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Growing population and water demand have increased pressure on water resources in various parts of the globe, including many transboundary river basins. While the impacts of upstream water use on downstream water availability have been analysed in many of these international river basins, this has not been systematically done at the global scale using coherent and comparable datasets. In this study, we aim to assess the change in downstream water stress due to upstream water use in the world's transboundary river basins. Water stress was first calculated considering only local water use of each sub-basin based on country-basin mesh, then compared with the situation when upstream water use was subtracted from downstream water availability. We found that water stress was generally already high when considering only local water use, affecting 0.95-1.44 billion people or 33%-51% of the population in transboundary river basins. After accounting for upstream water use, stress level increased by at least 1 percentage-point for 30-65 sub-basins, affecting 0.29-1.13 billion people. Altogether 288 out of 298 middle-stream and downstream sub-basin areas experienced some change in stress level. Further, we assessed whether there is a link between increased water stress due to upstream water use and the number of conflictive and cooperative events in the transboundary river basins, as captured by two prominent databases. No direct relationship was found. This supports the argument that conflicts and cooperation events originate from a combination of different drivers, among which upstream-induced water stress may play a role. Our findings contribute to better understanding of upstream-downstream dynamics in water stress to help address water allocation problems.

The differentiation of activated CD4(+) T cells into the T helper type 1 (T(H)1) or T(H)2 fate is regulated by cytokines and the transcription factors T-bet and GATA-3. Whereas interleukin 12 (IL-12) produced by antigen-presenting cells initiates the T(H)1 fate, signals that initiate the T(H)2 fate are not completely characterized. Here we show that early GATA-3 expression, required for T(H)2 differentiation, was induced by T cell factor 1 (TCF-1) and its cofactor beta-catenin, mainly from the proximal Gata3 promoter upstream of exon 1b. This activity was induced after T cell antigen receptor (TCR) stimulation and was independent of IL-4 receptor signaling through the transcription factor STAT6. Furthermore, TCF-1 blocked T(H)1 fate by negatively regulating interferon-gamma (IFN-gamma) expression independently of beta-catenin. Thus, TCF-1 initiates T(H)2 differentiation of activated CD4(+) T cells by promoting GATA-3 expression and suppressing IFN-gamma expression.

Therapeutic resistance remains a primary obstacle to curing cancer. Healthy cells exposed to genotoxic insult rapidly activate both p53-dependent and -independent non-genetic programs that pause the cell cycle and direct either DNA repair or apoptosis. Cancer cells exploit these same pathways as they respond to stresses induced by cancer therapies. In this study, we investigated a potential role for upstream stimulatory factor 1 (USF1) and USF2 in the p53-independent response of lymphoma cells to genotoxic therapy. We previously found that lymphocytes utilize the responsiveness of USF1 to double-stranded DNA breaks to coordinate T cell receptor beta ( Tcrb ) gene expression during V(D)J recombination. Here, microarray gene expression analysis of derivatives of the p53-deficient mouse B lymphoma cell line, M12, revealed that simultaneously depleting cells of both USF1 and USF2 altered the expression of 940 gene transcripts (>1.50-fold change, < 0.05 FDR), relative to cells expressing a scrambled control shRNA. Seven days after exposure to a single sublethal (5 Gy) dose of ionizing radiation, USF-depleted (USFKD) cells exhibited widespread and distinct transcriptional responses from those of irradiated controls (5035 and 5054 differentially expressed gene transcripts, respectively, with roughly half shared between both cell types). Gene ontology analyses revealed that USF knockdown induced numerous changes in the expression of genes critical for immune development and function while diminishing the responsiveness of genes linked to DNA damage pathways. Microarray findings were confirmed by RT-qPCR for a panel of genes responsive to USF knockdown and/or irradiation. These findings shed further light on transcriptional responses to ionizing radiation that manifest over time in transformed cells, identifying a novel p53-independent role in lymphocytic DNA damage stress responses for USF.

