bluuebunny/crossref_metadata_2025_split huggingface dataset:
CDK 7/9 inhibition amplifies mithramycin's suppression of Ewing sarcoma cell proliferation
Authors: Flores Guillermo , Everett Joel , Osgood Christy , Madaj Zachary , Grohar Patrick J 
Published: November 2023

BackgroundEwing sarcoma is driven by the fusion of the EWSR1 and FLI1 genes generated by a t(11:22) chromosomal translocation. Ewing sarcoma cells depend on the EWS‐FLI1 fusion product, a constitutively active oncogenic transcription factor, and become nonviable when EWS‐FLI1 driven transcription is inhibited. Mithramycin, a natural antineoplastic compound, has been identified as a potent EWS‐FLI1 inhibitor and is highly toxic to Ewing sarcoma cells. However, clinical trials of mithramycin in patients with Ewing sarcoma have been limited due to toxicity. To overcome this challenge, we sought to identify genes that sensitize Ewing sarcoma cells to mithramycin when inhibited. We identified RNA pol II inhibition as a potent sensitizer to mithramycin and phenocopied this with the dual cyclin dependent kinase (CDK) 7/9 inhibitors PHA‐767491 and SNS‐032. We show a marked suppression of cellular proliferation in response to mithramycin in combination with either CDK 7/9 inhibitor at concentrations significantly lower than mithramycin alone.MethodsWe utilized matrix drug screening with small molecule CDK inhibitors in combination with mithramycin. We measured cell viability using MTS assays and time lapse microscopy in Ewing sarcoma cells. Results were correlated with EWS‐FLI1 activity using western blot analyses and qPCR. Synergy between small molecule inhibitors and mithramycin was measured using Bliss independence.ResultsPHA‐767491 and SNS‐032 displayed strong synergy when combined with mithramycin. These combinations decreased the concentration of mithramycin necessary to block Ewing sarcoma cell proliferation, measured by multiple complementary assays. These results reflected effects on EWS‐FLI1 activity.ConclusionsWe have identified a synergistic relationship between the small molecules PHA‐767491 and SNS‐032 with mithramycin in Ewing sarcoma cells. Future work will include siRNA validation of the CDK 7/9 inhibition and mithramycin combination, analysis of genome wide expression changes, and characterization of the efficacy of a PHA‐767491 or SNS‐032 and mithramycin drug combination in vivo with the eventual goal of introducing it into the clinic.
The Role of Scleraxis in Neutrophil Activation
Authors: Awotunde Olanike , Nillas Andrea , Hammad Samar , Reaves Titus A. 
Published: November 2023

Scleraxis is a transcription factor in the basic helix loop helix (BHLH) family, and originally shown to regulate embryonic tissue into tendon and endothelium in blood vessels. More recently scleraxis has been linked to activation of fibroblasts and cardiac fibrosis. Organ fibrosis follows inflammation and is characterized by dysregulated and excessive production/release of extracellular matrix proteins—principally collagen. Within fibrotic organs there are a variety of immune cells. In particular, polymorphonuclear leukocytes (PMN) are components of the innate immune system and are first responder cells present in high numbers in organs undergoing fibrosis. PMN have two basic functions, they migrate to areas of inflammation/injury and destroy pathogenic organisms. Activated PMN release myeloperoxidase ((MPO), heme protease that is toxic to pathogens and is dependent on the leukocyte specific integrin CD11b) from their primary granules. We hypothesize that scleraxis is involved in the activation of PMN. Immunocytochemistry, flow cytometry, and Western blot analysis shows that PMN express scleraxis. Antibodies against scleraxis had minimal effects on cell attachment of PMN to fibrinogen, but induced substantial release of MPO. Taken together these results highlight scleraxis as a potential treatment protein for inflammatory conditions involving PMN.Support or Funding InformationNIH 4R25HL092611‐09 and National Science foundation (RII), 3T34GM008663‐20S1
CD2AP facilitates HCV production by targeting NS5A to lipid droplets and regulating lipid droplet biogenesis
Authors: Li Chaoyang 
Published: November 2023

