Characterization of the human DYRK1A promoter and its regulation by the transcription factor E2F1

<p>Abstract</p> <p>Background</p> <p>Overexpression of the human <it>DYRK1A </it>gene due to the presence of a third gene copy in trisomy 21 is thought to play a role in the pathogenesis of Down syndrome. The observation of gene dosage effects in transgenic mouse models implies that subtle changes in expression levels can affect the correct function of the <it>DYRK1A </it>gene product. We have therefore characterized the promoter of the human <it>DYRK1A </it>gene in order to study its transcriptional regulation.</p> <p>Results</p> <p>Transcription start sites of the human <it>DYRK1A </it>gene are distributed over 800 bp within a region previously identified as an unmethylated CpG island. We have identified a new alternative noncoding 5'-exon of the <it>DYRK1A </it>gene which is located 772 bp upstream of the previously described transcription start site. Transcription of the two splicing variants is controlled by non-overlapping promoter regions that can independently drive reporter gene expression. We found no evidence of cell- or tissue-specific promoter usage, but the two promoter regions differed in their activity and their regulation. The sequence upstream of exon 1A (promoter region A) induced about 10-fold higher reporter gene activity than the sequence upstream of exon 1B (promoter region B). Overexpression of the transcription factor E2F1 increased <it>DYRK1A </it>mRNA levels in Saos2 and Phoenix cells and enhanced the activity of promoter region B three- to fourfold.</p> <p>Conclusion</p> <p>The identification of two alternatively spliced transcripts whose transcription is initiated from differentially regulated promoters regions indicates that the expression of the <it>DYRK1A </it>gene is subject to complex control mechanisms. The regulatory effect of E2F1 suggests that DYRK1A may play a role in cell cycle regulation or apoptosis.</p

A multicenter open-label phase II trial to evaluate nivolumab and ipilimumab for 2nd line therapy in elderly patients with advanced esophageal squamous cell cancer (RAMONA)

Background: Advanced esophageal squamous cell cancer (ESCC) is frequently diagnosed in elderly patients. The impact of 2nd line chemotherapy is poorly defined. Recent data demonstrated effectiveness of checkpoint inhibitors in different squamous cell carcinomas. Therefore, we assess combined nivolumab/ipilimumab as 2nd line therapy in elderly ESCC patients. Methods: RAMONA is a multicenter open-label phase II trial. The primary objective is to demonstrate a significant survival benefit of nivolumab/ipilimumab in advanced ESCC compared to historical data of standard chemotherapy. Primary endpoint is therefore overall survival (OS). Major secondary objective is the evaluation of tolerability. Time to QoL deterioration will thus be determined as key secondary endpoint. Further secondary endpoints are tumor response, PFS and safety. We aim to recruit a total of n = 75 subjects that have to be &gt; 65 years old. Eligibility is determined by the geriatric status (G8 screening and Deficit Accumulation Frailty Index (DAFI)). A safety assessment will be performed after a 3 cycle run-in phase of nivolumab (240 mg Q2W) to justify escalation for eligible patients to combined nivolumab (240 mg Q2W) and ipilimumab (1 mg/kg Q6W), while the other patients will remain on nivolumab only. RAMONA also includes translational research sub-studies to identify predictive biomarkers, including PD-1 and PD-L1 evaluation at different time points, establishment of organoid cultures and microbiome analyses for response prediction. Discussion: The RAMONA trial aims to implement checkpoint inhibitors for elderly patients with advanced ESCC as second line therapy. Novel biomarkers for checkpoint-inhibitor response are analyzed in extensive translational sub-studies. Trial registration: EudraCT Number 2017–002056-86; NCT03416244, registered: 31.1.2018

Assimilation of phytate-phosphorus by the extracellular phytase activity of tobacco (Nicotiana tabacum) is affected by the availability of soluble phytate

