bluuebunny/crossref_metadata_2025_split huggingface dataset:
“The Flexible Nucleosome: A Cross‐Kingdom Perspective”
Authors: Mohamed Farzanna A. , Melters Daniël P. , Dalal Yamini 
Published: June 2021

In order to maintain genomic integrity, chromosomes must be segregated faithfully. The centromere is the region of the chromosome that seeds the kinetochore and is epigenetically marked by the centromere‐specific histone H3 variant CENP‐A/CENH3. Although centromeres are functionally essential, their underlying DNA sequences and associated proteins are rapidly evolving. This is known as the centromere paradox. CENP‐A's sequence is highly diverged across species, yet its function to seed the kinetochore remains intact. One possibility is that specific structural features of CENP‐A nucleosomes are conserved across the species. Indeed, in silico, in vivo, and in vitro studies have focused on the characterization of the dynamic structural behaviors of human and yeast CENP‐A nucleosomes. This dynamic behavior is thought to be predominantly driven by its αN‐helix, loop L1, and four‐helix bundle. Recently, we developed a technique to directly assess CENP‐A structural plasticity by using single‐molecule nano‐indentation force spectroscopy. We observed that human CENP‐A nucleosomes are twice as elastic compared to canonical H3 nucleosomes. In order to determine whether this observation is unique to human CENP‐A or represents a universal feature that defines a rapidly evolving CENP‐A nucleosome, we set out to investigate the elasticity of CENP‐As derived from 17 distinct species, including those that have holocentric chromosomes, spanning the three main eukaryotic kingdoms. Here, we present a progress report on our efforts to elucidate structural features underpinning the evolution of centromeric histones and its associated chromatin, which in turn serves as a model for the study of the evolution of epigenetic systems.Support or Funding InformationIntramural Research Program of CCRThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Determining if Mating Type Proteins Play a Role in Immaturity of the Ciliate Tetrahymena thermophila
Authors: Enerson Rebecca Leigh , Cervantes Marcella 
Published: June 2021

Tetrahymena thermophila is free‐living ciliate, commonly found in freshwater ponds, and is used as a model research organism. Mating type gene expression is induced in mature cells after they have been starved and put in the presence of cells of a different mating type (Tetrahymena Genome Database, November 10, 2018). T. thermophila are known to have developmental stages of maturity, and immature cells are unable to mate, although the scientific processes that prevent mating are unknown. It is known that mating type gene expression increases one‐hundred fold from logarithmic phase growth to three hours of starvation (Tetrahymena Genome Database, November 10, 2018). It is currently unknown if the mating type genes are expressed in immature progeny since immature cells do not mate.The goal of this study was to determine if immature progeny cells expressed mating type proteins under a variety of conditions. We hypothesized that mating type genes would not be expressed in immature progeny because T. thermophila will not mate until they reach an level of maturity, and the expression of the mating type genes is essential for mating (Rogers and Karrer, 1985). We plan to quantify the mating type gene expression using q‐RT PCR and RNA Seq transcriptase analysis.This study mated two strains, one mating type VI and II, of T. thermophila and isolated the progeny based on drug sensitivity differences from parental cells. For the q‐RT PCR experiment, mating type gene expression for the parental strain was tested at log phase, thirty minutes of starvation, and two hours of starvation. The mating type gene expression for the progeny strain was tested at the same three conditions as the parental strain. RNA samples from each of the six conditions were isolated and used to create cDNA. The cDNA was confirmed through a PCR containing a genomic control, mating type primers, starvation versus log primers, and negative control primers. Then qPCR was utilized to quantify the expression of mating type genes in the six previously described conditions. RNA Seq was utilized to quantify the expression of all genes for triplicate samples of parental cells and progeny, each starved for 24 hours.Mating type gene expression will be validated through the cross referencing of quantified results provided by RNA Seq and qPCR for T. thermophila and are hypothesized to be lower or absent in progeny samples. There may also be differences in expression levels of transcription factors between the progeny and mature cells. However, if there is no difference in either mating type gene or transcription factors, then prevention of mating in immature cells occurs after transcriptional processes and needs further investigation.Support or Funding InformationAlbion College Foundation for Undergraduate Research, Scholarship, and Creative ActivityThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The role of lncRNA CRNDE in obesity‐associated endothelial dysfunction
Authors: Nguyen Nghi Minh , Moran Matthew , Sun Xinghui 
Published: June 2021