Lipid droplets (LDs) play an important role in the life cycle of hepatitis C virus (HCV). In HCV infected cells, the surfaces of LDs are decorated with HCV non‐structure protein NS5A and structure protein, core. However, the targeting of these proteins to LDs and the biogenesis of LDs are complex and not completely understood. Here, we describe that CD2 associated protein (CD2AP), a scaffolding molecule that regulates the actin cytoskeleton, plays dual functions to affect HCV assembly. First, by interacting with HCV non‐structural protein NS5A, CD2AP transfers NS5A via an actin dependent manner to a cellular machinery which is then targeting to LDs via the microtubule dependent manner. The interaction between NS5A and CD2AP requires the SH3 domain of CD2AP and the domain I of NS5A. Second, CD2AP affects HCV assembly by modulating LDs biogenesis. Down regulation of CD2AP significantly reduces biogenesis of LDs, NS5A and core protein location on LDs, and thus virus assembly but not HCV genome replication, virus entry, and virus IRES dependent translation. We found that in hepatocytes CD2AP, cbl‐b E3 ligase and IRS1 form a protein complex and binding between CD2AP and cbl‐b enhances IRS1 ubiquitination. Knocking down CD2AP increases IRS1 total protein and phosphorylated IRS1 level, affects AMPK to HSL axis, and results in reduced LDs biogenesis. Thus, CD2AP affects HCV assembly via two distinct pathways. Because CD2AP regulates LD biogenesis, our results not only provide mechanisms to understand HCV assembly but also point out a potential approach to target insulin resistance.Support or Funding InformationMinistry of Science and Technology of China, Grant #: 2013CB911102
Intervertebral disc degeneration and low back pain: Pre‐clinical studies and clinical implications
Authors: Stone Laura S 
Published: November 2023

Chronic back pain is a major health problem throughout the world, affecting 10–15% of the global population. Current diagnostic and therapeutic approaches to chronic back pain are limited by our narrow understanding of the underlying biological mechanisms. While intervertebral disc degeneration has been linked to chronic back pain in humans, the relationship between disc pathology and pain symptoms remains a subject of debate. We are using mouse models of intervertebral disc degeneration to study this complex problem. Recent pre‐clinical findings on the relationship between progressive disc degeneration, pathological disc biochemistry and innervation and the emergence of behavioural signs of low back pain and radicular leg pain will be discussed. The therapeutic benefits of pharmacological and non‐pharmacological interventions (e.g. physical activity, diet) on these parameters will be presented and potential mechanisms underlying the low back pain and the beneficial effects of physical activity will be proposed.Support or Funding InformationSupported by the Canadian Institutes of Health Research
Patterning coronary artery development
Authors: Red‐Horse Kristy 
Published: November 2023

An important goal of developmental biology is to understand how millions of cells unite to produce a complex organ. For this process to be successful, the cells must take on proper fates at the appropriate times, in the correct proportions, and organize themselves so that the organ functions effectively. We are using the cardiovascular system as a model to understand the rules of these developmental events. Our laboratory is specifically focused on how cardiovascular progenitor cells behave in three dimensions and at the single cell level with inputs from physical forces. This allows us to study coronary development at high resolution to gain new information on how the cardiovascular system is assembled during embryogenesis and how developmental programs are reignited during disease.Support or Funding InformationThis research is supported by NHLBI of the National Institutes of Health under award number R01 HL128503.
Gross Anatomical Brain Region Approximation (GABRA): a new landmark‐based approach for estimating brain regions in dinosaurs and other archosaurs
Authors: Morhardt Ashley Caroline , Ridgely Ryan , Witmer Lawrence M 
Published: November 2023

