bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Time Course Study Examining the Effects of Tributyltin Exposures on eIF4E
Authors: Brown Shyretha , Whalen Margaret 
Published: June 2021

Tributyltin (TBT) is an environmental contaminant which is found in human blood (ranging as high as 261 nM) and other tissues due to its lipophilic nature, and it has been shown to increase incidences of tumors in mammals. TBT alters secretion of pro‐inflammatory cytokines such as interferon gamma (IFNγ), tumor necrosis factor alpha (TNFα), interleukin 1 beta (IL‐1β), and interleukin 6 (IL‐6) from human lymphocytes. Previous ex vivo studies have shown that when TBT is introduced as a stress inducer to the cellular environment, the production (secretion and/or intracellular levels) of IL‐1β and IL‐6 was increased in human lymphocytes, and ERK1/2 and p38 MAPK pathways were shown to be necessary for this TBT effect. An additional study indicated that TBT‐induced increases in IL‐1β and IL‐6 production cannot be explained by increases in their respective mRNA at concentrations below 50 nM. Thus, further studies were carried out to address whether the changes in IL‐1β and IL‐6 production induced by TBT might be due to alterations of mRNA translation. The ERK1/2 pathway has been noted to catalyze the phosphorylation (activation) of eukaryotic initiation factor 4E (eIF4E), which is a key player of binding the 5′ cap of mRNAs. As previous studies have indicated a role for the ERK1/2 pathway in the TBT‐induced increases in IL‐1β and IL‐6 production, we carried out initial studies to examine the levels and phosphorylation state of eIF4E after 10 min, 30 min, 6 h, and 24 h exposures to TBT in monocyte‐depleted peripheral blood mononuclear cells (MD‐PBMCs) using western blot analysis. Results suggested that increased phosphorylation (activation) of eIF4E was seen after 10 min exposure to TBT and maintained after 24 h exposure. These results may indicate that eIF4E is a key factor for regulating translation when MD‐PBMCs are exposed to TBT.Support or Funding InformationSupported by NIH grant U54CA163066.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification of Notch binding protiens and localization of Notch to focal adhesions
Authors: Havel Shelby Lee , Gazdik Tana Ranae , Wood Travis Lee , LaFoya Bryce , Albig Allan 
Published: June 2021

The Notch signaling pathway begins at the cell surface when Notch receptors interact with Notch ligands on adjacent cells triggering an intramolecular cleavage that releases the NICD (or Notch Intracellular Domain fragment) that then enters the nucleus to drive transcription. In addition to this cannonical Notch mechanism, our lab previously demonstrated that Notch signaling is modified by integrin ligation, via a previously unknown non‐cannonical mechanism. In unpublished research performed by LaFoya et al, SRC kinase has now been determined to bind to, and phosphorylate a small peptide near the ankyrin domain of Notch 1, resulting in modification of Notch signaling. The goal of our project is to expand upon these results by discovering other proteins that may interact with this region of Notch. To accomplish this goal, we constructed a fusion protein of the Notch peptide and the BirA protein, and performed a proximity biotin ligation screen in 293T cells for interacting proteins. Biotinylated proteins were isolated by streptavidin magnetic beads, and identified by MS/MS. Amongst the many hits, we identified members of the SRC family of kinases and several proteins commonly associated with focal adhesions and/or tight junctions. Western blotting confirmed the presence of focal adhesion proteins α‐actinin 1 and α‐actinin 4, and tight junction protiens cingulin, and angiomotin, in the biotin precipitated fractions. Subsequent co‐immunoprecipitation experiments in HMEC endothelial cells positively determined an interaction between the NICD domain and α‐actinin1, but failed to confirm interactions with the other proteins. The functional significance of this Notch‐actinin interaction is currently unknown, although given that SRC and actinin both localize to focal adhesions, we speculate that Notch may localize to focal adhesions where it is phosphorylated by SRC kinases.Support or Funding InformationNIH Grant 2R15GM102852‐02This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
DNA binding properties of the transcription factor optix
Authors: Rodriguez Jessica Marie 
Published: June 2021

