bluuebunny/crossref_metadata_2025_split huggingface dataset:
The WalRK two-component system is essential for proper cell envelope biogenesis in Clostridioides difficile
Authors: Müh Ute , Ellermeier Craig D. , Weiss David S. 
Published: April 2022

AbstractThe WalR-WalK two-component regulatory system (TCS) is found in all Firmicutes, where it regulates the expression of multiple genes required for remodeling the cell envelope during growth and division. Unlike most TCSs, WalRK is essential for viability, so it has attracted interest as a potential antibiotic target. Here we used overexpression of WalR and CRISPR interference to investigate the Wal system of Clostridioides difficile, a major cause of hospital-associated diarrhea in high-income countries. We confirmed the wal operon is essential and identified morphological defects and cell lysis as the major terminal phenotypes of altered wal expression. We also used RNA-seq to identify over 150 genes whose expression changes in response to WalR levels. This gene set is enriched in cell envelope genes and includes several predicted PG hydrolases and proteins that could regulate PG hydrolase activity. A distinct feature of the C. difficile cell envelope is the presence of an S-layer and we found WalR affects expression of several genes which encode S-layer proteins. An unexpected finding was that some Wal-associated phenotypic defects were inverted in comparison to what has been reported in other Firmicutes. For example, down-regulation of Wal signaling caused C. difficile cells to become longer rather than shorter, as in Bacillus subtilis. Likewise, down-regulation of Wal rendered C. difficile more sensitive to vancomycin, whereas reduced Wal activity is linked to increased vancomycin resistance in Staphylococcus aureus.ImportanceThe WalRK two-component system (TCS) is essential for coordinating synthesis and turnover of peptidoglycan in Firmicutes. Here we investigate the WalRK TCS in Clostridioides difficile, an important bacterial pathogen with an atypical cell envelope. We confirmed that WalRK is essential and regulates cell envelope biogenesis, although several of the phenotypic changes we observed were opposite to what has been reported in other Firmicutes. We also identified over 150 genes whose expression is controlled either directly or indirectly by WalR. Overall, our findings provide a foundation for future investigations of an important regulatory system and potential antibiotic target in C. difficile.
Building molecular band-pass filters via molecular sequestration
Authors: Zhang Yichi , Cuba Samaniego Christian , Carleton Katelyn , Qian Yili , Giordano Giulia , Franco Elisa 
Published: April 2022

AbstractEngineered genetic circuits with tailored functions that mimic how cells process information in changing environments (e.g. cell fate decision, chemotaxis, immune response) have great applications in biomedicine and synthetic biology. Although there is a lot of progress toward the design of gene circuits yielding desired steady states (e.g. logic-based networks), building synthetic circuits for dynamic signal processing (e.g. filters, frequency modulation, and controllers) is still challenging. Here, we provide a model-based approach to build gene networks that can operate as band-pass filters by taking advantage of molecular sequestration. By suitably approximating the dynamics of molecular sequestration, we analyze an Incoherent Feed-Forward Loop (IFFL) and a Negative Feedback (NF) circuit and illustrate how they can achieve band-pass filter behavior. Computational analysis shows that a circuit that incorporates both IFFL and NF motifs improves the filter performance. Our approach facilitates the design of sequestration-based filters, and may support the synthesis of molecular controllers with desired specifications.
Profound phenotypic and epigenetic heterogeneity of the HIV-1 infected CD4+ T cell reservoir
Authors: Wu Vincent H. , Nguyen Son , Nordin Jayme M.L. , Joy Jaimy , Mampe Felicity , Tebas Pablo , Montaner Luis J. , Bar Katharine J. , Vella Laura A. , Betts Michael R. 
Published: April 2022