Obesity, a risk factor for type II diabetes and cardiovascular disease affects 35.7% of the U.S. population and costs approximately $210 billion per year. Endothelial cells (ECs) are cells that line blood vessels, are important in signaling and transport and have been implicated in obesity. The role of long non‐coding RNA (lncRNA) in regulating endothelial function, especially under disease conditions, has been understudied. In an RNA‐seq analysis of palmitate‐treated ECs, the lncRNA Colorectal Neoplasia Differentially Expressed (CRNDE) was determined to be upregulated. The purpose of this study is to determine the role of CRNDE in EC function through examination of various signaling pathways and exploring how obesogenic stimuli induce CRNDE expression. To determine the effect of obesogenic stimuli on CRNDE expression, HUVECs were treated with albumin‐conjugated palmitic acids or albumin for 4 or 12 hours and CRNDE expression was determined by qPCR. CRNDE is significantly increased in palmitic acid‐treated HUVECs after 12 hours compared with cells treated with albumin. Bioinformatic analysis of CRNDE and its promoter reveals the presence of p53 binding sites at CRNDE promoter which mediate the induction of CRNDE. To test this, CRNDE expression will be examined in HUVECs in the presence or absence of p53 knockdown. Chromatin immunoprecipitation will be used to determine the binding of p53 at the CRNDE promoter in HUVECs. Loss‐of‐ and gain‐of‐function studies will be performed to examine the signaling pathway that is regulated by CRNDE. Understanding the role of CRNDE in obesity‐associated endothelial dysfunction could provide new targets to reduce vascular complications associated with obesity and diabetes.Support or Funding InformationUCAREThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Analysis of miR‐21‐5p and ‐3p expression and function during pathological neovascularization
Authors: Tripodi Giuseppe Daniele , Thounaojam Menaka C , Bartoli Manuela 
Published: June 2021

Retinal neovascularization (RNV) is a sight compromising complication of diabetes and prematurity. Previous studies from our laboratory have shown that the small noncoding microRNA, microRNA‐21‐5p (miR‐21‐5p), is an inducer of RNV. Emerging new evidence has shown that miRNA passenger strands, the 3p forms, which were thought to be degraded, possess biological functions. In this study, we have investigated the role of miR‐21‐3p in RNV. In particular, we have assessed the effect of miR‐21‐3p on the same molecular targets of miR‐21‐5p: the tissue inhibitor of matrix metalloproteinases 3 (TIMP3), peroxisome proliferator‐activated receptor alpha (PPAR‐α) and pigment epithelium‐derived factor (PEDF). Real‐time PCR (RT‐PCR) showed that the expression of miR‐21‐3p is significantly upregulated in retinas and blood spot samples of mice subjected to oxygen‐induced retinopathy (OIR), a model of RNV. Furthermore, we noted a significant upregulation of miR‐21‐3p levels, measured by RT‐PCR, in human retinal endothelial cells (HuREC) and pigment epithelial cells (ARPE‐19) when exposed to angiogenic stimuli such as 20ng/mL of vascular endothelial growth factor (VEGF) or to hypoxic conditions (pO2= 2+/−0.5) for different times (1, 3, 6, and 12 hours). Transfection of HuREC and ARPE‐19 with miR‐21‐3p mimic (50nM) for 24 and 48 hours had only marginal inhibition on the miR‐21‐5p target genes (TIMP3, PPARα, and PEDF). Additionally, blockade of miR‐21‐3p achieved by transfecting cells with a specific miR‐21‐3p inhibitor (50nM) did not rescue VEGF‐changes in TIMP3, PPARα, and PEDF expression. Our data demonstrated that the passenger strand, miR‐21‐3p, is co‐expressed with miR‐21‐5p in ischemic conditions, but targets different genes. Although further studies will be necessary to clarify miR‐21‐3p biological function, our studies suggest that, together with miR‐21‐5p, miR‐21‐3p may hold potential diagnostic and therapeutic value for treatment of RNV in ischemic retinopathies.Support or Funding InformationAll support for this research came from the Bartoli laboratory of the Ophthalmology Department of the Medical College of Georgia at Augusta University and the funding sources (EY0246, Knights Templar –GA Chapter).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MicroRNA‐92b is a potential therapeutic target for Glioblastoma treatment
Authors: Grafals‐Ruiz Nilmary , Lozada‐Delgado Eunice , Santana‐Rivera Yasmarie , Rivera‐Diaz Monica , Valiyeva Fatma , Vivas‐Mejía Pablo 
Published: June 2021

