bluuebunny/crossref_metadata_2025_split huggingface dataset:
Extracellular MIF and Wnt and eph/ephrin signaling are involved in WWOX‐regulated cell‐cell recognition and migration
Authors: Chang Nan‐Shan , Chou Pei‐Yi , Kuo Hsiang‐Ling , Sie Yong‐Da 
Published: June 2021

Loss of tumor suppressor WWOX enhances cancer cell metastasis. Here, we determined that WWOX‐deficient cells fail to recognize the parental wild type cells or any WWOX‐expressing cells of the same or different species. WWOX‐negative cells have loose cell‐cell contacts and migrate individually. WWOX‐positive cells group tightly and migrate collectively. Transiently overexpressed WWOX suppresses cancer cell migration. Knockdown of WWOX by small interfering RNA, or ectopic dominant‐negative WWOX, increases cell migration. WWOX/p53/TIAF1 triad is a known axis of tumor suppression. Ectopic expression of the protein triad suppresses cell migration, anchorage‐independent growth, and induces apoptosis. Mutant p53 at the Ser46 phosphorylation site causes functional loss of the triad. Among all the p53 isoforms, ectopic Δ133p53γ is most potent in inducing apoptosis due, in part, to its activation of the SMAD‐responsive promoter. Time‐lapse microscopy revealed that upon encountering, WWOX‐negative human breast MDA‐MB‐231 cells suppress the migration WWOX‐positive mouse L929 fibroblasts. Few L929 cells undergo apoptosis. Indeed, MDA‐MB‐231 accelerate their migration, contact L929 with protrusions, and then migrate in a retrograde manner. The accelerated migration of MDA‐MB‐231 is associated with rapid activation of the Wnt signaling‐mediated TCF/LEF‐responsive promoter in less than 2 hr. However, L929 exhibit the activation of CRE and NF‐kB promoters in 16 hr and later. Suppression of the Wnt signaling by XAV939 abolishes the retrograde migration of MDA‐MB‐231 and their physical merge with L929. Similarly, inhibition of cell surface ephrin A1, B1 and B2 and extracellular MIF with specific antibodies leads to anterograde migration of MDA‐MB‐231 and restoration of their merge with L929. Together, WWOX‐regulated cell‐to‐cell recognition is associated with cell surface ephrins, extracellular TGF‐β and MIF, and Wnt‐dependent TCF/LEF promoter activation, so as to to avoid unwanted surrounding microenvironment by retrograde migration.Support or Funding InformationResearch was supported by the Department of Defense USA (W81XWH‐08–1‐0682), the Ministry of Science and Education, Taiwan, ROC (NSC99‐2320‐B‐006‐012‐MY3, 102‐2320‐B‐006‐018‐, 102‐2320‐B‐006‐030‐, and 102‐3011‐P‐006‐005‐), and the National Health Research Institute, Taiwan, ROC (NHRI‐EX99–9704BI; NHRI‐EX107‐10734NI) to NS Chang.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of adenosine A1 receptor agonist induced‐hibernation on the arousal pathway in the Arctic Ground Squirrel
Authors: Jenkins Mackenzie , Frare Carla , Drew Kelly 
Published: June 2021

