bluuebunny/crossref_metadata_2025_split huggingface dataset:
Regulation of 2’‐5’‐oligoadenylate synthetase 1 (OAS1) by small double‐stranded RNAs
Authors: Schwartz Samantha L. , Conn Graeme L. 
Published: June 2021

The innate immune system comprises a broad collection of critical intra‐ and extracellular processes that limit the infectivity of diverse pathogens. The 2’‐5’‐oligoadenylate synthetase (OAS) family of enzymes are important sensors of cytosolic double‐stranded RNA (dsRNA), and play a critical role in limiting viral infection by initiating signaling to halt viral replication and establish an antiviral state in the cell. Further attesting to the importance of the OAS/RNase L pathway, diverse viruses have developed numerous distinct strategies to evade the effects of OAS activation. Like other innate immune proteins, OAS must be able to accurately distinguish “self” from foreign molecules. Misregulation of the innate immune system can cause increased persistence and susceptibility to viral infection and human diseases, such as interferonopathies. Several recent studies have provided important new insights into the molecular mechanisms of OAS activation by dsRNA as well as unanticipated findings of specific RNA sequences and structural motifs that strongly enhance activation of at least one form of OAS. Although X‐ray crystal structures have given some insight into how OAS1 is activated by dsRNA, we still do not completely understand how specific features of the dsRNA contribute to the level of OAS1 activation. To address this question, we designed dsRNAs to test the impact of specific sequence changes within a strongly activating short (18 base pair) dsRNA. Remarkably, while a single point mutation on one strand resulted in complete loss of OAS1 activity, the equivalent mutation on the opposite strand led to increased OAS1 activity. Despite these stark differences in ability to activate OAS1, both variants appear to bind OAS1 with similar affinity. Given these preliminary findings, I hypothesize that dsRNAs may contain competing OAS1 binding sites with remarkably different capacities to activate the protein in a context‐dependent manner. However, the molecular signatures defining these sites as activating and non‐activating are unknown. On‐going studies to define these features will enhance our understanding of host‐pathogen interactions, such as how viruses might circumvent the OAS1/RNase L pathway by masking activating RNA motifs to evade detection.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Light sensitivity of the photoreceptor cryptochrome of the Drosophila circadian clock and its interaction with other clock components
Authors: Lin Changfan 
Published: June 2021

Flavoprotein cryptochromes (CRYs) are blue light sensors that regulate the circadian clock in plants and animals. They consist of a highly conserved photolyase homology region (PHR) and a C‐terminal tail extension (CTE) of various sizes. In Drosophila CRY, photoreduction of the PHR bound cofactor FAD triggers conformational changes within CRY, which include undocking of C‐terminal Tail (CTT) helix. The mechanism of FAD photoreduction remains a key question in understanding CRY activation.Four tryptophan residues (W420, W397, W342, W394) mediate electron transfer for flavin photoreduction. We have substituted these residues to both redox‐inert phenylalanine residues (F) and redox‐active tyrosine (Y) residues to tune the light sensitivity of CRY. Light‐sensitivities of the variants correlate well with the extent of conformational change at the CTT. Interestingly, the surface tryptophan W394 is the most indispensable residue because it gates electron transfer from exogenous reductant to the photoreduction chain.In flies, CRY interacts with other signaling proteins of the circadian clock in a light depend‐manner to entrain the 24 hour‐cycle to environmental light. For example, in light CRY binds the protein timeless (TIM) and recruits the E3 ligase jetlag to facilitate TIM degradation. However, the molecular interactions between CRY and TIM are not well understood due to the low stability of this protein complex. We have utilized a chemical cross‐linking strategy to map the interfacing domain in this assembly and stabilize it for future structure study by structural methodsSupport or Funding InformationThis work was supported by grants from the NIH: R01GM054339 (to M.W.Y.) and R35GM122535 (to B.R.C.).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Kinetic Basis for Selective Inhibition of Hepatitis C Virus RNA‐Dependent RNA Polymerase by Nucleoside Analogs
Authors: Villalba Brian , Li Jiawen , Johnson Kenneth 
Published: June 2021