DNA binding proteins such as transcription factors are key determinants of cellular state and have been shown to control cell differentiation. Evaluating the specificity profile of DNA binding proteins is a nontrivial challenge that hinders the ability to decipher gene regulatory networks or engineer molecules that act on genomes. optix is a transcription factors that has been previously identified as a master regulator of red color patterns in the butterfly (Heliconius) developing wing. However the direct gene targets of optix and their effects in final wing coloration remain to be determined. Based on this, we want to determine the intrinsic DNA‐binding preferences of optix. We are currently working on cloning, over‐expressing and purifying full‐length optix and optix's DNA‐binding domain, the homeodomain (HD). We will determine the in vitro DNA‐binding specificity of optix protein of Heliconius erato using Cognate Site Identification by high‐throughput SELEX. Our results will enable us to bioinformatically predict the genomic targets of optix and to better understand its role in wing color patterning.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Expression and Characterization of the Receptor Tyrosine Kinase Tie2
Authors: Kennedy Madison Arza , Wu Yunjin , Grunseth Adam , Sohl Christal D. 
Published: June 2021

Nearly one‐third of human kinases are implicated in disease, and this enzyme class has proved to be an enormously successful drug target for combating many diseases. Kinases are key regulators in signaling pathways responsible for such processes as growth, proliferation, and angiogenesis. Tie2 is an endothelium‐specific receptor tyrosine kinase (RTK) responsible for angiogenesis and vasculature maintenance. Mutations in this enzyme can cause venous malformations for which no current therapies are available, and Tie2 has been proposed to be a target for anticancer therapies. However, the molecular mechanisms of Tie2 activity are not well understood. Here, we developed a robust purification strategy using heterologous expression in insect cells, optimizing expression and purification with a post‐infection incubation time of 72 hours and utilization of affinity chromatography. We have created autophosphorylation assays to establish catalytic rates of wild‐type Tie2 and mutants implicated in disease. We hypothesize that mutant Tie2 will have faster rates of autophosphorylation, likely through stabilization of the active conformation of the protein. These studies in kinase activity can improve our understanding of the catalytic features of wild‐type and mutant Tie2, and can inform the development of therapeutic targets for vascular disease.Support or Funding InformationNIHThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A synergistic combination of Huperzine A, Convolvulus pluricaulis and Celastrus paniculatus promote cognitive function and health
Authors: Ahmad Imtiaz , Swaroop Anand , Bagchi Debasis 
Published: June 2021

Approximately 56 billion dollars are spent as a result of traumatic brain injury and about 90,000 Americans suffer from long‐term disability. Uprising stress, busy lifestyle and unhealthy food habits are the major causes of traumatic brain injury. The majority of the treatment strategies are based on the improvement of cholinergic function in the brain and one of the emerging therapeutic target is to enhance the acetylcholine level in the brain. Standardized herbal extracts including Huperzia serrata, Convolvulus pluricaulis (Shankhapushpi; SP) and Celastrus paniculatus (Jyotismati; JY) are long been known to attenuate brain function and well‐being. Research studies have demonstrated the versatile roles of these novel phytochemicals as natural acetylcholinesterase inhibitors and neuroprotectants, and exhibited their efficacy in the management of neurological or memory impairment, dementia, Parkinson's and Alzheimer's disease. In the recent past, we developed a novel, standardized Huperzia serrata extract, CogniUp™ enriched with 1% Huperzine A, which demonstrated broad spectrum safety and neuroprotective abilities. In this study, we developed a unique combination of CogniUp, SP and JY (MZ001), which can effectively and synergistically inhibit acetylcholinesterase (AChE) inhibition. Based on our initial assessment, concentration‐dependent AChE inhibition kinetics was assessed individually using 0, 3.0, 6.0, 9.0, 12.0 and 24.0 mg SP/ml, while AChE inhibition kinetics was assessed individually using 0, 0.5, 1.0, 2.0, 4.0 and 8.0 mg JY/ml. Simultaneously, concentration‐dependent AChE inhibition kinetics was assessed individually using 0, 0.0625, 0.125, 0.25 and 0.5 μg CogniUp/ml. Several combinations were assessed and ultimately, we found that a combination of SP, JY and CogniUp (12 mg/ml, 4 mg/ml and 0.125 μg/ml) provided the most efficacious and synergistic AChE inhibition. No significant adverse events were observed. Further studies are in progress to establish the therapeutic efficacy of our unique proprietary combination‐MZ001 containing SP, JY and CogniUp.Support or Funding InformationThis project was funded by Cepham Inc., Piscataway, NJThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A luminal kinase regulates sarcoplasmic reticulum calcium cycling and heart disease
Authors: Pollak Adam , Wiley Sandra , Dixon Jack 
Published: June 2021