Hibernation is a seasonal phenomenon and in the arctic ground squirrel (AGS), the effect of seasonal changes is very evident. In late fall AGS decrease their metabolic demand followed by the entry into hibernation in winter. Previously in the lab it has been shown that N6‐cyclohexyladenosine (CHA), an A1 adenosine receptor agonist, induces hibernation in AGS in a seasonally dependent manner. In winter AGS treated with CHA experience a decrease in metabolic demand and temperature as seen during natural hibernation. However, in summer CHA triggers a brief and transient metabolic suppression that does not lead to hibernation onset.As adenosine is known to promote sleep, we decided to investigate if the sleep‐arousal pathways were affecting the seasonal difference in response to CHA. The tuberomammillary nucleus (TMN) is a nucleus of the ascending arousal pathway characterized by fast firing rates to promote wakefulness. Previously we showed a decrease in neuronal activation in the TMN in AGS treated with CHA in both summer and winter seasons.Here we test the hypothesis that seasonal response to CHA is associated with difference in activation in another nucleus of the ascending arousal pathway, the locus coeruleus (LC).AGS implanted with body temperature (Tb) transmitter, were treated with CHA or vehicle (0.5mg/kg, IP). AGS were perfused with 4% paraformaldehyde and brains were dissected and cryoprotected in 5%, 10%, 15%, 20%, 30% sucrose. We performed free‐floating immunohistochemistry on 40μm brain slices in four experimental groups: winter CHA, winter control, summer CHA and summer control. We sectioned brains from eight AGS per group for a total of 32 animals and identified active nuclei with cFos immunoreactivity (mouse anti‐cFos 1:20,000, Santa Cruz) and active tyrosine hydroxylase neurons (rabbit anti‐TH). Blinded analysis of cFos immunoreactive neurons was done using bright field microscopy and Metamorph software. Statistical analysis was performed on R.CHA produced a hibernation‐like response measured by a decrease in Tb in winter AGS but not in the summer. We did not observe any significant difference in neuronal activation in the LC, thus CHA is associated with a decreased neuronal activity in the TMN, resulting in the TMN being the major wakefulness center affected by CHA but not by seasons.Support or Funding InformationResearch was supported by NIH R03 NS081637, NSF IOS‐1258179, RL5GM118990 and Alaska INBRE P20GM103395. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The gamma‐aminobutyric acid (GABA)A receptor modulates the cytokine response to rhinovirus exposure in human monocytic cells
Authors: Hall David J , Hader Shelby N , Kondiles Bethany R 
Published: June 2021

Rhinoviral infections are a major cause of asthma exacerbations. Though productive rhinovirus infections occur predominantly in the bronchial epithelial cells of the upper airway, monocytic‐lineage cells, the most prominent immune cell in the lung, are important in establishing the inflammatory microenvironment observed during rhinoviral asthma exacerbations. Emerging evidence indicates that the gamma‐aminobutyric acid type A receptor (GABAAR) signaling system on leukocytes, airway epithelial cells and airway smooth muscle cells modulates the inflammation in the lung. We sought to determine if GABAAR signaling alters the release of inflammatory mediators from human monocyte‐derived macrophages (MDM) exposed to human rhinovirus (HRV). Results demonstrate that GABAARs on human MDMs are functional and stimulation with GABA or HRV caused GABAARs on the cell surface undergo protein kinase C (PKC)‐mediated endocytic removal and recycling, similar to that found in neurons. GABA treatment of MDMs during HRV exposure attenuated the production of HRV stimulated CCL2 and CXCL10 production by inhibiting the stress kinase June N‐terminal kinase (JNK) and increasing phosphorylation of extracellular regulated kinase (ERK). Interestingly, GABA stimulated the production of interferon‐b through the classical innate immune signaling molecules TANK binding kinase (TBK) and interferon regulated factor 3 (IRF3). Phosphorylation TBK occurred via the lysine deficient kinase (WNK) pathway after the influx of chloride ions when GABAARs are stimulated. Our findings suggest that GABA treatment of MDMs lead to a novel activation of the WNK‐TBK‐IRF3 axis implying the potential for GABA‐mediated treatment of HRV‐induced asthma exacerbations.Support or Funding InformationLawrence University, NIH grant R15 AI065505‐01A1 and NSF grant 0521112.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Drosophila CG1275 Expression, Antibody Production, and Western Blotting
Authors: Garcia Brandon Michael , Gorman Maureen J. , Ragan Emily J. 
Published: June 2021

