105,965 research outputs found
HINT-ATAC
No full textTransposase-Accessible Chromatin followed by sequencing (ATAC-seq) is a simple protocol for detection of
open chromatin. Computational footprinting, the search for regions with depletion of cleavage events due to
transcription factor binding, is poorly understood for ATAC-seq. We propose the first footprinting method
considering ATAC-seq protocol artefact. HINT-ATAC uses a position dependency model to learn the cleavage
preferences of the transposase. We observe strand-specific cleavage patterns around transcription factor
binding sites, which are determined by local nucleosome architecture. By incorporating all these biases,
HINT-ATAC is able to significantly outperform competing methods in the prediction of transcription factor
binding sites with footprints.</p
Current status of numerical flow prediction for separated nozzle flows
No full textThe European ‘Flow Separation Control Device’ group (FSCD) organized in collaboration with the French ‘Aérodynamiques des tuyères et Arrière-Corps’ group (ATAC) a CFD workshop with test cases on different nozzle flow topics. One of these test cases (1A) was managed by the German Aerospace Center (DLR) and Astrium ST. The objective was to compute the flow inside a strongly over-expanded truncated ideal contour nozzle with respect to the prediction of location and shape of the flow separation, the oblique shock and the Mach disc. Experimental data were provided by DLR.
An introduction to the test facility and the experimental setup is given. The numerical results are evaluated and compared to test data. A concluding synthesis illustrates the current status of nozzle flow computation
Functional characterization of C-class chemokine ATAC in vivo
No full text1\. EINLEITUNG 1 1.1 ÜBERBLICK ÜBER DAS IMMUNSYSTEM 1 1.2 DIE ANTIGEN-
PRÄSENTIERENDEN ZELLEN: DENDRITISCHE ZELLEN 2 1.2.1 Kreuzpräsentation von
Dendritischen Zellen 4 1.3 DIE CD8+ T-ZELL-VERMITTELTE IMMUNITÄT 6 1.3.1
Periphere Toleranz von CD8+ T-Zellen 7 1.3.2 CD8+ T-Zell-vermittelte
Zytotoxizität 9 1.3.3 Die Aktivierungsmarker auf CD8+ T-Zellen 10 1.4
CHEMOKINE 12 1.4.1 C-Klasse Chemokin: ATAC 12 1.4.2 Spezifischer ATAC
Rezeptor: XCR1 14 1.4.3 ATAC Funktionen 17 2\. ZIELSETZUNG DER ARBEIT 20 3\.
MATERIAL UND METHODEN 22 3.1 VERWENDETE GERÄTE, CHEMIKALIEN UND MATERIALIEN 22
3.1.1 Geräte 22 3.1.2 Chemikalien und Materialien 22 3.1.3 Puffer und Lösungen
für SDS-Page 22 3.1.4 Lösungen für Western Blot und Detektion 23 3.1.5 Puffer
für Affinitätschromatographie 24 3.1.6 Medien und Lösungen für die
Bakterienkultur 24 3.1.7 Medien und Lösungen für die Zellkultur 25 3.1.8
Lösungen für die Zellisolierung und FACS 26 3.2 MOLEKULARBIOLOGISCHE METHODEN
26 3.2.1 Quantitative Reverse-Transkriptions-PCR (qRT-PCR) 26 3.3 EXPRESSION
VON REKOMBINANTEM ATAC 28 3.3.1 Generierung des Expressionsvektors 28 3.3.2