The Hepatitis C Virus (HCV) infects an estimated 71 million people worldwide. Currently, there is no vaccine available to treat or prevent HCV infections. Nucleoside analogs (NA) with a modification on the 2′‐carbon are a group of drugs that target the HCV RNA‐dependent RNA polymerase, NS5B. Among these nucleoside analogs are the prodrug Mericitabine and Sofosbuvir which get metabolized to 2′‐fluoro‐2′‐C‐methylcytidine‐5′‐triphosphate and 2′fluoro‐2′‐C‐methyluridine‐5′‐triphosphate, respectively. These compounds work by mimicking natural nucleotides and competing for binding and incorporation during the replication of the HCV genome. Once incorporated they act as chain terminators by preventing further elongation of the nascent RNA strand. Recently, it has been shown that NA can be excised by a nucleotide‐mediated excision reaction which rescues RNA synthesis. In this study we use transient‐state kinetics to understand the mechanism of inhibition for two CTP analogs and a UTP analog. The results demonstrate that while CTP analogs are readily incorporated and excised, UTP and the UTP analogs efficiently incorporated but are resistant to excision.Support or Funding InformationThis work was supported by funding from the NIHThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Using Enzyme Kinetics and Small Angle X‐ray Scattering to Understand the Allosteric Regulation of SIRT1
Authors: Wang Ningkun 
Published: June 2021

SIRT1 is an NAD+ dependent deacetylase located in the nucleus and cytoplasm of the cell. Research has shown that SIRT1 has been implicated to be involved in many important cellular functions such as neurodegenerative diseases and aging. SIRT1 has also been studied with resveratrol, a polyphenol, which was shown to modulate SIRT's enzyme activity. While the mechanism of SIRT1 has been studied extensively, some exact details are still not known, specifically regarding the allosteric regulation by the N‐terminal domain. We hypothesize that there is a correlation between the activity of SIRT1 and the dynamics of the N‐terminal domain relative to the catalytic core. We are testing this hypothesis by enzyme kinetics and small angle X‐ray scattering (SAXS) methods, using resveratrol as a tool to vary SIRT1 activity. A recent study examined the activity of SIRT1 on peptides of different acetylation sites and found that resveratrol could either increase, decrease or not change SIRT1 activity. We chose peptides from each of these categories as substrates to produce a range of SIRT1 activity change upon resveratrol addition: increase, no change and decrease. First we examined the kinetic parameters (kcat and KM) of SIRT1 with these peptide substrates with and without resveratrol. We are using a continuous enzyme‐coupled assay, rather than an endpoint assay used in the previous study, to see if whether kcat or KM contributes more to the activity change. We also studied the conformational dynamics of SIRT1 in the presence of these peptides and resveratrol using SAXS. The SAXS data informed us about the folding state and overall conformation of the protein, where most changes in conformation would be due to large motions of the more dynamic N‐terminal domain. Combining the data from Michaelis‐Menten enzyme assays and SAXS, we hope to identify a correlation between the conformational dynamics of the N‐terminal domain and the enzyme activity of SIRT1. Learning more about the allosteric mechanism of SIRT1 could aid in the design of therapeutic targets in modulating SIRT1 activity in the cell.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
EU‐OPENSCREEN ‐ Innovative collaboration models for molecular biologists in early drug discovery
Authors: Stechmann Bahne 
Published: June 2021

As of now, the majority of molecular biologists do not have ready access to suitable drug screening platforms and compound collections, which are generally expensive to purchase, operate and maintain. This represents a major limitation in the field to advance innovative drug discovery projects in academia.With the aim to address this need, researchers from >20 institutions have joined forces to launch ‘EU‐OPENSCREEN’ (www.eu‐openscreen.eu), which offers international scientists access to a wide range of state‐of‐the‐art technology platforms, resources and expertise. This publically funded initiative integrates high‐capacity screening platforms and medicinal chemistry groups throughout Europe on a not‐for‐profit basis.The aim of EU‐OPENSCREEN is to collaboratively develop novel molecular tool compounds with external researchers from various disciplines of the life sciences and to support researchers seeking a better understanding on how fundamental molecular processes act to govern biological function at the organismal, tissue, cellular and pathway levels.Molecular biologists and biochemists who have a robust and suitable assay, and are interested in developing novel chemical tool compounds (inhibitors or activators) for their target of interest, will be able to access access the EU‐OPENSCREEN's screening platforms on a collaborative basis and to screen the rationally selected EU‐OPENSCREEN compound collection, comprising up to 140.000 commercial and proprietary compounds collected from European chemists. After the primary screening, EU‐OPENSCREEN's medicinal chemists optimise the identified ‘hit’ compounds during the hit‐to‐probe program.I will explain how international pharmacologists and molecular biologists can access and collaborate with EU‐OPENSCREEN to advance their research projects.Support or Funding InformationThis project has received fundng from the European Union's Horizon 2020 research and innovation programme under grant agreement No 654248.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chromosomal Instability Promotes cGAS‐Mediated Cytosolic DNA Response in Metastatic Cancer
Authors: D'Ausilio Morgan E. , Agrawal Anushka , Karande Mira , Kolarov Teodora , Kushen Jemma , Lin Jessica , Mandel Matthew , Patel Sahdev , Pollard Sophie , Bakhoum Samuel F. 
Published: June 2021