Studying brain evolution in extinct taxa can be challenging due to a lack of close correspondence between the morphology of the endocranial cavity and that of the underlying brain. Cranial endocasts may be faithful brain proxies in certain groups (mammals, birds) due to relatively complete filling of the cavity with neural tissue in life. However, the brain does not fill the endocranial cavity in many adult non‐avian archosaurs, making their endocasts less reliable indicators of brain size and shape. As such, previous studies of relative brain size and evolution in archosaurs relied on untested assumptions about brain‐endocast fidelity. We propose a new approach known as Gross Anatomical Brain Region Approximation (GABRA), which involves importing a digital endocast, derived from CT scanning and 3D visualization software, into modeling software (Maya). In Maya, brain regions underlying the endocast are modeled as 3D polygons. Discernable osteological correlates for soft‐tissue structures (e.g., neurovascular canals, dural sinuses, fossae formed by the brain itself) present on the endocast serve as anatomical landmarks that inform the location and size of the modeled general brain regions (e.g., cerebral hemispheres, cerebellum, optic lobes, olfactory bulbs). Together, the anatomical landmarks form a set of explicit criteria used to assess endocasts and model brain regions. GABRA criteria and resulting brain models were validated in extant diapsids (lizards, snakes, alligators, birds) via literature review, gross dissection, CT scanning of iodine‐stained specimens, and MRI studies. Therefore, GABRA models produced for extinct archosaurs are credible. Credible GABRA models for extinct dinosaurs provided volumetric estimates for general brain regions, which, when summed, offer a whole‐brain estimate. Such data from several dinosaurs have permitted analyses using modern comparative methods of relative brain‐size evolution (e.g., encephalization quotient), with results indicating that brain sizes for many dinosaurs were previously underestimated. Additionally, analyses of GABRA data show a mix of concerted and mosaic patterns of brain evolution, wherein the pituitary and olfactory bulbs emerge as evolving independently from the rest of the brain. In sum, GABRA provides insight into how brains evolved across dinosaur lineages. Future studies will examine within‐lineage changes, offering greater insight into how and why dinosaur brains evolved.Support or Funding InformationFunding for this project was provided by the follow sources: the National Science Foundation, Jurassic Foundation, Sigma Xi Grant‐in‐Aid of Research, Ohio Center for Ecology and Evolutionary Studies, Ohio University Student Enhancement Award, Ohio University Original Work Grant.
A new Paradigm for Catalysis of Nucleotidyltransfer Reactions
Authors: Yang Wei , Gao Yang , Samara Nadine , Wu Jinjun 
Published: November 2023

Enzyme catalysis has been studied for over a century. How it actually occurs has not been visualized until recently. By combining in crystallo catalysis with X‐ray diffraction analysis of reaction intermediates, we have observed the processes of DNA synthesis and RNA at unprecendented atomic detail. Contrary to the canonical view that enzyme‐substrate complexes and transition states have identical atomic composition and catalysis occurs by a two‐metal‐ion mechanism, we have discovered that it is essential for a third divalent cation to be captured en route to product formation. Unlike the canonical two metal ions, which are coordinated by DNA polymerases and nucleases, this third metal ion is not coordinated by the enzyme at all. Its location between the a‐ and b‐phosphates of dNTP in the case of DNA synthesis suggests that the third metal ion may drive phosphoryltransfer from the leaving group opposite to the 3′‐OH nucleophile. Experimental data indicate that binding of the third metal ion may be the rate‐limiting step in DNA synthesis, and the free energy associated with metal‐ion binding can overcome the activation barrier to the DNA synthesis reaction.Support or Funding InformationIntramural Research of NIDDK, National Institutes of HealthIATAP, National Institutes of Health
What lurks beneath (the membrane): a mechanistic exploration of rhomboid proteolysis
Authors: Urban Sinisa 
Published: November 2023