The mechanisms and machinery regulating calcium cycling through the sarcoplasmic reticulum (SR) remain incompletely understood, and further insight is critical for discerning normal and diseased heart function. An overlooked component is luminal SR protein phosphorylation. Our lab recently identified a protein kinase in the secretory pathway lumen, Fam20C, which phosphorylates Ser within Ser‐x‐Glu/pSer motifs. Fam20C is responsible for generating the majority of the secretory phosphoproteome. However, Fam20C's role in cardiac function is currently undefined.Here we show that cardiac histidine‐rich calcium binding‐protein (HRC) is a Fam20C substrate. Importantly, a common HRC Ser96Ala genetic mutation causes deadly arrhythmias in patients. The functional significance of this mutation is to block Fam20C phosphorylation at Ser96. Furthermore, several other calcium regulating luminal SR proteins are Fam20C substrates, suggesting that Fam20C plays a broad and impactful role in SR function.We developed a cardiac‐specific Fam20C mouse knockout (KO) model. Using transverse aortic constriction (TAC), we show that Fam20C protects against heart failure. Fam20C KO mice show altered levels of SR calcium handling proteins during TAC, suggesting that calcium cycling through the SR is perturbed. Furthermore, we demonstrate that contractility is altered in Fam20C KO mice.In summary, we introduce Fam20C as a novel yet critical regulator of SR calcium cycling and cardiac biology. The phosphorylation of SR luminal cardiac proteins by Fam20C plays an important role in SR calcium dynamics and heart disease.Support or Funding InformationNHLBI RUTH L. KIRSCHSTEIN NRSA F32 HL136122‐01This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Assessment of Visual Literacy Skill Gains in Graduate Biochemistry
Authors: Procko Kristen , Dean Diane M. 
Published: June 2021

Biochemistry II, a culminating course of the biochemistry master's degree program at the University of Saint Joseph, emphasizes protein structure, folding, catalysis, and mechanism. Accordingly, the course has a significant molecular modeling component. Students are taught to render and manipulate proteins in PyMOL through tutorials that span the basics of the program, measurement and evaluation of potential molecular interactions, assessment of mutations, and prediction of protein function based on structural features. To evaluate students' progress in their ability to interpret visual representations of macromolecules, we designed a study using several of the overarching visual literacy themes outlined by the BioMolViz group (BioMolViz.org). At the beginning of the course, students complete a pretest in which they view macromolecule structures and answer questions that encompass several of the overarching visual literacy themes to evaluate their baseline biomolecular visualization skills. At the end of the semester, students complete an exit evaluation, which contains unique questions that mirror the first assignment. Students demonstrated improvement in each of the visual literacy themes evaluated, with the most significant gains in the structure‐function relationship and structural model skepticism areas. Here, we present the results of the assessment from three unique sections of the course, along with an overview of the tutorials and assignments being used to build visual literacy skills. Likewise, we will discuss revised course materials addressing areas in which students did not initially achieve the expected gains.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Kaposi's Sarcoma‐Associated Herpesvirus (KSHV) role in preventing apoptosis: The vBCL‐2 and human BIK association
Authors: Yeager Chyenne , Canfield Jennifer 
Published: June 2021