Glioblastoma is the most aggressive type of primary brain tumor with an overall survival of 14.6 months with standard care treatment (surgery, radiotherapy, and Temozolomide chemotherapy). About 90% of patients develop recurrent tumors due to acquired resistance to temozolomide. Therefore, new multimodal treatments that target more than one molecular pathway are needed. MicroRNAs (miRNAs) are small (18–22 nucleotides) non‐coding RNAs that regulate more than 60% of protein coding genes post‐transcriptionally. Their dysregulation is implicated in many diseases including diabetes, heart disease, and cancer. MicroRNA‐92b (miR92b) is highly upregulated in glioblastoma tumors and its expression has an inverse correlation with patient overall survival. This study aims to determine the biological effect of targeting miR92b in glioblastoma. We hypothesized that its inhibition would lead to cell death and decreased proliferation by allowing the expression of messenger RNA (mRNA) targets needed for normal cell function. To determine miR92b targeting effect in vitro, we transfected glioblastoma cell lines with an anti‐miR92b and evaluated cellular apoptosis, clonogenicity, and proliferation. Results showed that by inhibiting miR92b there was an increase in apoptotic cells and reduction of cell proliferation. We next evaluated the miR92b targeting effect in vivo, administering intraperitoneal injections of anti‐miR92b encapsulated liposomes to glioblastoma‐xenograft mice. Inhibiting miR92b in mice xenografts led to a significant decrease in tumor volume and tumor weight. Furthermore, we performed bioinformatic analysis to identify mRNA potential targets of miR92b. We both inhibited and overexpressed miR92b in glioblastoma cells (U87 and CRL1620 respectively) and determined the expression of more than 90 potential mRNA targets by quantitative PCR. This analysis lead to the identification of five potential miR92b mRNA targets‐ASB5, TEF, KIAA1024, ZNF776, FBXW7 and BAZ2‐ which are currently under further validation using Western blot analysis. Our results show that miR92b is a good therapeutic target, since its inhibition leads to increased cell death and decreased cellular proliferation both in vitro and in vivo. MiR92b is a potential target for glioblastoma patients and its targeting could lead to the regulation of tumor suppressive protein coding genes.Support or Funding InformationNIGMS‐RISE Grant Number R25‐GM061838, RCMI grant U54 MD007600 (National Institute of Health and Health Disparities), and UPR Comprehensive Center Seed Funds.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
lncRNA Chronos Exacerbates Pathological Cardiac Dysfunction and Fibrosis
Authors: Neppl Ronald L 
Published: June 2021

