The Influence of Temperature on Coumarin 153 Fluorescence Kinetics

The influence of temperature varied in the range 183 K–323 K on the fluorescence quantum yield, fluorescence lifetime, absorption and emission transition moments and non-radiative deactivation rate was determined for the well known and largely used dye Coumarin 153, dissolved in 1-chloropropane. The Kennard-Stepanov relation connecting the absorption and emission spectra was used to check for the presence of more than one absorbing/emitting species and to investigate whether intramolecular vibrational redistribution completes in the C153 excited S1 state before the emission takes place. The emission spectrum corresponding to S1→S0 transition, was fitted at each temperature to the model function including the information on the dye vibrational modes coupling. In this way the displacement in equilibrium distance for the most active vibrational mode was determined for C153 in S1 and in S0. Using the temperature dependence of the fluorescence decay time and quantum yield, the non-radiative deactivation rate was determined. Its temperature dependence was compared to that calculated using the theoretical model with the most active vibrational mode displacement values taken from steady-state spectra analysis. The somewhat surprising dependence of the fluorescence decay time and quantum yield on temperature was related to non-trivial coupling between low-frequency vibrational modes of C153 in the excited and ground states

The NSL Complex Regulates Housekeeping Genes in Drosophila

MOF is the major histone H4 lysine 16-specific (H4K16) acetyltransferase in mammals and Drosophila. In flies, it is involved in the regulation of X-chromosomal and autosomal genes as part of the MSL and the NSL complexes, respectively. While the function of the MSL complex as a dosage compensation regulator is fairly well understood, the role of the NSL complex in gene regulation is still poorly characterized. Here we report a comprehensive ChIP–seq analysis of four NSL complex members (NSL1, NSL3, MBD-R2, and MCRS2) throughout the Drosophila melanogaster genome. Strikingly, the majority (85.5%) of NSL-bound genes are constitutively expressed across different cell types. We find that an increased abundance of the histone modifications H4K16ac, H3K4me2, H3K4me3, and H3K9ac in gene promoter regions is characteristic of NSL-targeted genes. Furthermore, we show that these genes have a well-defined nucleosome free region and broad transcription initiation patterns. Finally, by performing ChIP–seq analyses of RNA polymerase II (Pol II) in NSL1- and NSL3-depleted cells, we demonstrate that both NSL proteins are required for efficient recruitment of Pol II to NSL target gene promoters. The observed Pol II reduction coincides with compromised binding of TBP and TFIIB to target promoters, indicating that the NSL complex is required for optimal recruitment of the pre-initiation complex on target genes. Moreover, genes that undergo the most dramatic loss of Pol II upon NSL knockdowns tend to be enriched in DNA Replication–related Element (DRE). Taken together, our findings show that the MOF-containing NSL complex acts as a major regulator of housekeeping genes in flies by modulating initiation of Pol II transcription

Explicit referencing in chat supports collaborative learning

In Pfister & Mühlpfordt (2002) a study was presented showing that chat discussions with a strict turn order combined with the requirement to assign a type and an explicit reference to each message lead to a higher learning score than discussions in a normal chat or in a chat with strict turn order only. Due to the experimental design it was not possible to judge the role of explicit referencing. Now we present the "missing" data: The higher learning score can be explained just by the explicit referencing. We argue that this is an important design issue for chat applications, because it seems that explicit referencing leads to a more homogeneous discourse behavior (more homogeneous participation, more participation in parallel discussion threads) and a better grounding. A case study explored the use of the referencing function in a less restricted everyday collaborative situation

Lernprotokollunterstütztes Lernen - ein Vergleich zwischen unstrukturiertem und systemkontrolliertem diskursivem Lernen im Netz

Cooperative learning in net-based scenarios is often conducted using synchronous communication tools such as text-based chat. Knowledge acquisition by exchanging textual contributions frequently leads to suboptimal results, since the discourse suffers from a lack of coordination and from incoherent contributions. What we call learning protocols is a method to improve coordination and coherence in net-based discourses, and thus to increase learning gains. Learning protocols are implemented as system-controlled cooperation scripts. Participants are required to reference their contributions, to indicate the type of each contribution, and to follow a predefined sequence when submitting a contribution. An experimental study is reported presenting evidence which confirms the positive effect of a learning protocol for mutual explanations, especially with respect to the referencing function. However, it turns out that the usefulness of the learning protocol is highly domain specific

