bluuebunny/crossref_metadata_2025_split huggingface dataset:
Crosstalk between mTOR and Estrogen Signaling
Authors: Holz Marina 
Published: April 2020

Binding of estrogen to ERα triggers the regulation of the target genes (genomic pathway). Additionally, a cytoplasmic fraction of estrogen‐bound ERα activates oncogenic signaling pathways such as PI3K/AKT/mTOR (nongenomic pathway). The upregulation of the estrogenic and the PI3K/AKT/mTOR signaling pathways is frequently associated with a poor outcome in breast cancer. To better characterize the connection between these two pathways, we performed a phosphoproteome analysis of estrogen and mTORC1 signaling. We identified two inhibitory proteins, DEPTOR end eIF3f, as estrogen‐regulated mTORC1 targets. DEPTOR binds to mTOR and inhibits its kinase activity. Although estrogen enhances the phosphorylation of DEPTOR by mTORC1, DEPTOR levels increase in estrogen‐stimulated cells. We demonstrated that DEPTOR accumulation is the result of estrogen‐ERα‐mediated transcriptional upregulation of DEPTOR expression. Consequently, the elevated levels of DEPTOR partially counterbalance the estrogen‐induced activation of mTORC1/2. We also found that the expression of the translation pre‐initiation inhibitory protein eIF3f is tightly controlled by ERα at the transcriptional and translational levels. Specifically, estrogen‐bound ERα represses transcription of the EIF3F gene, while promoting eIF3f mRNA translation. To regulate translation, estrogen activates the mTORC1 pathway. These results underscore the critical role of estrogen‐ERα as a modulator of the PI3K/AKT/mTOR signaling pathway in ER‐positive breast cancer cells.Support or Funding InformationNIH R35GM128675
Anti‐glycation Effect and Advanced Glycation End‐products Cross‐link Breaking Ability of Sclerocarya birrea Stembark Extracts
Authors: Adeniran Oluwaseyefunmi Iyabo , Mogale Motetelo Alfred 
Published: April 2020

Advanced glycation end‐products (AGEs) are implicated in the pathogenesis of late microvascular complications of diabetes mellitus. Currently, there is no clinically approved anti‐glycation agent with minimal side effects. The objective of the study was to investigate the anti‐glycation effect and AGE‐cross‐link breaking ability of Sclerocarya birrea (S. birrea) stembark extracts. Bovine serum albumin (BSA) was incubated with fructose in the presence of different stembark extracts of S. birrea at 37°C for 40 days. Fluorescent AGEs (FAGEs), total immunogenic AGEs (TIAGEs) and carboxymethyl lysine (CML) formed were measured using spectrofluorometry and enzyme‐linked immunosorbent assay (ELISA). The ability of S. birrea stembark extracts to break BSA‐AGE‐collagen cross‐links was also investigated by means of ELISA.Ethyl acetate, methanol and water extracts significantly inhibited the formation of FAGEs than the standard inhibitor aminoguanidine. Hexane extract demonstrated significantly higher anti‐glycation activity than aminoguanidine against TIAGEs. The water extract exhibited higher inhibitory activity than aminoguanidine against the formation of CML. With regard to the ability to break down AGE‐cross‐links, S. birrea stembark methanol and water extracts demonstrated higher ability to break AGE‐cross‐links than the hexane and ethyl acetate extracts. In conclusion, crude S. birrea stembark extracts have the ability to inhibit the formation of AGEs and to breakdown AGEs‐protein cross‐links. The results of this study may lead to the isolation of bioactive phytochemicals from S. birrea stembark extracts that may be used for the prevention of vascular complications of diabetes.Support or Funding Information1. National Research Foundation (NRF), South Africa. 2. Sefako Makgatho Health Sciences University (SMU), South Africa
Investigating the impact of cell maturity on transplantation success for CNS injury
Authors: Payne Samantha L , Tuladhar Anup , Obermeyer Jaclyn , Varga Balazs V , Morshead Cindi M , Nagy Andras , Shoichet Molly S 
Published: April 2020