Affinitätsreinigung von ATAC 29 3.3.3 SDS-Polyacrylamidgelelektorphorese 30
3.3.4 Western Blot 30 3.3.5 Bestimmung der Proteinkonzentration 31 3.4
ZELLBIOLOGISCHE METHODEN 31 3.4.1 Kultivierung von permanenten Zelllinien 31
3.4.2 Isolation von murinen Splenozyten aus der Milz 32 3.4.3 Aktivierung von
T-Zellen in vitro 32 3.4.4 Isolierung von murinen Zellen aus Lymphknoten 33
3.4.5 Isolierung von murinen Zellen aus Lebern 33 3.4.6 Isolierung von murinen
Zellen aus Lungen 33 3.4.7 Isolierung von murinen Zellen aus dem Knochenmark
34 3.4.8 Anreicherung von T-Zellen mittels Nylonwolle 34 3.4.9 Gewinnung von
Dendritischen Zellen aus Milzen 35 3.4.10 Anreicherung von Milzzellen durch
magnetische Zellsortierung (MACS) 35 3.4.11 Zellproliferations-Assay 36 3.4.12
Apoptose-Assay 37 3.5 DURCHFLUSSZYTOMETRIE 37 3.5.1 Zellmarkierung 37 3.5.2
Messung am Durchflusszytometer 41 3.5.3 Auswertung der Messung am
Durchflusszytometer 43 3.6 TIEREXPERIMENTELLE METHODEN 43 3.6.1 Verwendete
Tiere 43 3.6.2 Adoptiver Transfer von OT-I-Zellen 44 3.6.3 Immunisierung der
Mäuse 44 3.6.4 Zytotoxischer Assay in vivo 44 3.6.5 Chemotaktische
Untersuchung in vivo 45 4\. ERGEBNISSE 47 4.1 PRODUKTION VON MURINEM ATAC-
PROTEIN MIT HILFE EINES SUMO-EXPRESSIONSSYSTEMS 47 4.2 EXPRESSION VON XCR1 IN
DER MILZ 50 4.2.1 Expression von XCR1 mRNA in verschieden Zellpopulationen 50
4.3 ROLLE VON ATAC IN DER T-DZ-INTERAKTION IN VITRO 52 4.4 FUNKTIONELLE
UNTERSUCHUNG VON ATAC IN VIVO DURCH EIN OVALBUMIN-SPEZIFISCHES OT-I-ZELL-
SYSTEM 56 4.4.1 Kinetik der ATAC-Expression in vivo 56 4.4.2 Expression von
XCR1 in naiven und aktivierten Milzzellen 60 4.4.3 ATAC ist ein wichtiger
Regulator der T-DZ-Interaktion in vivo 62 4.4.4 Proliferation von ATAC KO und
WT OT-I-Zellen nach Gabe von Ovalbumin in vivo 63 4.4.5 Proliferation und
Apoptose von OT-I-Zellen nach Gabe von αDEC-205:OVA in vivo 64 4.4.6 Expansion
von OT-I-Zellen 3 Tagen und 5 Tagen nach Aktivierung in vivo 66 4.4.7
Überleben von OT-I-Zellen 12 Tagen nach Aktivierung in vivo 69 4.4.8 Rolle von
ATAC bei CD8+ T-Zell-vermittelter Zytotoxizität 73 4.5 ATAC IST EIN CHEMOKIN
FÜR XCR1-TRAGENDE ZELLEN 77 4.6 DER PHÄNOTYP VON ATAC KO C57BL/6 MÄUSEN 80
4.6.1 T-Zell-Populationen in der Milz 80 4.6.2 Der Phänotyp von über ein Jahr
alten ATAC KO C57BL/6 Mäusen im Vergleich zu WT Mäusen 82 5\. DISKUSSION 89
5.1 IDENTIFIZIERUNG VON ATAC-TARGET-ZELLEN 89 5.2 DIE PUTATIVE ROLLE VON ATAC
WÄHREND DER KREUZPRÄSENTATION 90 5.2.1 Die Rolle von ATAC während der T-DZ-
Interaktionsphase 90 5.2.2 Die Rolle von ATAC während der Expansion von CD8+
T-Zellen 92 5.2.3 Die Rolle von ATAC für das Überleben von CD8+ T-Zellen nach
Aktivierung 94 5.2 DER EINFLUSS VON ATAC AUF DIE CD8+ T-ZELL-VERMITTELTE
ZYTOTOXIZITÄT 95 5.3 DIE CHEMOTAKTISCHE WIRKUNG VON ATAC IN VIVO 96 5.4
PHÄNOTYP VON NAIVEN ATAC-DEFIZIENTEN C57BL/6 MÄUSEN 97 6\.
ZUSAMMENFASSUNG/SUMMARY 100 6.1 ZUSAMMENFASSUNG 100 6.2 SUMMARY 101 7\.