AbstractUnderstanding the complexity of the long-lived HIV reservoir during antiretroviral therapy (ART) remains a major impediment for HIV cure research. To address this, we developed single-cell viral ASAPseq to precisely define the unperturbed peripheral blood HIV-infected memory CD4+ T cell reservoir from antiretroviral treated people living with HIV (ART-PLWH) via the presence of integrated accessible proviral DNA in concert with epigenetic and cell surface protein profiling. We identified profound reservoir heterogeneity within and between ART-PLWH, characterized by novel and known surface markers within total and individual memory CD4+ T cell subsets. We further uncovered novel epigenetic profiles and transcription factor motifs enriched in HIV-infected cells that suggest infected cells with accessible provirus, irrespective of reservoir distribution, are poised for reactivation during ART treatment. Together, our findings reveal the extensive inter- and intrapersonal cellular heterogeneity of the HIV reservoir, and establish an initial multiomic atlas to develop targeted reservoir elimination strategies.
Cotranslational folding and assembly of the dimeric E. coli inner membrane protein EmrE
Authors: Mermans Daphne , Nicolaus Felix , Fleisch Klara , von Heijne Gunnar 
Published: April 2022

AbstractIn recent years, it has become clear that many homo- and heterodimeric cytoplasmic proteins in both prokaryotic and eukaryotic cells start to dimerize cotranslationally, i.e., while at least one of the two chains is still attached to the ribosome. Whether this is possible also for integral membrane proteins is unknown, however. Here, we apply Force Profile Analysis (FPA) – a method where a translational arrest peptide (AP) engineered into the polypeptide chain is used to detect force generated on the nascent chain during membrane insertion – to demonstrate cotranslational interactions between a fully membrane-inserted monomer and a nascent, ribosome-tethered monomer of the E. coli inner membrane protein EmrE. Similar cotranslational interactions are also seen when the two monomers are fused into a single polypeptide. Further, we uncover an apparent intrachain interaction between E14 in TMH1 and S64 in TMH3 that forms at a precise nascent chain length during cotranslational membrane insertion of an EmrE monomer. Like soluble proteins, inner membrane proteins can thus both start to fold and start to dimerize during the cotranslational membrane-insertion process.Significance statementMany water-soluble proteins are known to fold and even dimerize cotranslationally, i.e., when still attached to the ribosome. Here, we show that an E. coli inner membrane protein can also start to fold and dimerize cotranslationally, establishing the generality of these cotranslational maturation processes.
Control of pancreatic islet function and glucose homeostasis by a novel microexon program misregulated in type 2 diabetes
Authors: Juan-Mateu Jonàs , Bajew Simon , Miret-Cuesta Marta , Íñiguez Luis P. , López-Pascual Amaya , Bonnal Sophie , Atla Goutham , Bonàs-Guarch Sílvia , Ferrer Jorge , Valcárcel Juan , Irimia Manuel 
Published: April 2022

AbstractPancreatic islets control glucose homeostasis by the balanced secretion of insulin and other hormones, and their abnormal function causes diabetes or hypoglycemia. Here, we uncover a conserved program of alternative microexons included in mRNAs of islet cells, particularly in genes involved in vesicle transport and exocytosis. Islet microexons (IsletMICs) are regulated by the RNA binding proteinSRRM3and represent a subset of the larger neural program that are particularly sensitive to the levels of this regulator. BothSRRM3and IsletMICs are induced by elevated glucose levels, and depletion ofSRRM3in beta cell lines and mouse islets, or repression of particular IsletMICs using antisense oligonucleotides, leads to inappropriate insulin secretion. Consistently,SRRM3mutant mice display defects in islet cell identity and function, leading to hyperinsulinemic hypoglycemia. Importantly, human genetic variants that influenceSRRM3expression and IsletMIC inclusion in islets are associated with fasting glucose variation and type 2 diabetes risk.
The optimal strategy of incompatible insect technique (IIT) using Wolbachia to control Malaria
Authors: Matsufuji Taiga , Seirin-Lee Sungrim 
Published: April 2022

AbstractFor decades, techniques to control vector population with low environmental impact have been widely explored in both field and theoretical studies. The incompatible insect method (IIT) using Wolbachia, based on cytoplasmic incompatibility, is a technique that Wolbachia-infected male mosquitoes are incapable of producing viable offspring after mating with wildtype female mosquitoes. While the IIT method experimentally ensured its effectiveness in several field works, the failure of female mosquito population control owing to the accidental contamination of Wolbachia-infected female mosquitoes has been a concern and an obstacle in implementing the IIT method in nature. In this study, we developed a population-based IIT mathematical model using cytoplasmic incompatibility and evaluated the effectiveness of the IIT method in scenarios where contamination was present or absent. In addition, by extending the model to assess the disease infection status of the human population with malaria, we evaluated the optimal release strategy and cost for successful disease control. Our study proves that IIT could be a promising method to control mosquito-borne diseases without perfect eradication of vector mosquito population regardless of contamination.
FCHO controls AP2’s critical endocytic roles through a PtdIns4,5P2 membrane-dependent switch
Authors: Zaccai Nathan R. , Kadlecova Zuzana , Dickson Veronica Kane , Korobchevskaya Kseniya , Kamenicky Jan , Kovtun Oleksiy , Umasankar Perunthottathu K. , Wrobel Antoni G. , Kaufman Jonathan G.G. , Gray Sally , Qu Kun , Evans Philip R. , Fritzsche Marco , Sroubek Filip , Höning Stefan , Briggs John A.G. , Kelly Bernard T. , Owen David J. , Traub Linton M. 
Published: April 2022