Globally, a total of 82 million people suffer from injuries to the central nervous system (CNS) and to date there is no cure. However, while humans and other mammals have a limited capacity for functional tissue repair, various other species exhibit near‐perfect CNS regeneration following injury. Understanding the factors that regulate the reparative response can therefore provide important insight into strategies for promoting regeneration in mammals. An emerging approach involves the localized delivery of exogenous neural stem cells (NSCs). Although preclinical studies have shown that NSCs can survive once transplanted, whether they differentiate into neurons and functionally integrate into the host tissue remains unclear. In addition, there are questions as to what stage of NSC maturity will provide the most therapeutic benefit. To address these questions, we investigated cell survival, differentiation, and long‐term sensorimotor recovery in a rat model of stroke following the delivery of human NSCs at three different stages of maturity: 1) undifferentiated NSCs (early stage), 2) neuronal progenitors (mid stage), and 3) mature neurons (late stage). To facilitate cell survival and integration into host tissue, cultured cells were encapsulated into a biocompatible, bioresorbable hyaluronan/methylcellulose hydrogel prior to delivery. Our results demonstrated that transplantation of early stage cells results in the greatest tissue and functional repair, relative to transplantation of more mature neurons. Surprisingly, we found that the transplantation process itself resulted in acute cell death in the late stage cells. This negatively impacted host tissue and negated any potential positive effects associated with cell maturity and the hydrogel vehicle, which itself showed some functional and tissue benefit. Whereas each transplant cell group (early, mid and late) started the experiment with a distinctive maturity profile that was maintained after 7 days post‐transplantation, by 50 days the pattern of neural marker expression between groups was nearly identical, highlighting the influence of the microenvironment on the terminal cell phenotype. Overall, we demonstrate that the phenotype of the cells before and after transplantation had an enormous impact on their survival and the consequent tissue and behavioral response, emphasizing the importance of characterizing injected cells in CNS regenerative strategies.Support or Funding InformationWe are grateful for funding from: the Canada First Research Excellence Fund to Medicine by Design at the University of Toronto (to MSS, CMM, AN), the Canadian Institutes of Health Research (Foundation grant to MSS, Operating grant to CMM) and the Natural Sciences and Engineering Research Council (CGS‐D to SLP).
Multifaceted Roles of Primary Cilia in Polycystic Kidney Disease and Obesity
Authors: Tran Pamela V. 
Published: April 2020

The primary cilium is a small, antenna‐like sensory organelle that protrudes from most mammalian cells and mediates signaling pathways. Cilia are built by intraflagellar transport (IFT) protein complexes B and A, which have separate roles. Dysfunction of IFT and cilia results in ciliopathies, disease syndromes which commonly cause renal cystic disease and can also lead to obesity.Intriguingly, in mice, deletion of Ift‐B genes, which causes loss of primary cilia, can markedly attenuate Autosomal Dominant Polycystic Kidney Disease (ADPKD), the most common renal cystic disease and a leading cause of renal failure. These data indicate that targeting primary cilia genes can have immense therapeutic potential in ADPKD through mechanisms that remain elusive. To help elucidate these mechanisms, we have ablated an Ift‐A gene in ADPKD mouse models. IFT‐A dysfunction partially rescues the ADPKD elongated renal ciliary phenotype and attenuates the disease in a renal tubular‐ and maturation‐dependent manner.In adult mice, Ift gene deletion causes hyperphagia‐induced obesity. We have found that prior to an obese phenotype, IFT‐A dysfunction misregulates expression of appetite‐controlling neuropeptides in the hypothalamus. Additionally, in a pre‐obese state and through a non‐cell autonomous mechanism, hyperphagia induces hepatic insulin resistance, which drives systemic insulin resistance. These data uncouple obesity and insulin resistance, and indicate that hyperphagia is a major driver of both phenotypes. Thus, we are examining methods to counter hyperphagia. In collaboration with Zafgen Inc., we have found that pharmacological inhibition of MetAP2, which cleaves the N‐terminal methionine of nascent proteins, reduces food intake and body weight, and substantially improves metabolic parameters in an Ift‐A mutant mouse. To our knowledge, this is the first demonstration of rescue of an obese phenotype in a ciliopathy mouse model.Support or Funding InformationThis research is supported by NIH/NIDDK R01DK103033
Retinoid Exposure Decreases Foxp3 Expression in Cutaneous T‐cell Lymphoma
Authors: Stahly Samuel L. , Powers Andrea M. , Cline Reid , Griffin Lauren A. , Sarcar Bryce , Siddiqui Nabila A. , Johnson Lucas R. , Danekas Alexis N. , Martin Sykes , Geis Abby L. , Bridges Lance C. 
Published: April 2020

