bluuebunny/crossref_metadata_2025_split huggingface dataset:
IN VITRO EVALUATION OF THE CYTOTOXIC POTENTIAL OF Securidacalongepedunculata ON HUMAN BREAST ADENOCARCINOMA (MCF‐7) CELLS
Authors: MINARI JOSEPH BAMIDELE 
Published: June 2021

The prevalence of cancer has increased interest in the role of complementary and alternative medicine, employing the use of plant products. Securidacalongepedunculata (SL) serves as a major component of anticancer decoctions in Nigeria. The present study evaluates the cytotoxic potential of the crude extract and fractions of SL root bark against human breast adenocarcinoma (MCF‐7) cell line. The root back of SL was pulverized and extracted with 80% methanol to yield a crude extract which was then submitted to liquid–liquid fractionation with dichloromethane (DCM) and butanol (BUT). The extract and fractions were subjected to Gas Chromatography‐Mass Spectrometry (GC‐MS). MCF‐7 cell line was treated with graded concentrations (9.77 – 5000 μg ml−1) of the aqueous methanol crude extract as well as the DCM and BUT fractions for 24 or 48 hours. Cell viability was thereafter measured by XTT proliferation assay. The activity of the aqueous methanolic crude extract was found to be both dose‐dependent and time independent with IC50 values of 2241μg ml−1 and 1808 μg ml−1 after 24 and 48 hours respectively. However, the DCM and BUT fractions were significantly (p < 0.05) cytotoxic with IC50 values of 86.27 μg ml−1 and 12.08 μg ml−1 respectively after 48 hours of treatment. The study lends credence that BUT fractions of SL may contain lead agent with anticancer properties and that informed its use in alternative medicine.Support or Funding InformationSELF FUNDINGThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterizing the Alternative Splicing Dynamics of Cav1.3, a Calcium Ion Channel Involved in Autism Spectrum Disorder
Authors: Mirkovic Jovan , Sabzanov Samuel , Wang Ruizhi , Hossain Faruk , Ruggiu Matteo 
Published: June 2021

Voltage‐gated calcium channels move calcium ions across cellular membranes in response to action potential stimuli, which mediate processes including cellular signaling, neurotransmitter discharge, muscle contraction, and gene expression. They are multimer proteins composed of alpha‐1, beta, alpha‐2/delta, and gamma subunits. Variety in calcium channels is afforded by different subtypes of alpha‐1, including the A, B, C, D, E, and S isoforms.Cav1.3 is a member of the L‐type calcium channels and is categorized in the high‐voltage activated family. It employs the alpha‐1 subunit encoded by the CACNA1D gene. The alpha‐1 subunit forms four homologous repeat domains, each consisting of six transmembrane helical segments. Cacna1d knock out mice suffer from bradycardia and congenital deafness, testifying to the significance of the alpha‐1 subunit in the sinoatrial node and in auditory hair cells. Naturally occurring missense mutations in alternatively spliced regions of CACNA1D, such as the mutually exclusive exons 8a and 8b encoding part of the sixth transmembrane segment of domain I, have been reported to be gain‐of‐function mutations, which cause aldosterone‐producing adenomas, aldosteronism, seizures, and autism spectrum disorder, highlighting the importance of alternative splicing for Cav1.3 function.Although the diseases derived from CACNA1D mutations have been characterized, the splicing dynamics of the CACNA1D pre‐mRNA that gives rise to the different alpha‐1 isoforms is not well understood. Here, we describe the alternatively spliced isoforms of Cacna1d and their distribution in mouse tissues.Support or Funding InformationNational Institute of General Medical Sciences Grant 1R15GM119099‐01This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
RNA Therapeutic Strategies to Block VEGFR2 Expression and Angiogenesis in Glioblastoma Multiforme
Authors: Muralidharan Koushik , Gawargi Flobater I , Fuoco Kerianne 
Published: June 2021