Long non‐coding RNAs (lncRNAs) are a recently discovered class of genes involved in epigenetic modifications, the regulation of transcription and translation, as well as RNA metabolism. Importantly, the expression of many lncRNAs are known to be developmentally regulated with cell and tissue type specificity and to be dysregulated under pathological conditions. Our previous work identified Chronos as a muscle enriched and aging‐induced, Akt1‐inhibited lncRNA that negatively regulates myogenesis. Testing the hypothesis that Chronos expression is deleterious, we generated Chronos deficient mice and subjected them to transverse aortic constriction (TAC) in order to study the role of Chronos in cardiac hypertrophy and myocardial remodeling. Preliminary data indicate that at early timepoints, Chronos deficient mice have less interstitial fibrosis, as indicated by Masson's Trichrome staining and qPCR analysis of extracellular matrix genes, as compared to WT controls, while still exhibiting hypertrophic growth. Pilot studies at late timepoints indicate no statistical difference between WT and Chronos deficient mice with regard to cardiac hypertrophy or interstitial fibrosis. These data suggest that lncRNA Chronos may play a role in the promoting the initial phases of interstitial fibrosis in response to cellular stress, rather than replacement fibrosis following cardiomyocyte apoptosis. Future studies will examine the role of Chronos in both the early and late stages of myocardial remodeling and compensatory cardiac hypertrophy.Support or Funding InformationThis work is supported by the Brigham and Women's Hospital Department of Orthopedic Surgery.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Single Amino Acid Substitutions Dramatically Shift Equilibria of Physiologically Relevant Alternate Protein Assemblies
Authors: Jaffe Eileen K , Saphire Erica Ollmann , Arturo Emilia C 
Published: June 2021

Increasing numbers of proteins are being revealed to “shape shift” in physiologically relevant ways that do not dramatically alter secondary structure. As each has been discovered they have been called morpheeins, transformers, metamorphic polymorphs, and others. In each case the alternate assemblies have different functions ranging from “on vs. off” to “moonlighting”. These examples continue to surprise their discoverers because they defy the “one sequence, one structure, one function” paradigm in an unexpected way. These alternate assemblies are likely physiologically relevant as their interconversion is thermodynamically feasible and the equilibrium of alternate assemblies is responsive to biological signals (e.g. pH, ligands). Experience now teaches that single amino acid substitutions can also dramatically shift an equilibrium of alternate assemblies. In some cases this shift is reasonably hypothesized, or established, to contribute to diseases such as inborn errors of metabolism and cancer.We present examples culled both from our work and the literature of such shape‐shifting proteins whose equilibria respond to single amino acid substitutions. Prime examples come from diverse branches of life, including viral proteins, bacterial proteins, and human metabolic enzymes. The growing number of such examples suggests that alternate assemblies, which have been called the fifth level of protein structure, may be a common factor in protein structure‐function relationships. Experience teaches that this fifth level of structure, for a given protein, may be revealed by the unexpected biophysical behavior of single amino acid variants.Support or Funding InformationNIH R01 NS100081 (EKJ)NIH R01 AI118016 (EOS)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In silico Analyses of the Binding of Experimentally Tested or Potentially Anti‐Prion Drugs to the PrPC Structures of Several Species of Mammals
Authors: Martinez Nelson 
Published: June 2021