AbstractClathrin-mediated endocytosis (CME) is the main mechanism by which mammalian cells control their cell surface proteome. Proper operation of the pivotal CME cargo-adaptor AP2 requires membrane-localised FCHO. Here, live-cell eTIRF-SIM shows that FCHO marks sites of clathrin- coated pit (CCP) initiation, which mature into uniform sized CCPs comprising a central patch of AP2 and clathrin corralled by an FCHO/Eps15 ring. We dissect the network of interactions between the FCHO interdomain-linker and AP2, which concentrates, orients, tethers and partially destabilizes closed AP2 at the plasma membrane. AP2’s subsequent membrane deposition drives its opening, which triggers FCHO displacement through steric competition with PtdIns4,5P2, clathrin, cargo and CME accessory factors. FCHO can now relocate toward a CCP’s outer edge to engage and activate further AP2s to drive CCP growth/maturation.125 character summaryFCHO primes AP2 for CCV incorporation, a process that triggers FCHO release to enable activation/recruitment of further AP2s
AcrNET: Predicting Anti-CRISPR with Deep Learning
Authors: Li Yunxiang , Wei Yumeng , Xu Sheng , Tan Qingxiong , Zong Licheng , Wang Jiuming , Wang Yixuan , Chen Jiayang , Hong Liang , Li Yu 
Published: April 2022

ABSTRACTAs an important group of proteins discovered in phages, anti-CRISPR inhibits the activity of the immune system of bacteria (i.e., CRISPR-Cas), showing great potential for gene editing and phage therapy. However, the prediction and discovery of anti-CRISPR are challenging for its high variability and fast evolution. Existing biological studies often depend on known CRISPR and anti-CRISPR pairs, which may not be practical considering the huge number of pairs in reality. Computational methods usually struggle with prediction performance. To tackle these issues, we propose a novel deep neuralnetwork foranti-CRISPR analysis (AcrNET), which achieves impressive performance. On both the cross-fold and cross-dataset validation, our method outperforms the previous state-of-the-art methods significantly. Impressively, AcrNET improves the prediction performance by at least 15% regarding the F1 score for the cross-dataset test. Moreover, AcrNET is the first computational method to predict the detailed anti-CRISPR classes, which may help illustrate the anti-CRISPR mechanism. Taking advantage of a Transformer protein language model pre-trained on 250 million protein sequences, AcrNET overcomes the data scarcity problem. Extensive experiments and analysis suggest that Transformer model feature, evolutionary feature, and local structure feature complement each other, which indicates the critical properties of anti-CRISPR proteins. Combined with AlphaFold prediction, further motif analysis and docking experiments demonstrate that AcrNET captures the evolutionarily conserved pattern and the interaction between anti-CRISPR and the target implicitly. With the impressive prediction capability, AcrNET can serve as a valuable tool for anti-CRISPR study and new anti-CRISPR discovery, with a free webserver athttps://proj.cse.cuhk.edu.hk/aihlab/AcrNET/.
Computational design and construction of an Escherichia coli strain engineered to produce a non-standard amino acid
Authors: Zomorrodi Ali R. , Hemez Colin , Arranz-Gibert Pol , Wu Terrence , Isaacs Farren J. , Segrè Daniel 
Published: April 2022