Retinoids, natural and synthetic vitamin A derivatives, activate members of the RAR and RXR subfamilies of transcription factors to promote cellular differentiation. Exploiting retinoids to induce differentiation embodies a milestone in cancer therapeutics. The success is championed by remarkable outcomes achieved upon the advent of retinoid regimens with acute promyelocytic leukemia (APL). While retinoids have been used for over 3 decades to treat cutaneous T‐cell lymphoma (CTCL), it remains unknown if retinoid‐dependent changes in differentiation accounts for the clinical benefit in CTCL as occurs with APL. Delineating the mechanism of action of retinoid‐based treatment in CTCL merits addressing considering that a lack of specificity contributes to metabolic sequelae that require ceasing treatment. The heterogeneity of the CTCL etiology further frustrates efforts as the exact classification of the T‐cell nature of CTCL is varied and largely debated. We first identify that CTCL lines express the hallmark Treg marker Foxp3, whereas lymphoma lines not of CTCL origin lack Foxp3 expression. We found that various synthetic and natural retinoids, including Bexarotene, an RXR agonist and the only FDA‐approved retinoid for CTCL treatment, induce a dose‐dependent decrease in Foxp3 expression in six different human CTCL cell lines. Changes in Foxp3 expression occurred at lower retinoid doses and shorter exposure times than are required to induce apoptosis. In addition, RAR and RXR receptors function synergistically to reduce Foxp3 expression at very low doses and abbreviated exposure times. Indeed, the decrement in Foxp3 expression is abrogated upon retinoid removal, demonstrating that diminished Foxp3 protein expression is not an artifact of cell death. Prior work by our group has shown that the RAR‐alpha receptor subtype prompts gut‐tropic adhesion and chemotactic changes with CTCL. We now show that the RAR‐alpha receptor subtype, assessed using selective receptor agonists and antagonists, transduces the observed changes in Foxp3 expression. This work establishes that the differentiation marker of Tregs is routinely expressed by CTCL cell lines, and that therapeutic retinoids decrease the abundance of this protein marker. It remains to be determined if the retinoid‐induced differentiation of CTCL forces a phenotype that is incompatible with cutaneous localization and predisposes the cells to apoptosis. Alternatively, retinoid‐forced differentiation of cells with a terminally differentiated phenotype, like CTCL, may trigger apoptosis. Furthermore, future work will determine if this newly identified consequence of retinoid exposure to alter differentiation markers in CTCL has clinical ramifications reminiscent of APL.Support or Funding InformationFunding provided by the BREA program at the Arkansas Colleges of Health Education
Deregulation of neuro‐developmental genes and primary cilium cytoskeleton anomalies in iPSc‐derived retinal sheets from human syndromic ciliopathies
Authors: Barabino Andrea , Flamier Anthony , Hanna Roy , Héon Elise , Freedman Benjamin , Bernier Gilbert 
Published: April 2020