Recent implication of chromosomal instability (CIN) in metastatic cancer cells has pointed to the frequency of chromosome missegregation as a key difference between primary and metastatic tumors. Understanding the cellular response therefore aids development of treatments to prevent metastasis. Chromosome missegregation creates micronuclei of double stranded DNA (dsDNA) fragments that are prone to rupture in the cytoplasm. The cGAS‐STING pathway responds to cytosolic dsDNA by activating inflammatory response pathways, a reaction useful to protect against viruses. Dimerization of cGAS, induced by dsDNA binding, catalyzes synthesis of cyclic GAMP (cGAMP) from ATP and GTP. Activation of cGAS results in upregulation of IFN‐β and NF‐κB stimulated immune response pathways. Cytosolic dsDNA associated with CIN has been proposed to induce cGAS‐STING mediated immune mimicry in cancer cells, allowing tumors to flourish in otherwise lethal environments. The Pingry School SMART Team, in conjunction with MSOE Center for BioMolecular Modeling, is using 3‐D modeling and printing technology to further examine the binding of cytosolic dsDNA to cGAS and the resultant production of the signaling molecule, cGAMP, by the enzyme. Stimulation of a cGAS‐STING inflammatory response due to CIN in cancer may contribute to the metastatic transition of primary tumor cells. The Pingry School SMART Team aims to utilize the cGAS structural model to better understand the role of this protein in responding to CIN and its link to metastasis through cytosolic dsDNA induced immune response.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Photobiomodulation on Apoptosis Process After Induced Myocardial Infarction in Rats: Analysis of Gene Expression
Authors: Maretti Joao Victor , Manchini Martha Trindade , Silva Jose Antonio , Serra Andrey Jorge , Feliciano Regiane Santos 
Published: June 2021

Cardiovascular Diseases (CVD) are the main cause of death in Brazil and in the rest of the world. Ischemic heart diseases represent 30% of deaths by CVD in occidental countries. It is well known that Myocardial Infarction (MI) harms contraction and is responsible of a hypertrophy process of the cardiomyocytes, besides the reduction of ventricle wall thickness, modifying extracellular matrix and promoting metabolic changes. Those metabolic processes stimulate a compensatory rearrangement by targeting three different morphologic regions: infarcted area, borderline area and the remote area to the MI. New treatments are aimed at the modulation of various cardiac processes, such as cardiac muscle contraction, metabolism, apoptosis, fibrosis formation and control of hypertrophy, with the objective of revert or prevent damage on those heart areas. Nowadays, Low‐Level Laser Therapy (LLLT) has become an alternative treatment by its capacity of modulation various biological processes and, depending on dose and wavelength parameters, anti‐inflammatory proprieties and faster cell proliferation. However, the action of the LLLT has not yet been elucidated and little is known about the remaining myocardium behaviour on this treatment, specially when related to apoptosis. Thus, the present study aims the evaluation of LLLT effects on genes related to apoptosis, by analysing TP53, MAP3K2, Bax, MAPK14 and Fas mRNA. The study, approved by the local ethics committee, was designed by the promotion of anterior descending coronary artery occlusion and posterior laser irradiation in the remote region to the myocardial infarction in rats, which were submitted to an echocardiography to evaluate myocardial function after therapeutics. Heart samples underwent an analysis by Real‐Time Polymerase Chain Reaction Array. Data showed that LLLT was capable of modulating apoptosis process, by downregulating TP53 gene, when compared to rats which underwent only anterior descending coronary artery occlusion. The study concludes that LLLT may stimulate cellular survival after MI, by the inhibition of apoptosis of remote region cells.Support or Funding InformationCNPq, CAPES, UNINOVEThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Photobiomodulation in mRNA Expression of Extracellular Matrix Components of Cardiac Fibrosis in Experimental Post‐infarcted Myocardial in Rats
Authors: Costa Thiago Oliveira D'Alcantara , Manchini Martha T , Serra Andrey J , Atum Allan B , Feliciano Regiane S , Silva Jose A 
Published: June 2021