Glioblastoma multiforme (GBM), a grade IV tumor of the central nervous system, is the most common malignant primary brain tumor, and has a median survival of only 14 months. Poor survival is due to a lack of efficacy in current therapies to pass the blood‐brain barrier (BBB). GBM tumors are characterized by angiogenesis, which is essential for tumor growth and survival. Endothelial cells form the walls of new blood vessels, bridging the gap between the growing tumor and the established vasculature. The membrane receptor that activates tumors to recruit endothelial cells to promote neo‐angiogenesis is vascular endothelial growth factor receptor 2 (VEGFR2). Changes in VEGFR2 expression to block its activation would inhibit the development of new blood vessels within the tumor microenvironment. We have designed novel therapies to bypass the BBB and deliver the genetic sequences of anti‐sense RNA molecules to alter the splicing pattern and expression of the VEGFR2 transcript. Critical splice sites, enhancers, silencers and intronic polyadenylation signals within the VEGFR2 pre‐mRNA transcript were analyzed using bioinformatics databases and RNA‐modeling tools. Based on this, nine different antisense sequences were generated to target and block these elements and were cloned into our therapeutic platform vector, pAAV‐U7‐smOPT. This vector directs the antisense RNA therapeutic to the spliceosome machinery. GBM cells were cultured and transfected with the therapeutic vectors, and the total protein and RNA was collected and analyzed. A172 and U87 GBM cells treated with antisense therapy revealed a greater than two‐fold reduction of VEGFR2. Current directions include direct delivery to the central nervous system using an adeno‐associated viral (AAV) gene therapy vector. In addition, we are isolating AAV‐exosomes with the potential to increase tropism toward the tumor microenvironment.Support or Funding InformationBristol‐Myers Squibb, Independent College Fund of New Jersey, Johnson and JohnsonThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Histone H4 Acetyltransferase NuA4 is Required for Appropriate RNA Splicing in Saccharomyces cerevisiae
Authors: Palfini Lawrence M , Maisner Rose , Gunning Thomas , Lin Mengjia M. , Kress Tracy L 
Published: June 2021

Eukaryotes undergo transcription and splicing simultaneously, allowing for coordination and regulation of these two processes. Currently, there are a few known connections between chromatin modification, which regulates transcription, and splicing found in yeast and metazoa1,2, yet there is still much to study. The NuA4 histone acetyltransferase works with Swr1 chromatin remodeling enzyme to promote transcription3–5. First, NuA4 acetylates histone H4, which is required for the recruitment of Swr1. Next, Swr1 inserts the histone variant H2A.Z, which is subsequently acetylated by NuA4. Interestingly, in Saccharomyces cerevisiae, both NuA4 and Swr1 play a key role in the transcription of the ribosomal protein genes7, which make up a large fraction of the intron containing genes. Thus, NuA4, Swr1, and H2A.Z are excellent candidates for coordinating transcription and splicing. A role for H2A.Z in splicing was recently identified6. Interestingly, data from our lab suggest that NuA4 may have functions in RNA splicing that are independent of H2A.Z deposition. Using directed genetic screens we identified positive (suppressive) interactions between splicing factor gene mutations and mutations that catalytically inactivate NuA4. Additionally, using MPE‐seq8 and RT‐qPCR we found that mutations that block the catalytic activity of NuA4 alter the splicing of a subset of RNAs. Our data suggest that NuA4 plays a role in regulating RNA splicing and we are currently performing chromatin immunoprecipitation experiments to determine whether ESA1 catalytic activity impacts the association of splicing proteins with nascent RNAs during transcription. Together, our data support a model in which NuA4 interacts with the splicing machinery to coordinate transcription and splicing.Support or Funding InformationResearch Corporation for the Advancement of Science (Cottrell College Science award no. 20186) National Institutes of Health (R15GM122026).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigating the Basis of Phenotypic Variation in the DNA Damage Response of Acinetobacter baumannii
Authors: DePamphilis Matthew , Ching Carly , Godoy‐Carter Veronica 
Published: June 2021