Modeling human developmental and degenerative diseases has been always logistically challenging in particular in the case of rare diseases where no exhaustive animal models are available. Generation of sustainable human disease models that allow in‐depth analysis of the molecular mechanism is one of the big challenges nowadays. Stem cell technology holds great potential in disease modeling and represents a new powerful tool for generating scalable and animal‐free models that can more accurately illustrate clinical phenotypes of complex human diseases. Ciliopathies are a group of heterogeneous genetic diseases affecting proteins involved in primary cilium structure and function. Syndromic ciliopathies have a broad spectrum of symptoms ranging from retinal degeneration to skeletal and neuro‐development anomalies, including polydactylism and mental retardation. Herein we describe the generation and molecular characterization of iPSC‐derived photoreceptor sheets from patients affected by ciliopathies. Photoreceptor sheets are characterized by a polarized, multi‐layered tissue expressing outer segment, connecting cilium, and nuclear photoreceptor markers. Ciliopathic photoreceptors displayed significant common alterations in the expression of hundreds of developmental genes. Moreover, they showed several anomalies in the formation and maintenance of cilia, the positioning of the mother centriole and the activation of a stress response to misfolded proteins. Furthermore, we observed genomic instabilities and accumulation of DNA damage in the photoreceptors progenitors of one of the patients. This study reveals how combining cell reprogramming and organogenesis technologies with nextgeneration sequencing enable the elucidation of molecular and cellular mechanisms involved in human ciliopathies. The same approach, combining photoreceptor sheet differentiation and wide genome expression profile could be applied to model many genetic, developmental and degenerative diseases affecting photoreceptors. These patient‐derived retinal sheets may be useful for elucidating the molecular mechanisms underlining these diseases, for drug screening of compounds with potential therapeutic effect and predicting drugs side effects.Support or Funding InformationStem Cell Network (SCN), Fondation de l’Hôpital Maisonneuve‐Rosemont (FHMR), Réseau de recherche en santé de la vision (RRSV), Université de Montréal (UdeM)
Developing Scientific Writing Skills in Upper Level Biochemistry Students through Extensive Practice and Feedback
Authors: Ponce Maria Laura Sosa , Moorhead Greg B.G. 
Published: April 2020

Effective communication is one of the most marketable and transferable skills a graduate can have. Unfortunately, science programs rarely develop effective writing skills due to the time‐consuming nature of evaluating these skills. Here, we try to adapt tools from specifications grading to simplify marking and maximize student success in a third‐year biochemistry lab techniques course. We provided feedback to students on whether or not they were writing to the expected level on short lab reports so that they could implement it in a cumulative lab report. Students struggled to accept the all or none nature of specifications grading and did better with a writing workshop and one‐on‐one feedback. Overall, writing improved the most in sections where students received the most practice. We observed moderate success in improving writing skills in class size of 35, which is larger than most previous exercises of this nature.Support or Funding InformationThank you to Kyle McDade and Ryan Toth for their help in grading.
Signaling Diversity Enabled by Rap1 and cAMP/PKA‐Regulated Plasma Membrane ERK with Distinct Temporal Dynamics
Authors: Keyes Jeremiah , Ganesan Ambhighainath , Molinar-Inglis Olivia , Hamidzadeh Archer , Ling Megan , Trejo JoAnn , Levchenko Andre , Zhang Jin 
Published: April 2020

A variety of different signals induce specific responses through a common, ERK‐dependent kinase cascade. It has been suggested that signaling specificity can be achieved through precise temporal regulation of ERK activity. Given the wide distrubution of ERK susbtrates across different subcellular compartments, it is important to understand how ERK activity is temporally regulated at specific subcellular locations. To address this question, we expanded the toolbox of FRET‐based ERK biosensors by creating a series of improved biosensors targeted to various subcellular regions via sequence specific motifs to measure spatiotemporal changes in ERK enzymatic activity. Using these sensors, we showed that EGF induces sustained ERK activity near the plasma membrane in sharp contrast to the transient activity observed in the cytopolasm and nucleus. Furthermore, EGF‐induced plasma membrane ERK activity is regulated by Rap1, a noncanonical activator, and the cAMP/PKA signaling axis. We report that plasma membrane ERK controls cell morphology and EGF‐induced membrane protrusion dynamics. Our work strongly supports that spatial and temporal regulation of ERK activity is integrated to control signaling specificity from a single extracellular signal to multiple cellular processes.Support or Funding InformationThis work is supported by NIH R35 CA197622, R01 DK073368 (to J.Z). J.K. and O.M.I are supported by a post‐doctoral fellowship from NIH/NIGMS (K12GM068524). The development of Rap1 sensors at CISMM was supported by 5P41‐EB002025.
Chemical Biology Tools to Elucidate the Solution Conformation and Cellular Interactions of Cyclic Adenosine 5′‐Diphosphate Ribose
Authors: Uwakwe Chiemeka David , Grega Sarah , Serrano Inis , Graham Steven 
Published: April 2020