Little is known about iron absorption in insects. One potential mechanism involves the action of a ferric reductase to reduce Fe(III) to Fe (II), which can enter the cell through a divalent metal transporter. CG1275 is a potential ferric reductase that is found in Drosophila melanogaster. By replicating, cloning, and expressing the N‐terminus of CG1275 we can develop a reagent to allow us to observe a protein knockdown after RNA interference (RNAi) experiments. We cloned a cDNA encoding the N‐terminal 107 amino acids of CG1275‐PA, the region before the first transmembrane helix, into pET‐32a. This was confirmed by restriction digestion. Expression of the N‐terminus of CG1275 along with a thioredoxin (Trx) tag and 6xHis tag from the vector was performed in Bl21 (DE3) E. coli. We then purified the protein using Nickel‐column affinity chromatography. The purified protein was separated by SDS‐PAGE and the protein bands of interest were cut out and sent for antibody production. The newly developed antibodies were then separated to remove antibodies that recognize the Trx tag. The antibodies to CG1275 were then used for western blots of S2 cell lysates to determine if they recognize CG1275 and are able to verify successful CG1275 knockdown after RNAi experiments.Support or Funding InformationNSF (IOS‐1656407)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigating a cytochrome b561 family member in insect cellular iron uptake
Authors: Holst Jessica , Arismendiz Omar Alexis Rodriguez , Ochoa Alma , Gorman Maureen J. , Ragan Emily J. 
Published: June 2021

Iron is an essential element to many functions of an animal's life, yet it is also toxic. Because iron is both essential and harmful, tight regulation of its concentration, location and ionization state is vital to animal livelihood. While we understand much about iron uptake in mammals, the mechanism for its transport is poorly understood in insects. We know iron is significant to many aspects of an insect's life, including: energy metabolism, detoxification of pesticides and as a nutrient for symbionts and commensals. We have identified a cytochrome b561 family member in Drosophila melanogaster, CG1275, that is similar to mammalian duodenal cytochrome b, a known ferric reductase involved in iron uptake. CG1275 is well expressed across all life stages of Drosophila, and we hypothesize that it may be involved in the uptake of iron to non‐intestinal insect cells. To test this hypothesis, we are performing RNA interference (RNAi) experiments to attempt knockdown in the expression of the gene coding for this protein. Previously we relied upon traditional polymerase chain reaction (PCR) to qualitatively observe whether knockdown had occurred. Now, by using real‐time PCR, we have quantified the CG1275 knockdown observed in our RNAi experiments. Next we will measure the iron concentration levels of our S2 cells to determine whether CG1275 plays a role in iron uptake to non‐intestinal insect cells.Support or Funding InformationNSF IOS‐1656407This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Quantitative Comparison of GM1 Gangliosidosis Afflicted Ovine Tissue using Thin‐Layer Chromatography, Mass Spectrometry and Nuclear Magnetic Resonance
Authors: Mata Cindy Venegas , Mitchell Jon C. 
Published: June 2021

GM1 ganglioside is a naturally occurring glycosphingolipid that cannot be synthesized in significant quantities; so it needs to be isolated as a natural product. Researchers at Glycoscience Research, Inc. (GRI – Toronto, SD) have developed a novel, verified, ovine source for producing GM1 ganglioside at levels needed for pharmaceutical applications. GM1 gangliosides have been shown to be an effective therapeutic against Huntington's disease as well as a potential promising lead against Parkinson's disease, Alzheimer's, and dementia (Holler, et al., 2016). Research presented here attempts to address the biochemical quantification of purified GM1 ganglioside isolated from afflicted/non‐afflicted lamb muscle to definitively show no significant differences in GM1 levels between the two meat sources. Chloroform:methanol extraction, anion exchange chromatography (AEC) and High Performance Thin‐Layer Chromatography (HPTLC) followed by Nuclear Magnetic Resonance (NMR) and MS/MS Spectrophotometry analysis compares similarities and differences in GM1 concentration levels between muscle tissue from lambs afflicted with the genetic disease of overproducing GM1 ganglioside and normal, wildtype lamb muscle.Support or Funding InformationThis material is based upon work supported by the National Science Foundation/EPSCoR Cooperative Agreement #IIA‐1355423 and by the State of South Dakota, SD‐GOED, NSF I‐Corps Team Program and Dr. Larry and Sue Holler, GlycoScience Research IncorporatedThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Synthesis and Development of peptidoglycan fragment microarray and probes to investigate innate immune signaling
Authors: Zhou Junhui , Lazor Klare , Grimes Catherine 
Published: June 2021