Classified as an “ESKAPE” pathogen, Acinetobacter baumannii is one of the leading causes of hospital‐acquired (nosocomial) infections. This Gram‐negative opportunistic pathogen is able to survive desiccation and rapidly acquire multi‐drug resistances. The bacterium's successful mutagenic and antibiotic resistant capabilities are linked to its unique regulation of the DNA Damage Response (DDR). In similarity to many other bacteria, the DDR in A. baumannii depends on RecA, a highly conserved recombinase. However, in A. baumannii, the DDR is bimodal and promotes phenotypic variation; meaning that a fraction of cells—with the same genetic information— respond very well to DNA damage while others do not. It is predicted that this bimodal gene expression is a bet‐hedging strategy that contributes to the persistent survivability of A. baumannii by providing a mechanism for acquisition of advantageous mutations, such as those for antibiotic resistance.We have shown that the recA gene transcript contains an extended structured 5′UTR that influences bimodal gene expression. One of the 5′UTR elements, an outer loop, possesses an influential role as a cis‐regulatory element. We have shown that RNA binding proteins (RBPs) bind specifically to the native UTR when DNA damage is induced. We have identified one of these proteins as a chaperonin known for its ability to bind to RNA and stabilize structure. We aim to further investigate these proteins and their sequence specific binding as we hypothesize that the outer loop is a binding site for RBPs, which would loosen and open the UTR to promote translation of the recA transcript. These findings provide insight to recA regulation, which in turn affects the DDR in A. baumannii. Understanding the DDR will permit to devise novel strategies to eliminate A. baumannii from clinical wards.Support or Funding InformationThis work is supported by grants from Northeastern University's Office of Undergraduate Research and Fellowships and Northeastern University's Honors Department.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Modifying Mouse Herpesvirus‐Induced Vasculitis and Serpin Anti‐Inflammatory Treatment ‐ Profound Effects of the Gut Microbiota
Authors: Lucas Alexandra , Yaron Jordan , Zhang Liqiang 
Published: June 2021

IntroductionInflammatory vascular syndromes (IVS) are rare but devastating arterial disorders. A lethal IVS model can be induced by mouse gamma herpesviral (MHV68) infection in interferon gamma receptor knockout mice (IFNγR−/−). Previously, a serine protease inhibitor (serpin), virus‐derived Serp‐1 protein and Serp‐1 reactive center loop (RCL) peptide S‐7 (G305TTASSDTAITLIPR319) significantly improved survival and reduced aortic inflammation after MHV68 infection. As the microbiome is reported to have profound effects on immune response, we investigated the role of the gut bacteria in MHV68‐infected IVS mouse models with and without treatment.MethodIFNγR−/− mice (N=56) were split into control or 10‐day antibiotic groups prior to infection with MHV68. Mice were treated with saline, Serp‐1 or RCL peptides S‐2, S‐7, S‐8 or modified S‐7 peptides MPS‐8 and MPS‐9. Fecal pellets were collected and 16S rRNA was Illumina sequenced from genomic DNA extracts.ResultsSuppression of gut bacteria accelerated mortality and reduced survival from 60 to 20 days (P=0.036) in MHV68 IVS. Survival was reduced from 70% at 150 days to 20% at 30 days with Serp‐1 treatment (P=0.003) and 0% at 30 days with S‐7 treatment (P<0.0001), while S‐2 was inactive and S‐8 trended towards improvement (N=14). In contrast, modified S‐7 RL peptides, MPS‐8 and MPS‐9, improved survival after suppression of gut bacteria (P<0.001, N=10). Both Serp‐1 and S‐7 significantly altered microbiome community structure (beta‐diversity), while S‐7 also altered community richness (alpha‐diversity), in the large intestine versus saline controls. Lactobacillales were significantly reduced and Streptophyta increased in MHV68 infection with saline treatment, while Bacillales were significantly reduced and Lactobacillales increased in Serp‐1 treatments. Treatment with normal gut micorbial gavage improved survival after antibiotic suppression of gut bacteria.ConclusionMicrobiome richness, structure, or sub‐population composition, play a crucial role in modulating lethal vasculitic syndromes and treatment response. Serp‐1 and S‐7 show significant therapeutic benefit in MHV68 infection‐induced IVS in mice, which is abolished by gut microbiome suppression. MPS‐8 and MPS‐9 recover efficacy in this model. Significant microbiome changes occurred in Serp‐1‐ and S‐7‐treated, gut bacteria‐suppressed mice.Support or Funding InformationArizona State University ‐ Start up funding American Heart Association Grant in aid National Institutes of Health RO1This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Structure of the WFIKKN2 Follistatin Domain and GDF8 Antagonism
Authors: McCoy Jason C , Thompson Tom 
Published: June 2021