Purpose Gastric cancer is an inflammation-driven disease often associated with a bad prognosis. Upstream stimulatory factors USF1 and USF2 are pleiotropic transcription factors, with tumor suppressor function. Low expression of USF1 is associated with low survival in gastric cancer patients. USF1 genetic polymorphism -202G > A has been associated with cancer susceptibility. Our aim was to investigate USF1 gene polymorphism and serum level with the risk of gastric cancer. Methods USF1-202 G/A polymorphism was analyzed by sanger sequencing, with the measure of USF1/USF2 serum levels by ELISA in H. pylori -positive patients with chronic gastritis, gastric precancerous lesions, gastric cancer and in healthy controls. Results Our results show that the presence of the USF1-202 A allele increased the risk of gastric cancer compared to G (OR = 2; 95% CI 1.07–3.9; P = 0.02). Genotypically and under the dominant mutation model, the combined USF1- GA/AA -202 genotypes corresponded to higher risk of gastric cancer (OR = 3.5; 95% CI 1.4–8.2; p-value = 0.005) than the GG genotype. Moreover, the G/A transition at USF1-202 was associated with lower USF1 serum level, and mostly observed in gastric cancer patients where the average serological level of USF1 were 2.3 and twofold lower for the AA and GA genotypes, respectively, compared to GG. Conclusion USF1-202 G/A polymorphism constitutes a gastric cancer genetic risk factor. Together with USF1/USF2 serum level, they can be proposed as promising biomarkers for gastric cancer detection/prevention.

A Linux distribution consists of thousands of packages that are either developed by in-house developers (in-house packages) or by external projects (upstream packages). Leveraging upstream packages speeds up development and improves productivity, yet bugs might slip through into the packaged code and end up propagating into downstream Linux distributions. Maintainers, who integrate upstream projects into their distribution, typically lack the expertise of the upstream projects. Hence, they could try either to propagate the bug report upstream and wait for a fix, or fix the bug locally and maintain the fix until it is incorporated upstream. Both of these outcomes come at a cost, yet, to the best of our knowledge, no prior work has conducted an in-depth analysis of upstream bug management in the Linux ecosystem. Hence, this paper empirically studies how high-severity bugs are fixed in upstream packages for two Linux distributions, i.e., Debian and Fedora. Our results show that 13.9% of the upstream package bugs are explicitly reported being fixed by upstream, and 13.3% being fixed by the distribution, while the vast majority of bugs do not have explicit information about this in Debian. When focusing on the 27.2% with explicit information, our results also indicate that upstream fixed bugs make users wait for a longer time to get fixes and require more additional information compared to fixing upstream bugs locally by the distribution. Finally, we observe that the number of bug comment links to reference information (e.g., design docs, bug reports) of the distribution itself and the similarity score between upstream and distribution bug reports are important factors for the likelihood of a bug being fixed upstream. Our findings strengthen the need for traceability tools on bug fixes of upstream packages between upstream and distributions in order to find upstream fixes easier and lower the cost of upstream bug management locally.

Autophagy, a highly conserved self-digestion process crucial for cellular homeostasis, is triggered by various environmental signals, including nutrient scarcity. The regulation of lysosomal and autophagy-related processes is pivotal to maintaining cellular homeostasis and basal metabolism. The consequences of disrupting or diminishing lysosomal and autophagy systems have been investigated; however, information on the implications of hyperactivating lysosomal and autophagy genes on homeostasis is limited. Here, we present a mechanism of transcriptional repression involving upstream stimulatory factor 2 (USF2), which inhibits lysosomal and autophagy genes under nutrient-rich conditions. We find that USF2, together with HDAC1, binds to the CLEAR motif within lysosomal genes, thereby diminishing histone H3K27 acetylation, restricting chromatin accessibility, and downregulating lysosomal gene expression. Under starvation, USF2 competes with transcription factor EB (TFEB), a master transcriptional activator of lysosomal and autophagy genes, to bind to target gene promoters in a phosphorylation-dependent manner. The GSK3β-mediated phosphorylation of the USF2 S155 site governs USF2 DNA-binding activity, which is involved in lysosomal gene repression. These findings have potential applications in the treatment of protein aggregation-associated diseases, including α1-antitrypsin deficiency. Notably, USF2 repression is a promising therapeutic strategy for lysosomal and autophagy-related diseases. Here, the authors show that USF2 epigenetically represses autophagy and lysosomal genes by recruiting NuRD complex and competing with TFEB. USF2 depletion enhances these processes, offering a potential therapeutic strategy for aggregate diseases.

Angiotensin II type 1 receptor-associated protein (ATRAP) is widely expressed in different tissues and organs, although its mechanistic role in breast cancer remains unclear. Here, we show that ATRAP is highly expressed in breast cancer tissues. Its aberrant upregulation promotes breast cancer aggressiveness and is positively correlated with poor prognosis. Functional assays revealed that ATRAP participates in promoting cell growth, metastasis, and aerobic glycolysis, while microarray analysis showed that ATRAP can activate the AKT/mTOR signaling pathway in cancer progression. In addition, ATRAP was revealed to direct Ubiquitin-specific protease 14 (USP14)-mediated deubiquitination and stabilization of Pre-B cell leukemia homeobox 3 (PBX3). Importantly, ATRAP is a direct target of Upstream stimulatory factor 1 (USF1), and that ATRAP overexpression reverses the inhibitory effects of USF1 knockdown. Our study demonstrates the broad contribution of the USF1/ATRAP/PBX3 axis to breast cancer progression and provides a strong potential therapeutic target.