DANKSAGUNG 102 8\. LITERATURVERZEICHTNIS 104 9\. ANHANG 123 9.1 LEBENSLAUF 123
9.2 BESCHEINIGUNG 124ATAC ist ein Mitglied der Chemokin-Familie, welches hauptsächlich von
aktivierten CD8+ T-Zellen und NK-Zellen freigesetzt wird. Das
Expressionsmuster des ATAC-Rezeptors XCR1 war jedoch umstritten, weshalb die
physiologische Funktion von ATAC bislang unklar blieb. In dieser Arbeit wurde
die Rolle von ATAC und XCR1 im murinen System untersucht. Um die
XCR1-exprimierenden Zellen in der Milz zu definieren, wurden ruhende und
aktivierte Milzzellen aus unterschiedlichen Mausstämmen isoliert und durch
quantitative RT-PCR analysiert. XCR1 konnte eindeutig in CD8+ dendritischen
Zellen (DZ), geringfügig in CD8- DZ, aber nicht in den anderen
Zellpopulationen nachgewiesen werden. Der Einfluss von ATAC auf die antigen-
spezifische Aktivierung von CD8+ T-Zellen wurde untersucht, indem WT oder ATAC
KO OT-I-Zellen adoptiv transferiert und CD8+ DZ der syngenen Rezipienten über
das Molekül DEC-205 selektiv mit Antigen (DEC205:OVA) beladen wurden. In
diesem Modell wurde eine sehr frühe (ab 6 h) und dauerhafte (bis 48 h) ATAC-
Sekretion in WT OT-I-Zellen beobachtet. In Abwesenheit des ATAC-Signals
erhöhte sich die Expression von CD69 und 4-1BB auf transferierten OT-I-Zellen
18-24 h nach Stimulation, aber ein deutlicher Einfluss auf die Proliferation
und Apoptose der transferierten OT-I-Zellen 2-6 Tage nach Stimulation konnte
nicht festgestellt werden. Dies galt sowohl für eine Stimulation mit DEC:OVA
alleine („Toleranz“) als auch für DEC:OVA plus anti-CD40-Antikörper
(„Entzündung“). Jedoch wiesen transferierte ATAC KO OT-I-Zellen eine deutlich
verringerte Populationsgröße 12 und 40 Tage nach Stimulation mit DEC:OVA plus
anti-CD40-Antikörper auf. Funktionell zeigten diese ATAC KO OT-I-Zellen eine
beeinträchtige zytotoxische Aktivität an den Tagen 7 und 12. Weiterhin konnte
in vivo eine chemotaktische Wirkung von ATAC nachgewiesen werden. Nach
Injektion in den Peritonealraum wanderten bestimmte DZ-Subpopulationen
(CD8+CD205+CD11b-CD4- DZ und CD8-CD205+CD11bniedrigCD4- DZ) ein. Bei der
Untersuchung von alten ATAC KO C57BL/6 Mäusen fiel darüber hinaus auf, dass
Speicheldrüse und Milz deutlich vergrößert waren, ohne dass ein Verlust von
Körpergewicht und eine Schädigung der Organstruktur feststellbar waren. Dies
ging mit der Aktivierung und Expansion vieler Immunzellen, insbesondere von
CD4+ T-Zellen, follikulären B-Zellen und Granulozyten einher. Zusammenfassend
konnte gezeigt werden, dass ATAC die Kooperation von Antigen-erkennenden CD8+
T-Zellen mit XCR1-tragenden DZ optimiert und so die Poolgröße und zytotoxische
Kapazität dieser CD8+ T-Zellen erhöht. Zusammen mit der bereits früher
nachgewiesenen chemotaktischen Aktivität von ATAC auf XCR1+ DZ ergeben diese
funktionellen Daten zum ersten Mal ein Konzept der physiologischen Wirkung von
ATAC.ATAC, a member of the chemokine family of molecules, is secreted mostly by
activated CD8+ T cells and NK cells. Mainly due to the disputed expression
pattern of its receptor XCR1, the physiological function of ATAC remained
elusive to date. In this work, the role of ATAC and XCR1 was investigated in
the murine system. To identify the XCR1-expressing cells, the splenic cell
populations were sorted from different mouse strains and analysed in both
steady and activated states through quantitative RT-PCR. The results showed
that XCR1 is essentially expressed on CD8+ dendritic cells (DC) and to a small
extent on CD8- DC cells, but not on other cell types. Furthermore, the
influence of ATAC on the antigen-specific activation of CD8+ T cells was
examined by adoptive transfer of WT or ATAC KO OT-I cells and selective
application of antigen into the CD8+ DC of recipient mice with the help of an
DEC-205-specific antibody coupled to the antigen ovalbumin (DEC:OVA). In this
model, WT OT I T cells continued to secrete ATAC from an early time point (6
h) for up to 48 h. The absence of ATAC enhanced the expression of CD69 and
4-1BB on the transferred OT-I T cells 18-24 h after stimulation, but did not
overtly influence their proliferation or apoptosis in the 2-6 days following
stimulation with DEC:OVA alone (“tolerization”) or DEC:OVA plus anti-CD40
(“immunization”). However, the pool size of transferred ATAC KO OT-I cells was
substantially reduced on days 12 and 40 upon application of DEC:OVA in
combination with anti-CD40. Functionally, ATAC KO OT-I cells exhibited a
reduced cytotoxicity on days 7 and 12 after antigen delivery. Further, a
chemotactic activity of ATAC could be demonstrated in vivo: after i.p.