SummaryIntroducing heterologous pathways into host cells constitutes a promising strategy for synthesizing nonstandard amino acids (nsAAs) to enable the production of proteins with expanded chemistries. However, this strategy has proven challenging as the expression of heterologous pathways can disrupt cellular homeostasis of the host cell. Here, we sought to optimize the heterologous production of the nsAA para-aminophenylalanine (pAF) in Escherichia coli. First, we incorporated a heterologous pAF biosynthesis pathway into a genome-scale model of E. coli metabolism, and computationally identified metabolic interventions in the host’s native metabolism to improve pAF production. Next, we explored different ways of imposing these flux interventions experimentally and found that the upregulation of flux in chorismate biosynthesis pathway through the elimination of feedback inhibition mechanisms could significantly raise pAF titers (∼20 fold) while maintaining a reasonable pAF yield-growth rate trade-off. Overall, this study provides a promising strategy for the biosynthesis of nsAAs in engineered cells.
Smartphone‐based colorimetric determination of gamma‐butyrolactone and gamma‐hydroxybutyrate in alcoholic beverage samples
Authors: Procida Anselmo , Honeychurch Kevin C. 
Published: April 2022

AbstractGamma‐hydroxybutyrate (GBH) is a popular recreational drug. Its strong sedative and amnesic effects have led to drug‐facilitated sexual assaults, poisonings, overdose, and death. As a result, legislation has restricted its availability leading to GHB, consumers switching to its pro‐drug, gamma‐butyrolactone (GBL). Consequently, there is a growing need for methods capable of their determination in complex samples such as beverages. Previous studies have shown the possibility to colorimetrically qualitatively determine both GBH and GBL by the formation of the lactone and its reaction with hydroxylamine and ferric chloride to give a purple‐colored complex. In this present investigation, we have shown the possibility of using this approach to both quantify GBL and GHB using both UV/Vis spectrometry and by the application of the camera of a smartphone to record images of the purple color developed. Via subsequent use of a downloadable free App, to extract the numerical values of the Red, Green, and Blue (RGB) color components, it was shown possible to construct a calibration curve and to quantitatively determine the concentration of the drugs present in fortified alcoholic beverage samples. It was found that by simple mathematical normalization of the RGB values the effects of camera distance and elimination could be readily overcome. Using the smartphone approach, GBL determinations on a sample of lager beer gave a mean recovery of 103% (%CV = 0.70%, n = 5) at a concentration of 0.56 mg/ml indicating the method holds promise for the determination of GBL and GHB in such samples.
Phytochemical profile of differently processed tea: A review
Authors: Wong Melody , Sirisena Sameera , Ng Ken 
Published: April 2022

AbstractFresh tea leaves (Camellia sinensis (L.) O. Kuntze) are processed by various techniques to produce different types of tea. The most common way to classify tea types is based on the similarities in processing methods resulting in the five commonly recognized tea types: white, green, oolong, black, and pu‐erh teas. The differences in the degree and nature of fermentation of tea leaves lead to different chemical changes depending on the processing method. Understanding the phytochemical profile of differently processed tea is important, as tea types classified by processing methods are currently not well defined because the chemical parameters for these tea types are still not established. Therefore, any significant characteristics found for a tea type due to processing may be helpful in defining tea types. However, the evidence on the impact of tea processing on phytochemical profile and contents in differently processed tea is currently unclear. Therefore, this review aims to examine (1) the processing techniques of white, green, oolong, black, and pu‐erh tea, (2) the impact of tea processing on tea phytochemicals, and (3) the key characteristics associated with the phytochemical profiles of differently processed tea.Practical ApplicationTea (Camellia sinensis (L.) O. Kuntz) is the most widely consumed beverage in the world. Tea consumption has been demonstrated through in‐vitro experiments and in animal and human intervention studies to exhibit potential in preventing various oxidative stress‐related chronic diseases, such as cardiovascular diseases, Alzheimer.s disease, diabetes and certain cancers. Based on the processing methods, tea is commonly categorized into white, green, oolong, black and pu‐er tea. However, there are large overlap in processing methods between some teas and, more importantly, the chemical compositions of differently processed teas are highly variable. This review aims to examine (1) how white, green, oolong, black and Pu‐erh tea are processed, (2) what are the effects of tea processing on tea phytochemicals and (3) to identify whether there are key characteristics associated to the phytochemical profiles of differently processed teas. The review will contribute to tea research in collating in one article the state of knowledge on the chemical changes and composition of the differently processed teas, and point to future direction in this area of research.
In vitro digestion effect on CCK and GLP‐1 release and antioxidant capacity of some plant‐based milk substitutes
Authors: Aly Esmat , Sánchez‐Moya Teresa , Darwish Aliaa A. , Ros‐Berruezo Gaspar , López‐Nicolás Rubén 
Published: April 2022

