Oct4 differentially regulates chromatin opening and enhancer transcription in pluripotent stem cells

The transcription factor Oct4 is essential for the maintenance and induction of stem cell pluripotency, but its functional roles are not fully understood. Here, we investigate the functions of Oct4 by depleting and subsequently recovering it in mouse embryonic stem cells (ESCs) and conducting a time-resolved multiomics analysis. Oct4 depletion leads to an immediate loss of its binding to enhancers, accompanied by a decrease in mRNA synthesis from its target genes that are part of the transcriptional network that maintains pluripotency. Gradual decrease of Oct4 binding to enhancers does not immediately change the chromatin accessibility but reduces transcription of enhancers. Conversely, partial recovery of Oct4 expression results in a rapid increase in chromatin accessibility, whereas enhancer transcription does not fully recover. These results indicate different concentration-dependent activities of Oct4. Whereas normal ESC levels of Oct4 are required for transcription of pluripotency enhancers, low levels of Oct4 are sufficient to retain chromatin accessibility, likely together with other factors such as Sox2

Genome-wide activity of unliganded estrogen receptor-\u3b1\ua0 in breast cancer cells

Estrogen receptor-\u3b1 (ER\u3b1) has central role in hormone-dependent breast cancer and its ligand-induced functions have been extensively characterized. However, evidence exists that ER\u3b1 has functions that are independent of ligands. In the present work, we investigated the binding of ER\u3b1 to chromatin in the absence of ligands and its functions on gene regulation. We demonstrated that in MCF7 breast cancer cells unliganded ER\u3b1 binds to more than 4,000 chromatin sites. Unexpectedly, although almost entirely comprised in the larger group of estrogen-induced binding sites, we found that unliganded-ER\u3b1 binding is specifically linked to genes with developmental functions, compared with estrogen-induced binding. Moreover, we found that siRNA-mediated down-regulation of ER\u3b1 in absence of estrogen is accompanied by changes in the expression levels of hundreds of coding and noncoding RNAs. Down-regulatedmRNAs showed enrichment in genes related to epithelial cell growth and development. Stable ER\u3b1 down-regulation using shRNA, which caused cell growth arrest, was accompanied by increased H3K27me3 at ER\u3b1 binding sites. Finally, we found that FOXA1 and AP2\u3b3 binding to several sites is decreased upon ER\u3b1 silencing, suggesting that unliganded ER\u3b1 participates, together with other factors, in the maintenance of the luminal-specific cistrome in breast cancer cell

CDK12 globally stimulates RNA polymerase II transcription elongation and carboxyl-terminal domain phosphorylation

Cyclin-dependent kinase 12 (CDK12) phosphorylates the carboxyl-terminal domain (CTD) of RNA polymerase II (pol II) but its roles in transcription beyond the expression of DNA damage response genes remain unclear. Here, we have used TT-seq and mNET-seq to monitor the direct effects of rapid CDK12 inhibition on transcription activity and CTD phosphorylation in human cells. CDK12 inhibition causes a genome-wide defect in transcription elongation and a global reduction of CTD Ser2 and Ser5 phosphorylation. The elongation defect is explained by the loss of the elongation factors LEO1 and CDC73, part of PAF1 complex, and SPT6 from the newly-elongating pol II. Our results indicate that CDK12 is a general activator of pol II transcription elongation and indicate that it targets both Ser2 and Ser5 residues of the pol II CTD

CPX-351 treatment in secondary acute myeloblastic leukemia is effective and improves the feasibility of allogeneic stem cell transplantation: results of the Italian compassionate use program

Secondary acute myeloid leukemia (sAML) poorly responds to conventional treatments and allogeneic stem cell transplantation (HSCT). We evaluated toxicity and efficacy of CPX-351 in 71 elderly patients (median age 66 years) with sAML enrolled in the Italian Named (Compassionate) Use Program. Sixty days treatment-related mortality was 7% (5/71). The response rate at the end of treatment was: CR/CRi in 50/71 patients (70.4%), PR in 6/71 (8.5%), and NR in 10/71 (19.7%). After a median follow-up of 11 months relapse was observed in 10/50 patients (20%) and 12 months cumulative incidence of relapse (CIR) was 23.6%. Median duration of response was not reached. In competing risk analysis, CIR was reduced when HSCT was performed in first CR (12 months CIR of 5% and 37.4%, respectively, for patients receiving (=20) or not (=30) HSCT, p = 0.012). Twelve-months OS was 68.6% (median not reached). In landmark analysis, HSCT in CR1 was the only significant predictor of longer survival (12 months OS of 100 and 70.5%, for patients undergoing or not HSCT in CR1, respectively, p = 0.011). In conclusion, we extend to a real-life setting, the notion that CPX is an effective regimen for high risk AML patients and may improve the results of HSCT