Transmissible Spongiform Encephalopathies (TSE's) or prion diseases are a group of rare neurodegenerative diseases that include Creutzfeldt–Jacob Syndrome in humans and Bovine Spongiform Encephalopathy (BSE) in cattle, and which are inevitably fatal. We have made in silico analyses on the docking of several drugs with either experimentally determined or potential anti‐prion activity, to the experimentally determined structures of the cellular isoform of the prion protein (PrPC) from several species of mammals, including humans.ObjectivesThe objectives of our study are, in the first place to identify differences, if any, between the predicted affinities and/or binding regions of the drugs tested to the PrPC structures from several species of mammals including humans, cattle, mouse, rabbit, and the Bank vole and, secondly, to determine whether these differences, if found, have any correlation with the different susceptibilities to infection with scrapie (PrPSc) prion protein from other species that different mammals exhibit.MethodsWe used free web–based software in our analyses, especially AUTODOCK VINA to model the binding of the drugs tested to experimentally determined crystal or NMR structures that are already available in the Protein Data Bank (PDB). We also used the web–based PRODRG platform to generate the structures of the drugs tested in PDB format, the NCBI Blast software for sequence alignments, and the Statdisk software to perform statistical analyses.ResultsSo far, we have not found yet any significant differences in the predicted affinities and/or binding regions for the drugs tested between binding to the Bank vole prion protein structure and binding to PrPC structures from other mammals which are less susceptible to infection with PrPSc from other species.ConclusionsWe have successfully modeled the binding of several drugs with potential anti‐prion activity to the experimentally determined NMR or crystal structures of the cellular prion protein of several mammals, including humans. Our results suggest that the differential susceptibilities to infection between the Bank vole and other species of mammals that have been found by experiments cannot be explained, at least alone, by structural differences between their PrPC isoforms.Support or Funding InformationThe author is a PhD candidate at the Department of Biochemistry, University of Puerto Rico, Medical Sciences Campus, and has received financial support for his studies from the grants NIMHD‐G12MD007600 and 8U54MD007587 to the University of Puerto Rico, and also from grant NIGMS‐RISE‐R25GM061838 and NIGMS‐INBRE.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Macromolecular crowding modulates actin bundle formation induced by actin crosslinking proteins
Authors: Park Jinho , Lee Myeongsang , Kang Hyeran 
Published: June 2021

Bundles of filamentous actin generate high ordered cellular structures including filopodia and lamellipodia with the help of actin binding proteins. α‐actinin and fascin are essential actin crosslinking proteins that affect the mechanical properties of bundles. While the roles of both actin binding proteins in bundling have been well studied in dilute buffer conditions, how they affect bundle formation in crowded environments is not established. Here, we investigate the effects of macromolecular crowding on actin bundles crosslinked by α‐actinin and fascin in vitro and in silico. Time‐dependent bundling processes by both proteins were monitored in the presence of the polymeric crowding agent polyethylene glycol (PEG) using total internal reflection fluorescence (TIRF) microscopy. Biophysical analysis show that PEG reduces the amount and average lengths of α‐actinin‐induced bundles but promotes the formation of thicker bundles induced by fascin. Utilizing molecular dynamics (MD) simulations, we evaluated the interaction between actin filaments and bundling proteins by measuring the contact area and the number of hydrogen bonds. The simulation results demonstrate that PEG increases the binding affinities between actin and binding domain of fascin, while lowering the affinities between actin and α‐actinin. Taken together, our work suggests that crowded environments may modulate actin bundling induced by actin binding proteins in vivo.Support or Funding InformationThis study was supported by the UCF start‐up fund and In‐House grant for Hyeran Kang. We acknowledge the computational time from UCF stokes cluster.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Towards revealing the molecular mechanism of STF
Authors: Pathak Prabhat Kumar 
Published: June 2021

Leaf is a primary organ for the plants to sustain their life and support life on earth by assimilating solar energy and carbon dioxide into chemical energy. STENOFOLIA (STF), a WUSCHEL (WUS)‐like homeobox transcriptional regulator, plays a key role in embryonic patterning, blade outgrowth, and leaf vascular patterning. STF has also been shown to help in increasing biomass and solubilized sugar yield. STF does not affect leaf outgrowth directly but is critical for their development. It is known that transcriptional repressor activity mediated by the WUSbox of STF acts to promote blade outgrowth, and STF executes its functions with its interactions with other protein and DNA. STF binds to GA rich DNA sequence and there is specific DNA binding domain present in the sequence. Although the DNA binding domain (HD) of the protein is conserved across the species, the WOX family HD seems to be unique and cannot be replaced by another HD protein from the plant. WOX HD domain is longer than usual HD domains and has additional protein interaction sites to achieve stable repressive activity. The molecular mechanism of STF function and how WOX genes have adopted the WUS in evolutionary development are unknown. Here we are working towards determining the atomic structure of STF in complex with DNA by using x‐ray crystallography.Support or Funding InformationNIHThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.