IntroductionMyocardial infarction (MI) is due to irreversible cell death (necrosis) of the heart muscle, secondary to a prolonged inhibition of oxygen supply (ischemia). Fibrosis is the major histologic marker, resulting in heart failure from myocardial infarction. Recently, low intensity laser (LBI) has become an alternative therapy to modulate various biological processes and depending on the wavelength, dosage and condition of the irradiated tissue, has anti‐inflammatory effect, reduces pain and speeds up cell proliferation.ObjectiveTo evaluate the effect of photobiomodulation on genes involved in the formation of post‐MI cardiac fibrosis by analyzing the mRNA of Col1a1, Col3a1, MMP‐1, MMP‐2, MMP‐9 and TIMP‐1 and TIMP‐2.Materials and MethodsFifteen female Wistar rats were divided into three groups: Sham (5), IM (5) and IM + Laser (5) (Ethics Committee: 0015–2012). The infarction induction was made by occlusion of the descending left coronary artery, the laser being applied for 60 seconds at the occlusion site immediately after the infarction. Left ventricle samples from the experimental animals were collected 72h after infarction. Modifications in gene expression of genes: Edn1, Ace, Agtr1a and Ace2 were analyzed by Real‐Time PCR.ResultsThere was a significant increase in the expression of Col1a1 and Col3a1 in 1270.0% and 605.0%, respectively, in the IM group compared to the SHAM group. In contrast, the expression of Col1a1 and Col3a1 was decreased by 51.68 %% and 42.70%, respectively, in the IM + Laser group, when compared to the IM group. There was a significant increase in the expression of MMP‐1, MMP‐2 and MMP‐9 by 240%, 166% and 295%, respectively, in the IM group compared to the SHAM group. Photobiomodulation was able to reduce the expression of all genes mentioned above in 64.12%, 62.78 and 76.71% respectively. We also observed a reduction in TIMP‐1 and TIMP‐2 mRNA expression in 32% and 35%, respectively, in the IM group compared to the SHAM group.ConclusionPhotobiomodulation was able to reduce the expression of COL1A1 and COL3A1 in the IM + Laser group when compared to the IM group, and also related to the reduction of MMPs expression and increase of TIMP‐1 and TIMP‐2 inhibitors, demonstrating the effects of laser therapy in reducing collagen deposition.Support or Funding InformationFAPESP, CNPq, CAPES, UNINOVEThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Comparative Microbiome Analysis of House Dust Mites, the Most Common Cause of Allergens
Authors: Yong Tai‐Soon , Lee Jinyoung , Kim Ju Yeong , Yi Myung‐hee , Lee In‐Yong , Nam Sung‐Hyun , Yong Dongeun 
Published: June 2021

Three species of mites, Dermatophagoides farinae, D. pteronyssinus, and Tyrophagus putrescentiae which are most popular in Korean homes, grown under the same condition would show different patterns of microbiome, and induce different immune reactions. Three species of mites (D. farinae, D. pteronyssinus, and T. putrescentiae), known to cause allergies, are cultured in s same condition (autoclaved media, 80%RH, 25_) and, analyzed for microbiome of each species. Using the next generation sequencing, the difference of microbiome between the dust mite species, and the level of endotoxin were investigated. The extracts of three species of mites were treated to human bronchial epithelial cells (BEAS 2B cells) to measure the expression level of proinflammatory cytokines. Three species of mites, D. farinae, D. pteronyssinus, and T. putrescentiae, grown under the same condition showed significantly different patterns of microbiome. In D. pteronyssinus grown on the autoclaved medium, only a few bacteria were seen in contrast to the other two species. The concentration of endotoxin derived from the extract of mites was proportional to the distribution of the microbiome. The expressed cytokine levels from BEAS 2B cell were comparable among three species of mites. The microbiome pattern of laboratory cultured dust mites differ not on the culture conditions but on the species. The extent to induce Th2 responses in vitro which may closely be related in allergic responses in humans also showed difference.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Metabolomic analysis of a mouse model of NASH and correlation with cytokine expression
Authors: McDermott Caitlin , Eudy Brandon , da Silva Robin 
Published: June 2021

Non‐alcoholic fatty liver disease (NAFLD) is a prevalent condition that begins with excess lipid accumulation, termed steatosis. NAFLD is one of the hallmarks of metabolic syndrome with a complex etiology, and over time can progress to nonalcoholic steatohepatitis (NASH), characterized by inflammation and fibrosis. There is no biomarker that can be used to detect fibrosis and therefore the condition can go undetected for a long time. In addition, there are no accepted medical treatments for NASH and extreme cases may progress to cirrhosis and hepatocellular carcinoma (HCC). Past work has identified associations between Glycine N‐methyltransferase (GNMT) and both fibrosis and HCC. The GNMT knockout mouse phenotype ties one‐carbon metabolism to hepatic inflammation and immune function. In humans NASH typically occurs in the obese type 2 diabetic population, but GNMT mice are neither overweight, nor insulin resistant. Therefore, we used a high‐fat diet to investigate the impact of diet on fibrosis. We used a metabolomic approach to identify candidate molecules that may be used to identify hepatic fibrosis. We also use Luminex cytokine analysis to identify altered pathways and immune response that correlate to metabolic changes. Intriguingly we find that GNMT mice receiving a high‐fat diet do not gain weight compared to the wild‐type mice. Our metabolomic analysis has identified several important pathways that contribute to the GNMT phenotype that warrant further investigation. We find interesting correlations with several cytokines including VEGF, M‐CSF, GM‐CSF, IFN gamma, IL‐2, IL‐6 and IL‐10.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.