At the turn of the millennium an extraordinary class of enzymes that cut proteins inside the cell membrane was discovered. Rhomboid proteases are the largest family of these ancient membrane‐immersed enzymes. Although deceptively simple, the act of hydrolyzing a single peptide bond inside the water‐excluding environment of the cell membrane has evolved to trigger cell signaling events during bacterial growth and animal embryogenesis, as well as mitochondrial quality control throughout the life of a cell. The membrane is a challenging environment for catalysis. We have been pioneering new quantitative methods to study rhomboid catalysis directly inside the membrane and combining them with modern X‐ray crystallography approaches. I will discuss an integrated but unexpected model for how rhomboid proteases function, and the promising new opportunities that it raises for combating devastating parasitic diseases that have plagued mankind for millennia, including malaria.Support or Funding InformationThis work was supported by NIH grants 2R01AI066025 and R01AI110925, the Howard Hughes Medical Institute, and the David & Lucile Packard Foundation.
Pre‐mRNA splicing, histone modification, and the coordinated control of gene expression
Authors: Johnson Tracy L. , Hossain Munshi Azad 
Published: November 2023

The emergence of whole genome and whole transcriptome sequencing has highlighted the need to explore new models to explain complex patterns of gene expression. For example, there has been a deepened appreciation for the role of diverse RNAs and their highly regulated processing in key cellular functions. Every step in the life cycle of an RNA—from its synthesis and processing to its export and translation—is subject to regulation. Recent studies have revealed that an added layer of regulation comes from the coupling and coordination of these reactions. RNA splicing, for example, occurs co‐transcriptionally, raising important questions about the interplay between transcription, chromatin, and the spliceosome as the cell regulates gene expression to adapt to a variety of environmental conditions.Despite its relatively streamlined genome, there are important examples of regulated splicing in Saccharomyces cerevisiae that provide mechanistic insights into the coordination of nuclear processes. We previously demonstrated that intron retention (a phenomenon that is common in higher eukaryotes) plays a crucial role in the yeast cell's ability to execute the proper gene expression program in response to nutrient deprivation (Hossain, M.A., Claggett, J. et al. Molecular Cell 2016). The glycolytic transcription factor Gcr1 (GlyColysis regulator 1) controls expression of over 75% of the genes in actively growing yeast, and GCR1 RNA contains a highly conserved intron, which allows the cell to generate multiple RNA isoforms whose levels and ratios change upon glucose depletion. One of these isoforms containing the retained intron is translated from a start site within the intron, raising important questions about regulation of GCR1 splicing, export of the intron‐containing GCR1 RNA, escape of the RNA from the cytoplasmic surveillance machinery, and the mechanism of its translation from different translation start sites. Additionally, synthesis of GCR1 is tightly regulated, including the regulated utilization of alternative transcription start sites that direct the synthesis of RNAs to make functional proteins. Hence, GCR1 regulation reveals intriguing RNA processing mechanisms. Here we show a role for chromatin modification in both the synthesis and processing of GCR1 RNA and define the features of GCR1 that influence its regulation. Based on these results, we have analyzed the yeast transcriptome under a variety of conditions to assess the pervasiveness of these mechanisms in yeast cells. These results reveal common themes that govern the coordinated regulation of gene expression processes.Support or Funding InformationThis work was funded by:Howard Hughes Medical Institute Professors Program grant # 52008140The National Science Foundation MCB‐1051921NIH GM‐085474
Orchestrating aging across a troubled soma
Authors: Dillin Andrew 
Published: November 2023