The innate immune system's interaction with bacteria plays a pivotal role in a variety of human diseases, such as Crohn's disease, asthma, and rheumatoid arthritis. In order to develop better therapies, small molecule probes derived from bacterial cell wall peptidoglycan (PG) are needed. Consequently, we are synthesizing new chemical probes based on PG to better understand their interactions with innate immune receptors. The objective here is to develop novel techniques for the production of cell wall derived libraries and then use those molecules as tools for the investigation of innate immune signaling. It has long been known that a small piece of the bacterial cell wall, muramyl‐dipeptide (MDP), has immune stimulatory properties. More specifically, MDP has been found to bind to Nucleotide Oligomerization Domain‐containing protein 2 (NOD2) to initiate a Nod2‐dependent immune response. The power of microarrays lays in their unprecedented capacity to simultaneously interrogate tens to hundreds of thousands of immobilized probes. Here we developed a synthesis for amine functionalized PG derivatives with varying linker length and position, and applied them to NHS‐functionalized glass slides to investigate their interactions with Nod2, and other nod‐like proteins. N‐acetyl muramic acid (NAM) serves as a key building blocks PG fragments. We report a synthetic strategy to access biorthogonal 3‐lactic acid NAM derivatives in large quantities. By utilizing metabolic cell wall recycling and biosynthetic machineries reported previously in our lab, the NAM derivatives were installed into the backbone of Gram‐positive and Gram‐negative bacteria. The whole cells are labelled via click chemistry and visualized using super‐resolution microscopy.Support or Funding InformationCobre (Center of Biomedical Research Excellence at the University of Delaware);NSF (National Science Foundation);U01(NIH) grant.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of the Interaction between ISTL1 and Exo70E2 in Arabidopsis
Authors: Camacho Lizbeth 
Published: June 2021

Internal and external stimuli at the plasma membrane are the cause of dynamic changes that affect the composition of the plasma membrane which is responsible for the proper responses, survival, and development of an organism. The Endosomal Sorting Complex Required for Transport, otherwise known as ESCRT, is a highly conserved pathway in all eukaryotes. This pathway allows for endocytosis, and endosomal trafficking which tags proteins for degradation. It is also responsible for the sorting of endocytic vesicles to the vacuole. This mediates the responses of plants. It has also been noted that there is a possible relationship between the ESCRT pathway and the exocyst. The exocyst is an octameric protein complex that directs vesicles after the Golgi complex to specific destinations of the the plasma membrane. It has recently been proposed that Exo70 proteins localize vesicle traffic.Support or Funding InformationNational Science FoundationNIH Minority Access to Research (MARC) 5 T34 GM 8395‐29 California State University NorthridgeOtegui LabThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Transdermal Detection of Hydrogen Sulfide in Humans
Authors: Wiest Elani Fourie , Brandenburg Jacquelyn , Clark Ross , Friedrichsen Debra , Silaski Grayson , Qeadan Fares , Howdieshell Thomas , Shekarriz Reza , Kanagy Nancy 
Published: June 2021

We have developed a novel approach to measure circulating hydrogen sulfide (H2S) non‐invasively as a potential way to diagnose and monitor endothelial dysfunction and peripheral artery disease (PAD). PAD is a life‐threatening condition caused by arterial constriction and obstruction of blood flow leading to limb ischemia. Current methods to diagnose and monitor PAD lack sensitivity, are expensive, and technically difficult. Recent studies indicate that decreased H2S production is an underlying cause of PAD. Reduced plasma H2S also correlates with endothelial dysfunction in individuals with untreated hypertension, diabetes, sleep apnea, and other cardiovascular diseases. The Transdermal Gas Sensor (TAGS) device was designed to measure transdermal H2S to test the hypothesis that the diffusion rate (and therefore gas phase concentration) of H2S is directly proportional to dermal blood flow in individuals without endothelial disease. Healthy volunteers between the ages of 21–65 were recruited. Exclusion criteria included subjects currently treated for hypertension, hyperlipidemia, and diabetes. Smokers and pregnant women were also excluded. We observed a linear association between TAGS readings and Laser Speckle Contrast Imaging (LSCI), a gold standard measure of skin blood flow, when readings were adjusted for age in the model (r = 0.5717, p = 0.0132, n = 18). Therefore the TAGS device is not inferior to LSCI in measuring skin blood flow in healthy individuals using a non‐inferiority margin of 5 (p = 0.0394). The TAGS device has the potential, therefore, to serve as a more sensitive, economical, and easy to use diagnostic tool to use H2S detection as a predictor of dermal blood flow.Support or Funding InformationNSF: R41 NSF FP00000502. NHLBI: R41HL121871This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigating the Evolution of the Allosteric Regulation of Glutamate Dehydrogenase
Authors: Lekova Virzhiniya , Bell Ellis 
Published: June 2021