The movement of calcium ions (Ca2+) is intimately involved in regulating many physiological processes, and so there is great interest in the structure of molecules, especially second messengers, that regulate this flow. One such messenger is cyclic adenosine 5′‐diphosphate ribose (cADPR), which is suspected to play a role in Ca2+ release in cardiomyocytes. The speculated mechanism of action involves the binding of cADPR to FKBP12.6, releasing FKBP12.6 from RyR2 channels, through which Ca2+ enters the sarcoplasm from the sarcoplasmic reticulum during ventricular systole. Guided by the principle that structure determines function, we aim to advance the investigation of regulatory cADPR interactions by synthesizing and studying cADPR and its analogs using the combined tools of liquid chromatography‐mass spectrometry (LCMS) and nuclear magnetic resonance (NMR) spectroscopy. The endogenous cyclization of nicotinamide adenine dinucleotide (NAD+) to cADPR by CD38 is mimicked in vitro using ADP‐ribosylcyclase (ADP‐RC, sourced from Aplysia californica). The low substrate specificity of ADP‐RC conveniently permits the cyclization of various cADPR analogs. Kinetic assays were prompted by an observed feedback inhibition of ADP‐RC by nicotinamide (IC50 ~ 1mM), and performed using LCMS. Following synthesis and purification, NMR experimental data provided insight into favored structural conformations of cADPR (Saatori et al., 2018) Photoaffinity labeling (PAL) was aimed at identifying the cognate binding partners of cADPR, some of which have been previously described by Walseth et al. (1993). Conformational analyses of cADPR analogs revealed structural trends with correlations to the biological activity of the analogs.Support or Funding InformationDepartment of Chemistry, Saint John’s University
Expression and Purification of Human Circadian Protein hRORγ for Structural and Functional Studies
Authors: Van Sanchez Sebastian , Madej Alexander , Moreno Brenda , Xian Yuejiao , Yoo Seung-Hee , Chen Zheng , Xiao Chuan 
Published: April 2020

The circadian rhythm is an internal biological clock that runs in many living organisms. It regulates the sleep wake cycle among other physiological functions. Long term disruption of this internal clock can cause sleep disorders, metabolic diseases, and high risks of cancer. The circadian rhythm is controlled at molecular level by the transcriptional‐translational feedback loop (TTFL) that involve positive transcriptional activators CLOCK/BMAL1 and negative feedback suppressors PERs/CRYs. The Retinoic Acid Receptor‐Related Orphan Receptors (RORs) function as ligand‐dependent transcription factors which regulate BMAL1 production to improve circadian rhythm stability and robustness. It was shown that the naturally occurring flavonoid Nobiletin can bind to human ROR gamma (hRORγ) and increase its function through a yet unknown mechanism. Understanding the interaction between hRORγ and Nobiletin would provide insights in enhancing the circadian rhythm and ameliorating its related diseases. This project focuses on solving the three‐dimensional structure of hRORγ/Nobiletin complex to establish the mechanism of their interaction. The hRORγ gene was cloned and expressed using E. coli BL21 cells. Expression protocols were optimized to produce large amounts of hRORγ recombinant proteins. The soluble portion of expressed protein was purified using affinity chromatography and size exclusion chromatography, and the purified recombinant hRORγ is properly folded indicating from circular dichroism data. Once the required concentration is reached, the purified hRORγ will be co‐crystalized with Nobiletin. The crystals would be used for structural determination using X‐ray crystallography. The structural information will shed light on Nobiletin’s enhancement mechanism on circadian rhythm and provide guidance in related drug development.Support or Funding InformationResearch reported is supported by: National Research of General Medical Sciences of NIH under award number SC3GM109870. Researcher has used instruments sponsored by the BUILDing SCHOLARS program funded by the National Institute of General Medical Sciences of NIH under linked Award Numbers RL5GM118969, TL4GM118971, and UL1GM118970. Researcher is supported by Maximizing Access to Research Careers Grant # T34GM008048