As an organism ages, its proteins face an increasing severity in the challenges they receive from extrinsic and intrinsic environmental perturbations [1, 2]. Chaperones become dysregulated, while the degradation machineries stop working properly. The protein accumulates damage and starts to misfold. At this point, the cell needs to mount a response to restore its homeostasis; however, the stress response machinery that it typically relies upon when faced with such challenges has lost its capacity to function [1, 2].This breakdown, however, does not lead to complete disorder. As the organism ages, it exhibits a degree of correlated, recognizable, and predictable changes to its physiology over time. These changes can occur synchronously across multiple tissues and organs. The phenotypic changes of aging occur in a type of concert, rather than in isolation, suggesting the residual participation of the endocrine system in the onset of age‐related phenotypes [3, 4]. The demise of the cell thus most often occurs within the context of the simultaneous demise of the whole organism.Our lab focuses on the questions of why an aging organism begins to lose control over the integrity of its proteome, and how this loss is communicated across its various tissues. We have taken the unique approach of breaking down a cell into its small and canonically‐autonomous parts – its suborganelles and subcompartments – such that we can take a larger step back to ask how those smaller portions can communicate both with each other and with the organism as a whole. Our approaches have required us to diversify the contexts in which we ask questions: we work on model systems ranging from stem cells and nematodes to mice. We have developed and applied techniques that allow us to manipulate signaling pathways or proteins within a single tissue, cell, or an organelle within a single cell so that we can observe how that small perturbation might reverberate and effect the physiology of the whole of the organism.In our research, we have found that the stress response pathways that ensure proteome maintenance are not restricted to cell autonomous events, but can be communicated across an organism. We have reported that this is found for the unfolded protein response of the mitochondria (UPRMT), and now find distinct signaling events for the UPR of the endoplasmic reticulum (UPRER) and the cytosolic heat shock response (HSR). Collectively, we have discovered at least three distinct signaling events that report upon proteome form and function across an organism. Our evidence strongly suggests that intracellular and even organelle‐specific stress can be communicated extracellularly via a regulated, endocrine‐like process to impinge on the aging process and health
Electron cryomicroscopy of rotary ATPases
Authors: Rubinstein John L. 
Published: November 2023

Ion‐translocating rotary ATPases serve either as adenosine triphosphate (ATP) synthases, using energy from a transmembrane ion motive force to create the cell's supply of ATP, or as transmembrane ion pumps that are powered by ATP hydrolysis. The members of this family of enzymes each contain two rotary motors: one that couples ion translocation to rotation and one that couples rotation to ATP synthesis or hydrolysis. Our group uses electron cryomicroscopy (cryo‐EM) of single protein particles to study the structures of rotary ATPases. We also work to develop new methods for cryo‐EM to facilitate these studies. Our recent studies have not only illuminated the structures of these fascinating molecular motors at unprecedented resolution, but have also started to uncover their dynamics through computational isolation of the different conformations of the enzymes that exist simultaneously in solution. This lecture will describe the latest structures we have determined for the mitochondrial ATP synthase and proton pumping V‐ATPase, and what we have learned about how these enzymes function and how they interact with molecules that affect their activities.Support or Funding InformationCanadian Institutes of Health Research, Canada Research Chairs
Innovations in Biotechnology Graduate Education
Authors: Jamison‐McClung Denneal S. 
Published: November 2023

Today's graduate students in biotechnology have a range of career options in the public and private sectors, including academia, industry, government and non‐profit organizations. It is not enough to be excellent researchers and university classroom teachers in today's job market; graduate students must develop a suite of transferrable professional skills that transcend STEM disciplines and academic norms.The trend toward interdisciplinary graduate education and public‐private research collaborations roughly 30 years prompted the formation of the UC Davis Biotechnology Program in 1986, and the subsequent launch of a formal doctoral degree program, the Designated Emphasis in Biotechnology (DEB) in 1997. Since that time the DEB has grown into the largest interdisciplinary degree program on campus, with a “steady‐state” of over 200 students from 29 different disciplines.Doctoral students see value in the DEB training model, which includes exposure to the “business of biotech”, including bioethics, intellectual property, regulatory affairs, project management and entrepreneurship. Through coursework, seminars, one‐on‐one career mentoring and industrial internships, DEB students gain a thorough understanding of non‐academic STEM career paths and an appreciation for the importance of interdisciplinary “team science” in developing new technologies.An overarching theme of DEB training is the development of strong communication and professional networking skills. DEB participants are presented with numerous opportunities to hone science communication skills for non‐specialist audiences (e.g. business plan competitions, K‐14 stem outreach, policy brief workshops, etc.), which is key for success in all STEM fields. This talk will elaborate on the mentoring tools and strategies we employ to help DEB doctoral students successfully transition from students to professional colleagues across a range of career paths.