Growth Differentiation Factor 8 (GDF8), also known as Myostatin, is a member of the TGF‐β superfamily and is a potent negative regulator of skeletal muscle growth. Disruption of GDF8 signaling or injection of GDF8 inhibitors results in muscle hypertrophy. Muscle mass can also be stimulated through the up‐regulation of secreted GDF8 protein antagonists, such as Follistatin. In contrast, transgenic overexpression of GDF8 causes muscle wasting. Thus, in animals the balance of muscle mass is highly regulated by the appropriate levels of active GDF8, both during development and in the adult. Given that GDF8 plays a profound role on muscle biology there has been a strong interest in developing therapeutics that target GDF8 to alleviate muscle wasting in a variety of diseases or to treat cancer cachexia where tumors have been shown to upregulate GDF8. As such, anti‐GDF8 therapeutics are within phase II and III clinical trials that directly target GDF8 or block signaling receptors. GDF8 is inhibited by a number of extracellular antagonists, one such antagonists, WFIKKN2, exhibits exceptional specificity for GDF8 and GDF11 (90% identity). Similar to the less specific Follistatin family of antagonists, WFIKKN2 utilizes a number of small modular domains including at least one follistatin (FSD) domain. Although there is vast knowledge regarding the Follistatin family FSDs and how they contribute to GDF8 antagonism very little is known about how the WFIKKN2 FSD interacts with GDF8. To address this, we isolated the WFIKKN2 FSD and characterized its binding interaction with GDF8 and GDF11. In addition, we solved the crystal structure of the WFIKKN2 FSD to 1.39Å. The structure revealed conserved, surface exposed, hydrophobic residues that when mutated to alanine reduced full‐length WFIKKN2 antagonism of GDF8. Comparison of the WFIKKN2 FSD to follistatin and FSTL3 indicates that WFIKKN2 utilized a different surface of the FSD to interact with GDF8. Further comparison reveals drastic differences in the domain arrangement classified as either an open or closed conformation. Collectively, the results highlight that although FSDs have conserved domain architecture they exhibit unique functions in biology.Support or Funding InformationNIH RO1: GM114640 American Heart Association: 18PRE33990312This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Dimerization of Yeast Ste14 Isoprenylcysteine Carboxyl Methyltransferase
Authors: Ratliff Anna , Bains Sahej , Funk Amy , Griggs Amy , Hrycyna Christine 
Published: June 2021