Photophysics of 3-hydroxyflavone in supercritical CO2: a probe to study the microenvironment of SCF

The excitation of 3-hydroxyflavone (3HF) to its second excited singlet state (S2) gives rise to dual fluorescence in supercritical carbon dioxide. The ultraviolet fluorescence originated from the S2 state of 3HF is well separated from the green emission emanating from the tautomeric form, produced via the excited state intramolecular proton transfer. The relative intensity of the S2 to the tautomer fluorescence (S2/T) has been studied as a function of pressure and temperature. It is shown that this ratio reflects the microheterogeneity of the supercritical CO2, and confirms the value of fluorometric probes in disclosing the microscopic properties of supercritical fluids.http://www.sciencedirect.com/science/article/B6TFN-4BVP7G9-3/1/02dd61c567fe3e9c8d6ac86a01f79ce

Summary model: NSL-dependent Pol II recruitment to promoters of housekeeping genes.

<p>The majority of the NSL-bound targets are constitutively expressed or “housekeeping” genes. These genes are characterized by prominent enrichment of particular histone modifications (H4K16ac, H3K9ac, H3K4me2, H3K4me3) as well as specific core promoter elements (such as DRE, E-box and motif 1; indicated by colored squares). In contrast, tissue-specific or developmentally regulated genes (small inlay) usually contain the TATA-box as the most prominent core promoter element. We propose that the NSL complex acts as a regulator of constitutively expressed genes by facilitating stable recruitment of the pre-initiation complex (PIC) members such as Pol II, TBP and TFIIB on target genes. NSL complex may therefore serve as an important link between specific promoter architecture and PIC assembly.</p

NSL proteins preferably associate with the promoters of constitutively active genes.

<p>(A) Metagene profiles of histone modifications reveal higher ratios of active chromatin marks H3K4me2/3, H4K16ac and H3K9ac for active genes bound by the NSL complex compared to active NSL-non-bound and inactive genes. On the contrary, the repressive mark H3K27me2 is not enriched on gene promoters bound by the NSL complex. Active genes were defined according to the expression data from <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Cherbas1" target="_blank">[28]</a> (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#s3" target="_blank">Materials and Methods</a>). The expression levels of NSL-bound and NSL-non-bound active genes are similar (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736.s002" target="_blank">Figure S2A</a>). The log₂ ratios ( = log₂FC (ChIP/input)) of the histone modifications were obtained from modENCODE, extracted for 200 bp bins, and normalized to H4 Chip-chip signals. (B) The chromatin color model contains <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Filion1" target="_blank">[29]</a> two states of euchromatin: “yellow” and “red”. NSL-bound TSSs are predominantly associated with “yellow”, but not “red” chromatin. NSL-non-bound genes display chromatin color ratios that resemble the pattern seen by Filion et al. for tissue-specific genes. (“Green” and “blue” correspond to classical and PcG heterochromatin, respectively, while “black” denotes regions of repressive chromatin). (C) For a different model of chromatin states devised by Kharchenko et al., similar results as in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen-1002736-g002" target="_blank">Figure 2B</a> were obtained: The pie chart depicts that 93% of all peaks of NSL1, MCRS2, NSL3 and MBD-R2 that localize within +/−200 bp of the nearest TSS associate with regions of chromatin state 1. This is defined as the state of actively transcribed TSSs <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Kharchenko1" target="_blank">[30]</a>. Complementary, as shown in the bar chart, NSL-bound TSSs of expressed genes are significantly enriched in chromatin state 1 and depleted of chromatin state 9 (p-values<2.2e-16; binomial test) while NSL-non-bound genes are more equally distributed between states of active TSSs (1) and elongation marks (states 2, 3, 4). (D) Heatmaps of ChIP-seq signals (log₂FCs) demonstrate the strong enrichment of NSL binding around the TSSs of constitutively transcribed genes. In contrast to the Pol II signal that is present in both constitutive and regulatory (not constitutive) active genes, the NSL proteins are predominantly found around the TSSs of constitutively transcribed genes. As indicated on the left-hand side, genes were sorted according to their genomic location. The proteins' binding intensities can be directly compared between the different panels. The inlay (right) illustrates the findings of the heatmap with a focus on genes that are bound by all NSLs concomitantly: 85.5% of NSL-bound promoters are constitutively expressed (light blue area). Active (not constitutive) and inactive genes are represented by dark blue and white areas, respectively.</p