Biopharmaceuticals have revolutionized the field of medicine in the types of active ingredient molecules and treatable indications. Adoption of Quality by Design and Process Analytical Technology (PAT) frameworks has helped the biopharmaceutical field to realize consistent product quality, process intensification, and real-time control. As part of the PAT strategy, Raman spectroscopy offers many benefits and is used successfully in bioprocessing from single-cell analysis to cGMP process control. Since first introduced in 2011 for industrial bioprocessing applications, Raman has become a first-choice PAT for monitoring and controlling upstream bioprocesses because it facilitates advanced process control and enables consistent process quality. This paper will discuss new frontiers in extending these successes in upstream from scale-down to commercial manufacturing. New reports concerning the use of Raman spectroscopy in the basic science of single cells and downstream process monitoring illustrate industrial recognition of Raman’s value throughout a biopharmaceutical product’s lifecycle. Finally, we draw upon a nearly 90-year history in biological Raman spectroscopy to provide the basis for laboratory and in-line measurements of protein quality, including higher-order structure and composition modifications, to support formulation development.

Epigenetic modifications of DNA play important roles in many biological processes. Identifying readers of these epigenetic marks is a critical step towards understanding the underlying mechanisms. Here, we present an all-to-all approach, dubbed digital affinity profiling via proximity ligation (DAPPL), to simultaneously profile human TF-DNA interactions using mixtures of random DNA libraries carrying different epigenetic modifications (i.e., 5-methylcytosine, 5-hydroxymethylcytosine, 5-formylcytosine, and 5-carboxylcytosine) on CpG dinucleotides. Many proteins that recognize consensus sequences carrying these modifications in symmetric and/or hemi-modified forms are identified. We further demonstrate that the modifications in different sequence contexts could either enhance or suppress TF binding activity. Moreover, many modifications can affect TF binding specificity. Furthermore, symmetric modifications show a stronger effect in either enhancing or suppressing TF-DNA interactions than hemi-modifications. Finally, in vivo evidence suggests that USF1 and USF2 might regulate transcription via hydroxymethylcytosine-binding activity in weak enhancers in human embryonic stem cells. Identifying readers of epigenetic marks is a critical step for understanding the role of epigenetic marks in biology. Here, the authors applied DAPPL, an all-to-all approach to profile the interactions between TFs and epigenetic modified DNA libraries.

Autophagy has been involved in protection of ischemia/reperfusion (I/R)-induced injury in many tissues including the brain. The upstream stimulatory factor 2 (Usf2) was proposed as a regulator in aging and degenerative brain diseases; however, the its role in autophagy during cerebral I/R injury remains unclear. Here, the middle cerebral artery occlusion (MCAO) operation was applied to establish an I/R mouse model. We showed that Usf2 was significantly upregulated in I/R-injured brain, accompanied by decreased levels of autophagy. Then, oxygen-glucose deprivation/recovery (OGD/R) treatment was used to establish a cellular I/R model in HT22 neurons, and lentiviral interference vector against Usf2 (LV-sh-Usf2) was used to infect the neurons. Our results showed that Usf2 was significantly upregulated in OGD/R-treated HT22 neurons that displayed an increased level in cell apoptosis and decreased levels in cell viability and autophagy, and interference of Usf2 largely rescued the effects of OGD/R on cell viability, apoptosis, and autophagy, suggesting an important role of Usf2 in neuron autophagy. In the mechanism exploration, we found that, as a transcription factor, Usf2 bound to the promoter of YTHDF1, a famous reader of N6-Methyladenosine (m6A), also induced by OGD/R, and promoted its transcription. Overexpression of YTHDF1 was able to reverse the improvement of Usf2 interference on viability and autophagy of HT22 neurons. Moreover, YTHDF1 suppressed autophagy to induce HT22 cell apoptosis through increasing m6A-mediated stability of Cdc25A, a newly identified autophagy inhibitor. Finally, we demonstrated that interference of Usf2 markedly improved autophagy and alleviated I/R-induced injury in MCAO mice.