mRNA. MS-275-mediated re-expression of LAT2 was associated with an increase in acetylation of histone H3, H3K9, H4 and trimethylation of H3K4. The increase of activating histone modifications was associated with the release of HDAC1, HDAC2 and HDAC3 from the LAT2 promoter. In conclusion, the epigenetic changes of the LAT2 promoter caused by AML1/ETO could be pharmacologically reverted by inhibition of histone acetylation.

scholarly journals Aldosterone directly affects apelin expression and secretion in adipocytes

2013 ◽  
Vol 51 (1) ◽  
pp. 37-48 ◽  
Author(s):  
He Jiang ◽  
Xiao-Ping Ye ◽  
Zhong-Yin Yang ◽  
Ming Zhan ◽  
Hai-Ning Wang ◽  
...  
Keyword(s):  
Target Genes ◽  
Direct Interaction ◽  
Serum Levels ◽  
Mapk Signaling ◽  
Dependent Manner ◽  
Beneficial Effects ◽  
Underlying Mechanisms ◽  
Plasma Apelin ◽  
Dose Dependent

There is a high incidence of metabolic syndrome among patients with primary aldosteronism (PA), which has recently been associated with an unfavorable cardiometabolic profile. However, the underlying mechanisms have not been clarified in detail. Characterizing aldosterone (Ald) target genes in adipocytes will help us to elucidate the deleterious effects associated with excess Ald. Apelin, a novel adipokine, exerts beneficial effects on obesity-associated disorders and cardiovascular homeostasis. The objective of this study was to investigate the effects of high Ald levels on apelin expression and secretion and the underlying mechanisms involved in adipocytes. In vivo, a single-dose Ald injection acutely decreased apelin serum levels and adipose tissue apelin production, which demonstrates a clear inverse relationship between the levels of plasma Ald and plasma apelin. Experiments using 3T3-L1 adipocytes showed that Ald decreased apelin expression and secretion in a time- and dose-dependent manner. This effect was reversed by glucocorticoid receptor (GR) antagonists or GR (NR3C1) knockdown; furthermore, putative HREs were identified in the apelin promoter. Subsequently, we verified that both glucocorticoids and mineralocorticoids regulated apelin expression through GR activation, although no synergistic effect was observed. Additionally, detailed potential mechanisms involved a p38 MAPK signaling pathway. In conclusion, our findings strengthen the fact that there is a direct interaction between Ald and apelin in adipocytes, which has important implications for hyperaldosteronism or PA-associated cardiometabolic syndrome and hoists apelin on the list of potent therapeutic targets for PA.

scholarly journals LncRNA–ENST00000446135 is a novel biomarker of cadmium toxicity in 16HBE cells, rats, and Cd-exposed workers and regulates DNA damage and repair

2020 ◽  
Vol 9 (6) ◽  
pp. 823-834
Author(s):  
Zhiheng Zhou ◽  
Zhijie Huang ◽  
Baoxin Chen ◽  
Qian Lu ◽  
Linlu Cao ◽  
...  
Keyword(s):  
Dna Damage ◽  
Noncoding Rna ◽  
Target Genes ◽  
Cadmium Toxicity ◽  
Dependent Manner ◽  
Dna Damage And Repair ◽  
Exposed Workers ◽  
Chromatin Immunoprecipitation Sequencing ◽  
Interfering Rna ◽  
Dose Dependent

Abstract Cadmium (Cd) and its compounds are well-known human carcinogens, but the mechanisms underlying the carcinogenesis are not well understood. This study aimed to investigate whether long noncoding RNA (LncRNA)–ENST00000446135 could serve as a novel biomarker of Cd toxicity in cells, animals, and Cd-exposed workers and regulate DNA damage and repair. LncRNA–ENST00000446135 expression increased gradually in cadmium chloride-transformed 16HBE cells. Small interfering RNA-mediated knockdown of LncRNA–ENST00000446135 inhibited the growth of DNA-damaged cells and decreased the expressions of DNA damage-related genes (ATM, ATR, and ATRIP), whereas increased the expressions of DNA repair-related genes (DDB1, DDB2, OGG1, ERCC1, MSH2, XRCC1, and BARD1). Chromatin immunoprecipitation-sequencing showed that MSH2 is a direct transcriptional target of lncRNA–ENST00000446135. Cadmium increased lncRNA–ENST00000446135 expression in the lung of Cd-exposed rats in a dose-dependent manner. A significant positive correlation was observed between blood ENST00000446135 expression and urinary/blood Cd concentrations, and there were significant correlations of LncRNA–ENST00000446135 expression with the DNA damage cell and the expressions of target genes in the lung of Cd-exposed rats and the blood of Cd-exposed workers and significantly correlated with liver and renal function in Cd-exposed workers. These results indicate that the expression of LncRNA–ENST00000446135 is upregulated and may serve as a signature for DNA damage and repair related to the epigenetic mechanisms underlying the cadmium toxicity and become a novel biomarker of cadmium toxicity.

scholarly journals RAC1 plays an essential role in estrogen receptor alpha function in breast cancer cells

Oncogene ◽  
2021 ◽  
Author(s):  
Jun Sun ◽  
Gabriel Gaidosh ◽  
Ye Xu ◽  
Adnan Mookhtiar ◽  
Na Man ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Estrogen Receptor Alpha ◽  
Target Genes ◽  
Dependent Manner ◽  
Rna Pol Ii ◽  
Receptor Alpha ◽  
Rac1 Activity ◽  
Er Positive ◽  
Dose Dependent

AbstractThe activity of Rho family GTPase protein, RAC1, which plays important normal physiological functions, is dysregulated in multiple cancers. RAC1 is expressed in both estrogen receptor alpha (ER)-positive and ER-negative breast cancer (BC) cells. However, ER-positive BC is more sensitive to RAC1 inhibition. We have determined that reducing RAC1 activity, using siRNA or EHT 1864 (a small molecule Rac inhibitor), leads to rapid ER protein degradation. RAC1 interacts with ER within the ER complex and RAC1 localizes to chromatin binding sites for ER upon estrogen treatment. RAC1 activity is important for RNA Pol II function at both promoters and enhancers of ER target genes and ER-regulated gene transcription is blocked by EHT 1864, in a dose-dependent manner. Having identified that RAC1 is an essential ER cofactor for ER protein stability and ER transcriptional activity, we report that RAC1 inhibition could be an effective therapeutic approach for ER-positive BC.

Sign in / Sign up

Export Citation Format

Share Document