Phytate, the major organic phosphorus in soil, is not readily available to plants as a source of phosphorus (P). It is either complexed with cations or adsorbed to various soil components. The present study was carried out to investigate the extracellular phytase activities of tobacco (Nicotiana tabacum variety GeXin No.1) and its ability to assimilate external phytate-P. Whereas phytase activities in roots, shoots and growth media of P i-fed 14-day-old seedlings were only 1.3-4.9% of total acid phosphatase (APase) activities, P starvation triggered an increase in phytase secretion up to 914.9 mU mg -1 protein, equivalent to 18.2% of total APase activities. Much of the extracellular phytase activities were found to be root-associated than root-released. The plants were not able to utilize phytate adsorbed to sand, except when insoluble phytate salts were preformed with Mg 2+ and Ca 2+ ions for supplementation. Tobacco grew better in sand supplemented with Mg-phytate salts (31.9 mg dry weight plant -1; 0.68% w/w P concentration) than that with Ca-phytate salts (9.5 mg plant -1; 0.42%), presumably due to its higher solubility. We conclude that insolubility of soil phytate is the major constrain for its assimilation. Improving solubility of soil phytate, for example, by enhancement of citrate secretion, may be a feasible approach to improve soil phytate assimilation. © Springer 2006.postprin

Web Searching: A Quality Measurement Perspective

The purpose of this paper is to describe various quality measures for search engines and to ask whether these are suitable. We especially focus on user needs and their use of web search engines. The paper presents an extensive literature review and a first quality measurement model, as well. Findings include that search engine quality can not be measured by just retrieval effectiveness (the quality of the results), but should also consider index quality, the quality of the search features and search engine usability. For each of these sections, empirical results from studies conducted in the past, as well as from our own research are presented. These results have implications for the evaluation of search engines and for the development of better search systems that give the user the best possible search experience

Characterization of the human promoter and its regulation by the transcription factor E2F1-7

Onserved in the mouse, dog, and platypus genomes and comprises a full CRE sequence (with one mismatch to the canonical octamer) and a CRE half-site (boxed). B) Forskolin stimulates phosphorylation of CREB on Ser133. PC12 cells were treated for 30 min with forskolin (10 μM) or DMSO (Ctrl) before nuclear extracts were prepared. Phospho-Ser133 in CREB was detected by Western blot analysis with the help of a phosphospecific antibody. Migration of molecular mass standards is indicated in kDa. C) Northern blot. PC12 cells were treated for 24 h with forskolin (10 μM) or DMSO as the vehicle (Ctrl) before total RNA was isolated and subjected to Northern blot analysis with probes for DYRK1A, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and . Duplicate lanes contain RNA samples from parallel plates. Migration of the ribosomal RNAs is indicated in the right margin (28S, 18S). D) Luciferase assays. The indicated cell lines (PC12, PC3, Saos2) were transfected with the reporter gene constructs as schematically depicted. Luciferase activity was determined from cells treated with forskolin (10 μM) for 24 h and from untreated cells. Data were normalized to β-galactosidase activity and are presented as fold induction relative to untreated cells. Bars reflect the means +/- SD of 3 independent experiments (Saos2, PC3) or the means of duplicate wells for the experiment with PC12 cells.<p><b>Copyright information:</b></p><p>Taken from "Characterization of the human promoter and its regulation by the transcription factor E2F1"</p><p>http://www.biomedcentral.com/1471-2199/9/30</p><p>BMC Molecular Biology 2008;9():30-30.</p><p>Published online 26 Mar 2008</p><p>PMCID:PMC2292204.</p><p></p

Characterization of the human promoter and its regulation by the transcription factor E2F1-3

Iferase cDNA in the vector pGL3basic. The position of the promoter fragments relative to TSS B (+1) is indicated. Segments upstream of TSS A and TSS B are highlighted in red and blue, respectively. Saos2 cells were co-transfected with the indicated reporter gene constructs and a β-galactosidase reporter control plasmid. Cells were lysed 48 h after transfection, and luciferase activities were determined from duplicate wells. Data were normalized to β-galactosidase activities and are presented as the ratio relative to the activity of the -742 construct. The diagram integrates results of 2–6 independent experiments. Bars reflect means +/- SD. The inset shows a magnified representation of the weakest signals.<p><b>Copyright information:</b></p><p>Taken from "Characterization of the human promoter and its regulation by the transcription factor E2F1"</p><p>http://www.biomedcentral.com/1471-2199/9/30</p><p>BMC Molecular Biology 2008;9():30-30.</p><p>Published online 26 Mar 2008</p><p>PMCID:PMC2292204.</p><p></p