injection of ATAC, certain DC subsets (CD8+CD205+CD11b-CD4- DC and
CD8-CD205+CD11blowCD4- DC) migrated into the peritoneal cavity. The analysis
of old ATAC KO C57BL/6 mice revealed enlarged salivary glands and spleens, but
no reduction of body weight or any visible damage in organ structure. However,
immune cell populations were substantially expanded in these mice, especially
CD4+ T cells, follicular B cells, and granulocytes. In summary, we could
demonstrate that ATAC optimizes the cooperation between antigen-recognizing
CD8+ T cells and XCR1-expressing DC, which resulted in an increased pool size
and cytotoxic capacity of these CD8+ T cells. Together with the previously
observed in vitro chemotactic activity of ATAC on XCR1+ DC, this work provides
for the first time a concept for the physiological function of ATAC
Identification of transcription factor binding sites using ATAC-seq
No full textAbstract Transposase-Accessible Chromatin followed by sequencing (ATAC-seq) is a simple protocol for detection of open chromatin. Computational footprinting, the search for regions with depletion of cleavage events due to transcription factor binding, is poorly understood for ATAC-seq. We propose the first footprinting method considering ATAC-seq protocol artifacts. HINT-ATAC uses a position dependency model to learn the cleavage preferences of the transposase. We observe strand-specific cleavage patterns around transcription factor binding sites, which are determined by local nucleosome architecture. By incorporating all these biases, HINT-ATAC is able to significantly outperform competing methods in the prediction of transcription factor binding sites with footprints
CoRE-ATAC predictions overlap with experimentally detected enhancers.
No full text(A) Overlap of FANTOM enhancer annotationswith CoRE-ATAC (C) and ChromHMM (H) predictions in MCF7, A549, CD4+ T and PBMC samples. CoRE-ATAC predicted the majority of FANTOM enhancers as enhancers or promoters, recapitulating these experimentally identified enhancers. CoRE-ATAC annotations were similar toChromHMM annotations. (B) CoRE-ATAC predictions for active regulatory regions identified by STARR-seq in A549 cell line. The majority of active enhancers identified by STARR-seq were predicted as promoter or enhancer by CoRE-ATAC. (C) MIN6 MPRA log fold change values for genomic regions predicted as losing or gaining cis-RE function based on CoRE-ATAC probabilities for reference and alternative alleles. Significance for predicted loss and predicted gain categories was calculated using student’s t-test for MPRA log fold change values being less than or greater than 0 respectively. Significance comparing the predicted loss and predicted gain of MPRA fold change distributions was calculated using Mann-Whitney U test. We observed concordant direction of effect both for CoRE-ATAC predictions and MPRA activity levels when alternative and reference alleles are compared. (D) Genome browsers of 19 islet samples highlighting a loss of enhancer activity for rs11205653 (also highlighted in (C)) for the alternative allele (G). Values for enhancer and other represent the probability assigned to those classes of cis-REs by CoRE-ATAC. We observe that for 5 out of 7 individuals with the reference allele (TT) CoRE-ATAC predicted enhancer activity, reflecting ChromHMM reference annotations, while for the individuals with GT or GG alleles, we observed an enhancer activity loss for all but one individual based on CoRE-ATAC predictions.</p
Research and development of Controlled Auto-Ignition (CAI) combustion in a 4-stroke multi-cylinder gasoline engine
No full textCopyright © 2001 SAE International. This paper is posted on this site with permission from SAE International. Further use of this paper is not permitted without permission from SAEControlled Auto-Ignition (CAI) combustion has been achieved in a production type 4-stroke multi-cylinder gasoline engine. The engine was based on a Ford 1.7L Zetec-SE 16V engine with a compression ratio of 10.3, using substantially standard components modified only in design dimensions to control the gas exchange process in order to significantly increase the trapped residuals. The engine was also equipped with Variable Cam Timing (VCT) on both the intake and exhaust camshafts. It was found that the largely increased trapped residuals alone were sufficient to achieve CAI in this engine and with VCT, a range of loads between 0.5 and 4 bar BMEP and engine speeds between 1000 and 3500 rpm were mapped for CAI fuel consumption and exhaust emissions. The measured CAI results were compared with those of Spark Ignition (SI) combustion in the same engine but with standard camshafts at the same speeds and loads. The comparison showed more than 30% reduction in BSFC and up to 99% reduction in NOx at low loads
ATAC/RNA CONGAS+ analysis versus scDNA-seq.