AbstractRecently, plant‐based milk substitutes, as an emerging industry, are receiving more attention. Despite that, these dairy alternatives have not been adequately studied for their functional properties. Thus, the current research was devoted to study the satiety potential through in vitro secretion of cholecystokinin (CCK) and glucagon‐like peptide‐1 (GLP‐1), and the antioxidant capacity of these dairy alternatives after in vitro digestion. The enteroendocrine cell line, STC‐1, was used to measure satiety hormones release (CCK and GLP‐1) by enzyme‐linked immunoassay (ELISA). Also, total phenolic and flavonoid contents and antioxidant capacity (using oxygen radical absorbance capacity [ORAC], ferric reducing antioxidant power [FRAP], and 2,2′‐azino‐bis(3‐ethylbenzothiazoline‐6‐sulfonic acid [ABTS] assays) were measured before and after in vitro digestion. The results demonstrated that CCK secretion was significantly (p < 0.05) higher for cow's milk (350.64 pg ml–1) as compared to plant‐based milk substitutes. Among the plant‐based milk substitutes, tiger nut milk showed the highest CCK stimulant (228.96 pg ml–1), followed by hazelnut milk (220.04 pg ml–1). Concerning GLP‐1 release, the data exhibited that spelt milk was the food with the highest induction of GLP‐1 hormone secretion, followed by cow's milk (910.17 and 876.59 pg ml–1, respectively), but without any significant differences between them. total phenolic content (TPC) values strongly increased after in vitro digestion, cow's milk and soymilk being the samples with the highest TPC values after in vitro digestion (165.76 and 153.71 mg GAE/100 ml, respectively). In line with TPC values, soymilk had the highest ORAC, ABTS, and FRAP values after in vitro digestion (25.41, 8.17, and 2.51 µmol TE/ml, respectively). Thus, these dairy alternatives could be an adequate substitute for cow's milk, according to its satiety and antioxidant capacity.Practical ApplicationThe first study deals with the efficacy of plant‐based milk substitutes as satiating foods. Spelt milk was the highest stimulant for GLP‐1 secretion. Tiger nut milk showed the highest CCK stimulant, followed by hazelnut milk. TPC and total antioxidant capacity increased after digestion.
Collagen peptides from Acaudina molpadioides prevent CCl4‐induced liver injury via Keap1/Nrf2‐ARE, PI3K/AKT, and MAPKs pathways
Authors: Li Jie , Li Yan , Lin Saijun , Zhao Wei , Chen Yan , Jin Huoxi 
Published: April 2022

AbstractCollagen peptide from Acaudina molpadioides (AMP) showed antioxidative activity in H2O2‐induced RAW264.7 cells in our pervious study. In this study, it was observed that AMP could effectively improve the morphology and function of liver in CCl4‐induced mice. After 200 mg/kg AMP treatment, the content of MDA in liver decreased by 62.3%, and the level of antioxidant enzymes (SOD, GSH‐Px, and CAT) increased by more than 65%. Western blot results disclosed that AMP (200 mg/kg) upregulated the Nrf2 level by 73.8% and downregulated Keap1 by 41.0% in CCl4‐induced mice liver. The levels of p‐ERK, p‐JNK, and p‐p38 in 200 mg/kg AMP treatment groups decreased by 57.3%, 40.9%, and 40.6%, but the levels of p‐PI3K and p‐AKT increased by 162.6% and 60.3%, respectively. Furthermore, the trends of Nrf2, Keap1, p‐ERK, p‐JNK, p‐p38, p‐PI3K, and p‐AKT levels in H2O2‐induced RAW264.7 cells after AMP treatment were similar to the results in CCl4‐induced mice liver. These findings provided evidence that AMP exerted antioxidant activity via Keap1/Nrf2‐ARE, PI3K/AKT, and MAPKs pathways in vivo and in vitro. Therefore, the collagen peptide from A. molpadioides might represent a novel functional food to prevent acute liver injury via attenuation of oxidative stress.