Critical literacy as a pedagogical goal in English language teaching

In this chapter, the authors provide an overview of the area of critical literacy as it pertains to second language pedagogy (curriculum and instruction). After considering the historical origins of critical literacy (from antiquity, and including in first language education), they consider how it began to penetrate the field of applied linguistics. They note the geographical and institutional spread of critical literacy practice as documented by published accounts. They then sketch the main features of L2 critical literacy practice. To do this, they acknowledge how practitioners have reported on their practices regarding classroom content and process. The authors also draw attention to the outcomes of these practices as well as challenges that practitioners have encountered in incorporating critical literacy into their second language classrooms

Internal combustion engine analysis using EngineEnvelope (EE)

Many common industrial activities, like energy production, people and goods transportation, heavily rely on internal combustion engines\u2019 operations. The capabilities of reducing the total cost of operation and maximizing the uptime of these machines are a very important topic for the their owner and much energy has to be put on the purpose. In many cases, scheduled maintenance programs are adopted. After a defined amount of time, decided by the engine producer, parts are overhauled and updated regardless of the real usage level and damage. Instead, it is clear that being able to substitute only the damaged parts, at due time, will reduce the engine ownership and maintenance costs. While on the market several maintenance tools dedicated to the on-line monitoring of the thermodynamic aspects (mixture characteristics, exhausts analysis, miss-firing) are present, none are available for the evaluation of the kinematics links problems. The paper propose a novel technique, named EngineEnvelope (EE), focused on the online monitoring of the mechanical potential issues like bolt looseness, pistons clearance and bearings damages. The paper describe the monitoring function, its capabilities of detecting damages on real data acquired on a two strokes, single cylinder engine

Global donor and acceptor splicing site kinetics in human cells.

RNA splicing is an essential part of eukaryotic gene expression. Although the mechanism of splicing has been extensively studied in vitro, in vivo kinetics for the two-step splicing reaction remain poorly understood. Here we combine transient transcriptome sequencing (TT-seq) and mathematical modeling to quantify RNA metabolic rates at donor and acceptor splice sites across the human genome. Splicing occurs in the range of minutes and is limited by the speed of RNA polymerase elongation. Splicing kinetics strongly depends on the position and nature of nucleotides flanking splice sites, and on structural interactions between unspliced RNA and small nuclear RNAs in spliceosomal intermediates. Finally, we introduce the 'yield' of splicing as the efficiency of converting unspliced to spliced RNA and show that it is highest for mRNAs and independent of splicing kinetics. These results lead to quantitative models describing how splicing rates are encoded in the human genome

Identification of novel LTR retrotransposons in the genome of Aedes aegypti

We have detected seventy-six novel LTR retrotransposons in the genome of the mosquito Aedes aegypti by a genome wide analysis using the LTR_STRUC program. We have performed a phylogenetic classi!cation of these novel elements and a distribution analysis in the genome of A. aegypti. These mobile elements belong either to the Ty3/gypsy or to the Bel family of retrotransposons and were not annotated in the mosquito LTR retrotransposon database (TEfam). We have found that !1.8% of the genome is occupied by these newly detected retrotransposons that are distributed predominantly in intergenic genomic sequences and introns. The potential role of retrotransposon insertions linked to host genes is described and discussed. We show that a retrotransposon family belonging to the Osvaldo lineage has peculiar structural features, and its presence is likely to be restricted to the A. aegypti and to the Culex pipiens quinquefasciatus genomes. Furthermore we show that the ninja-like group of elements lacks the Primer Binding Site (PBS) sequence necessary for the replication of retrotransposons. These results integrate the knowledge on the complicate genomic structure of an important disease vector