Kaposi's Sarcoma‐Associated Herpesvirus (KSHV) is a harmful oncogenic virus that leads to skin cancer commonly found in immunocompromised individuals. KSHV evades host defense systems by targeting signals transduction pathways to suppress autophagy and apoptosis. The BCL‐2 family of proteins controls commitment to apoptosis by regulating mitochondrial outer membrane permeabilization (MOMP). Research has been conducted examining the specific mechanisms and localization of different proteins in the BCL‐2 family such as BAK and BAX, but other proteins like BIK ( BCL‐2 interacting killer) have been far less studied. BIK is a pro‐apoptotic tumor suppressor that is critical n apoptosis induced by cytokines, toxins and viral invasion. Anti‐apoptotic Bcl‐2 family members competitively bind pro‐apoptotic proteins, like BIK, and suppress apoptosis. Although BIK is a known therapeutic target, little is known about the association of BIK with the KSHV BCL‐2 homology, vBCL‐2. In order to learn more about this association, we overexpressed vBCL‐2 in HEK293 cells and carried out subcellular fractionation to localize the protein. We analyzed the physical interaction human BIK and vBCL‐2 using immunoprecipitation under different cellular conditions. We examined the apoptosis mechanism using isolated mitochondria to measure the induction of cytochrome c release in the presence another BCL‐2 family member, BIM, from cells expressing MCL‐1 and MCL‐1 andvBCL‐2. By titrating in BIM and BIK, we can analyze the BIK/vBCL‐2 interactions. Results of these experiments will help elucidate the unique relationship between vBCL‐2 and BIK in the context of mitochondria mediated apoptosis. Definitively elucidating the mechanism, localization, and protein interactions in apoptotic and non‐apoptotic cell death in the presence and absence of vBCL‐2 will guide future therapeutic approaches against KSHV and possibly elucidate other non‐resistant mechanisms to kill cancer cells.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Apoptotic Effect of Methylglyoxal in Pancreatic RINm5F Cells; Reversal by INGAP‐P
Authors: Gonzales Kerald , Su Jing , Mungre Sue 
Published: June 2021

Diabetes mellitus is a metabolic disease characterized by high level of blood sugar. In a healthy person, the hormone insulin from β‐cells of pancreatic islets regulates blood sugar level. In diabetic patients, there is a continuous loss of pancreatic β‐cells mass, dysfunction of β‐cells that result in deficiency of insulin. Methylglyoxal (MG), a normal by product of glycolysis, is in high in a diabetic person. MG causes oxidative stress to β‐cells leading to cell death. A linear pentadecapeptide segment of Islet Neogenesis Associated Protein ‐ Peptide (INGAP‐P), Ac‐IGLHDPSHGTLPNGS‐NH2, has been shown to induce proliferation of β‐cells through transdifferentiation of matured ductal cells. The goal of this research was to determine whether INGAP‐P protects RINm5F cells, an islet β‐cell line, from the detrimental effects of MG. Cells were treated with different concentrations of MG in the absence or presence of INGAP‐P. Cell viability of RINm5F cells after treatment was measured by MTT, [3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide], assay at 570 nm. MG at 40μM caused 53% cell death in 24 hours. When treated with INGAP‐P at 1μM for 24 hours cell viability increased by 20%. When cells were treated with MG and INGAP‐P together, RINm5F cells showed no cell death suggesting that INGAP‐P protects the cells from MG. We are currently determining whether the cell death is due to apoptosis and how INGAP‐P protects RINm5F cells from the effect of MG. We will study the expression of Bax and Bcl2 in RINm5F cells treated with MG and INGAP‐P. The findings from this research will provide insights into the mode of action of INGAP‐P and guide the design of new synthetic drug molecules for diabetes that act in a similar mechanism.Support or Funding InformationUS Department of Education Title III GrantThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Exploration of Novel Markers of Posterior Capsular Opacification
Authors: Jackson Erin , Shihan Mahbubul , Wang Yan , Duncan Melinda 
Published: June 2021