Isoprenylcysteine carboxyl methyltransferase (Icmt) is an integral membrane protein localized to the endoplasmic reticulum that is responsible for the post‐translational α‐carboxyl methylesterification of the C‐terminus in CaaX proteins. CaaX proteins (where C is Cys, a is generally an aliphatic amino acid and X is one of several amino acids) include a variety of regulatory proteins, most notably Ras. The Icmt from S. cerevisiae, Ste14, is comprised of six transmembrane (TM) domains in which TM1 contains a putative dimerization motif, G31XXXG35XXXG39, where G is a glycine amino acid residue and X is a subset of hydrophobic amino acids. Ste14 has been shown biochemically to form functional homodimers or higher order oligomers, yet further studies must be performed to better understand this mechanism. To explore these elements as possible sequence and/or structural determinants for dimerization, we used cysteine‐scanning mutagenesis to generate a library of single cysteine mutants for each residue in TM1 (amino acids 25–42). These TM1 cysteine mutants were characterized with immunoblot analyses to assess protein expression, activity, stability, and the ability to form dimers. We determined that residues S27, Y28, L30, G31, G35, and G39 are critical for both methyltransferase activity and the structural stability of the protein. Interestingly, several of the TM1 cysteine mutants that are proposed to lie on the same face of an α‐helix in a helical wheel diagram formed dimers upon the addition of sulfhydryl specific cross‐linkers. We further validated the homodimerization of Ste14 by size exclusion chromatography, multi‐angle light scattering and small‐angle X‐ray scattering. Together, these data suggest that the active form of the Ste14 Icmt is, in fact, a homodimer and these data will ultimately be useful as we further explore the mechanism of action of Ste14.Support or Funding Information1. Purdue Research Foundation2. National Institutes of Health/NIGMS (Grant #: 5R01GM106082‐05)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Genotype on the Ergogenic Effects of Caffeine in Collegiate Distance Runners
Authors: Kline Gabriel M , Millard Julie T 
Published: June 2021

Although caffeine is the world's most popular stimulant, used in both athletic and cognitive contexts, its effects vary widely between individuals. In athletics, caffeine effects depend on dose but may include increases in lipolysis, pain tolerance, and time to exhaustion. While the effects of caffeine on cognitive performance are mixed, caffeine consumption has been linked to increased performance on visual processing tasks and reaction time tasks. Despite the evidence supporting the beneficial effects of caffeine on athletic and cognitive performance, variation in genes related to caffeine metabolism and mechanism of action may moderate its performance‐enhancing effects. For example, a single nucleotide polymorphism (rs762551) of the gene for the liver enzyme CYP1A2 has been reported to influence caffeine outcomes, with “slow” metabolizers having reduced benefits of caffeine during exercise and increased risk of heart attack. Similarly, a single nucleotide polymorphism (rs5751876) of the adenosine receptor gene ADORA2A has been reported to modulate caffeine consumption habits, caffeine‐induced anxiety, and the ergogenic effects of caffeine. In this study, 12 collegiate distance runners were genotyped at CYP1A2 and ADORA2A loci. To assess the effects of caffeine on athletic and cognitive performance, participants underwent two treadmill exhaustion tests and two cognitive battery tests, with one trial being under the influence of low dose caffeine and the other a placebo trial. This work may contribute to a better understanding of the variable effects of caffeine consumption on athletic performance, as well as inform the timing and use of caffeine for optimal performance enhancement.Support or Funding InformationSupported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM0103423.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
CYP 2D6* 4 polymorphism in Polycythemia vera patients in Turkish population
Authors: Cifci Kezban Ucar , Okat Zehra 
Published: June 2021

Studies have shown the association of susceptibility to several cancers with gene encoding enzymes' polymorphisms which engage in xenobiotics' biotransformation. In this study, our main motivation is to determine the relation between cytochrome P450 (CYP) 2D6* 4 polymorphisms and Polycythemia vera (PV) incidence in Turkish population. When patients and controls were compared in terms of CYP 2D6* 4 poor metabolizer (PM) and heterozygous extensive metabolizer (HEM) genotype frequency, it has been found that the patients have 1.35% PM and 32.43% HEM frequency, and controls have 2.63% PM and 21.05% HEM frequency. The correlation between the results of CYP 2D6 genotype analysis and the risk of disease in patients and controls was examined. We found that PM, HEM and extensive metabolizer (EM) genotypes were unrelated with the risk of PV (OR 0.51, 95% CI 0.04–5.71, OR 1.80, 95% CI 0.86–3.75, χ2 1.93, P: 0.164, OR 0.61, 95% CI 0.30–1.24, χ2 1.40, P: 0.235). Our results suggested that the CYP 2D6* 4 polymorphism, in the studied population, does not play an important role in PV etiology.Support or Funding InformationThe funding for the research came from Marmara University Scientific Research Projects Coordination Unit under the project SAG‐C‐DRP‐130515‐0162. Part of the study was published in Marmara Medical Journal in 2018.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.