No full textA,B. Mapping among scDNA-seq clones (ground truth) detected from a gastric cancer cell line (SNU601 [20]), and clusters inferred by CONGAS+ (λ = 0.5) from independent ATAC/RNA data, using the segmentation of the most prevalent clone from scDNA-seq. The largest cluster per mapping is highlighted to denote that there is almost a one-to-one mapping between the analyses, as is also suggested by the absolute mean absolute deviation between copy number profiles of the two analyses. C. CNA profiles for the matched analyses are in large agreement, excluding small segments on chromosomes 3, 4 and 20. D,E. A UMAP low-dimensionality representation shows good overlap between analyses. F,G. Comparison between the ATAC count distribution on the p-arm of chromosome 20, coloured by ground truth clones and CONGAS+ clusters. H. RNA distribution on the p-arm of chromosome 20 as in panels F-G shows concordance among ATAC and RNA. I,J. Differential gene expression volcano plot (Wilcoxon test) for two CONGAS+ clusters. and binding motifs associated with differently expressed ATAC peaks in both clusters.</p
Dilution effects on the controlled auto-ignition (CAI) combustion of hydrocarbon and alcohol fuels
No full textCopyright © 2001 SAE International. This paper is posted on this site with permission from SAE International. Further use of this paper is not permitted without permission from SAEThis paper presents results from an experimental programme researching the in-cylinder conditions necessary to obtain homogenous CAI (or HCCI) combustion in a 4-stroke engine. The fuels under investigation include three blends of Unleaded Gasoline, a 95 RON Primary Reference Fuel, Methanol, and Ethanol. This work concentrates on establishing the CAI operating range with regard to Air/Fuel ratio and Exhaust Gas Re-circulation and their effect on the ignition timing, combustion rate and variability, Indicated thermal efficiency, and engine-out emissions such as NOx. Detailed maps are presented, defining how each of the measured variables changes over the entire CAI region.
Results indicate that the alcohols have significantly higher tolerance to dilution than the hydrocarbon fuels tested. Also, variations in Gasoline blend have little effect on any of the combustion parameters measured
CoRE-ATAC predicted enhancers significantly overlap FANTOM and STARR-seq enhancers.
No full text(A) Distribution of CoRE-ATAC predictions for FANTOM enhancers in test chromosomes (chr3 and chr11). (B) Distribution of CoRE-ATAC predictions for test chromosomes. Pairs of bars represent comparisons between ChromHMM (H) and CoRE-ATAC (C), where each pair represents a sample/replicate for the respective cell type. (C,D) Histogram of distances to the nearest TSS for STARR-seq enhancers predicted as promoters by CoRE-ATAC for all chromosomes (C) and test chromosomes (D). Majority of predicted promoters are within 1kb of a TSS. (e,f) Histogram of distances to the nearest TSS for STARR-seq enhancers predicted as enhancers by CoRE-ATAC for all chromosomes (E) and test chromosomes (F). Majority of predicted enhancers are distal (> = 20kb) from the nearest TSS. STARR-seq enhancers annotated as promoters result from the close proximity these enhancers are to a TSS. (G,H) ChromHMM annotation (relabeled to 10 classes) distribution for STARR-seq enhancers in A549 for all chromosomes (G) and test chromosomes (H). Majority of STARR-seq enhancers are annotated as promoter or enhancer by ChromHMM. (TIF)</p
Going Beyond Counting First Authors in Author Co-citation Analysis
No full textThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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