Cataract, the leading cause of blindness worldwide, is defined as the clouding of the ocular lens. While cataract surgery is commonly performed by removing the lens and replacing it with an intraocular lens (IOL), it only temporarily cures cataract and actually causes a fibrotic condition called posterior capsular opacification (PCO). PCO, or secondary cataract, results from the migration and proliferation of lens capsule cells remaining post cataract surgery (PCS). While laser treatments can cure PCO, this incurs risks to vision. Thus, a comprehensive understanding of the signaling pathways leading to lens epithelial cell (LEC) fibrosis must be reached in order to prevent PCO. Previous research discovered that over‐activation of transforming growth factor beta (TGF‐β) signaling leads to increased fibrosis. The bone morphogenetic protein (BMP) signaling pathway is important for normal cell behavior and, in other cell types, controls fibrosis by inhibiting TGF‐β signaling. This work explores possible relationships between BMP and TGF‐β signaling during PCO pathogenesis. RNA sequencing and immunostaining revealed that many fibrotic markers are upregulated by 48 hours PCS including alpha smooth muscle actin (αSMA), collagen I, and lysyl oxidase, an extracellular matrix (ECM) protein that plays a role in the cross‐linking of collagen‐1. These upregulations occur concurrently with increases in phosphorylated SMAD3 (pSMAD3), a marker of the TGF‐β signaling pathway which is known to mediate LEC fibrosis. Additionally, a decrease in pSMAD1/5/8, major effectors of BMP signaling, was observed at the same time point consistent with the known requirement for BMP signaling in normal lens epithelial cell biology. Notably, the mRNA for Gremlin, a BMP antagonist, upregulates several hundred‐fold in LECs by 48 hours PCS, while Gremlin protein levels upregulate in lens cells sharply within six hours PCS and reach high levels at 48 hours PCS, providing a possible explanation for the attenuation of pSMAD1/5/8 observed. These data suggest that the upregulation of Gremlin PCS may be a critical factor in the downregulation of BMP signaling and the upregulation of TGF‐β signaling necessary for the fibrotic response leading to PCO.Support or Funding InformationThe National Eye Institute grant EY015279 as well as the University of Delaware Summer Scholars program funded this work.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification and characterization of protein changes in the Drosophila brain upon Wolbachia infection
Authors: Lucas Amber , Adams Haili , Winegardner Nicole , Sullivan William , Minden Jonathan S. 
Published: June 2021

Many bacterial hosts in recent years have been shown to have a profound effect on host behavior, and the endosymbiotic alphaproteobacteria Wolbachia is no exception. Present in at least 40% of the insect population, the maternally transmitted Wolbachia modulates behaviors such as male aggression, mate selection, and migration. Wolbachia have also been shown to change host response to olfactory cues, influence life span, and prevent infection by certain pathogens. This last feature of Wolbachia, specifically the prevention of infection by viral pathogens such as Dengue, Zika, and Chikungunya in mosquitos, has become of recent interest as a potential mechanism to suppress transmission of these disease‐causing viruses to humans.To tease apart the mechanisms by which Wolbachia influences host behavior and viral suppression, Drosophila, a genetically tractable model organism that is commonly used to delve into the molecular and cell biology of the symbiont behavioral, was used. A proteomic screen of changes in the Drosophila brain upon Wolbachia infection was performed using 2D Differential Gel Electrophoresis and LC‐MS. Protein abundance changes and post translational modification changes were observed and compared in both Drosophila simulans and Drosophila melanogaster as well as between male and female populations. Post translational modification changes were observed in multiple metabolic proteins and proteins responsible for neurotransmitter synthesis. This study describes the protein and post translational changes identified after Wolbachia infection and how they might contribute to behavioral changes in the host.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.