Glutamate dehydrogenase exhibits no allosteric regulation in bacteria and archaea, while in mammals it has complex allosteric regulation. The major activator is ADP, and inhibitor is GTP. The enzyme is a homohexamer in all organisms. Ciliates and animals have an antenna domain, not present in other unicellular eukaryotes, or in bacteria and archaea. Using GDH from E. coli, S. cerevisiae, N. tabacum, the unicellular eukaryotes D. discoideum, T. pyriformis, and P. tetraurelia, D. melanogaster, and Bos taurus sequence alignments show the presence of an antenna domain in GDH enzymes from D. discoideum, T. pyriformis, and P. tetraurelia. Conserved residues in all living organisms have been idntified as well as conserved residues present only in eukaryotes or prokaryotes allowing comparisons to be made between residues involved in structure and catalysis as compared to regulation. Homology models of GDH from these organisms have been used to examine the structural differences of GDH from various evolutionary levels. The antenna domain showed similarity to a methyl transferase of D. aromatica, suggesting that its appearance results from an insertion mutation. These observations suggests complex allosteric regulation is linked to multicellularity and the metabolism of ammonia, as well as usage of glutamate as a neurotransmitter.Supported by NSF Grant MCB 0448905 to Ellis Bell
Gene Profiles of Dengue Virus Targeted Host Biological Processes and Pathologic Markers
Authors: Muhie Seid , Hammamieh Rasha , Yang David , Jett Marti 
Published: June 2021

Dengue is one of the most prevalent arthropod‐borne viral illnesses in humans with neither selective therapy nor an approved vaccine. Understanding its molecular targets and biological processes may contribute towards developing sensitive diagnostic markers and/or effective therapeutics. Leucocytes obtained from 15 US Army Ranger Cadets before and after 9 weeks training were in vitro treated with dengue virus serotype 2. cDNAs of isolated RNAs were profiled using glass microarray chips containing ∼10,000 human gene targets. Statistical analyses gave ∼180 genes specifically regulated by dengue both in the presence and absence of severe physiological stress. Ontological‐enrichment showed that genes forming spliceosome, ribosome, translation initiation and proteosome complexes were up regulated while those involved in translation elongation, protein modification and targeting were down regulated. It seems that dengue virus mainly targets host protein biosynthesis with the net effect of blocking formation of functional host proteins while it gears the host's machineries towards promoting its own replication and pathogenesis. Subsets of 180 genes were also specific to dengue even in the presence of infections with other pathogens. These have the potential to be used as diagnostic markers irrespective of the physiological and pathologic conditions of the host.We are grateful to DTRA for funding.
Conserved hydrophobic ensembles in protein kinases: their integrating and regulatory roles
Authors: Kornev Alexandr P. , Ten Eyck Lynn F. , Taylor Susan S. 
Published: June 2021

Protein kinases represent a large protein superfamily which regulates numerous processes in living cells. Structures of several serine‐threonine and tyrosine kinases were investigated by recently developed bioinformatics method: Local Spatial Patterns alignment. Previously this method proved to be an effective tool, which is capable to detect highly conserved spatial patterns of amino acid residues. These residues do not form traditional motifs in terms of sequence or protein fold and can not be detected by traditional methods. An example of that was discovery of “the spine” in protein kinases, an important regulatory element. In this work we report detailed analysis of the protein kinase catalytic core. A set of conserved ensembles comprised mostly by hydrophobic residues was detected. The formations traverse protein kinase molecules and play integrating and regulatory roles. They are anchored to the F‐helix located in the middle of the large lobe, which acts as an organizing “hub” providing exact positioning of the key catalytic and regulatory elements.This work was supported by National Science Foundation Grant NSF‐DBI 99111196 and National Institute of General Medical Sciences Grants GM70996 (to L.F.T.E.); GM19301 and National Science Foundation Grant DBI0217951 (to S.S.T.).