bluuebunny/crossref_metadata_2025_split huggingface dataset:
Nicotinic acetylcholine receptor silent agonists modulate inflammation
Authors: Simard Alain , Richter Katrin , Roy Danika , Espinosa Eduardo Soto , Roy Patrick , Godin Jean‐Rémi , Grau Veronika , Quadri Marta , Horenstein Nicole , Papke Roger 
Published: June 2021

Nicotinic acetylcholine receptors (nAChRs) have been increasingly studied for their immune regulatory properties. nAChRs are best known to function as ligand‐gated ion channels in neurons, however, recent evidence suggests that nicotine modulates inflammation by desensitizing non‐neuronal nAChRs, rather than by inducing channel opening. The objective of this study was to determine whether nAChR‐dependent anti‐inflammatory properties are due to channel desensitization. To this end, we made use of various nAChR ligands, including: 1) nicotine, a nAChR non‐selective agonist that also desensitizes the receptors; 2) AR‐R17779, an α7 nAChR subtype‐selective agonist; 3) α‐bungarotoxin, an α7 and α9 nAChR‐selective competitive antagonist, 4) mecamylamine, a non‐selective and non‐competitive antagonist; 5) N,N‐diethyl‐N’‐phenylpiperazine (DEPP)‐based silent agonists that selectively induce the desensitized state of α7 nAChR while producing little or no channel opening. We first evaluated the effects of these various nAChR ligands on the modulation of mouse and human monocyte/macrophage numbers, phenotype and cytokine production. Monocyte phenotype, cellular viability, proliferation and cytokine production were assessed by flow cytometry. In bone marrow‐derived monocyte/macrophage (BMDM) cell cultures, nicotine, mecamylamine and silent agonists reduced pro‐inflammatory monocyte numbers by affecting their viability and proliferation. The silent agonists also significantly reduced both LPS‐ and ATP‐induced cytokine production by mouse BMDMs. In human peripheral blood mononuclear cells (PBMCs), the silent agonists tested had modest effects on monocyte numbers and phenotype. However, LPS‐ and ATP‐induced cytokine production by human blood‐derived immune cells (PBMCs and whole blood) was inhibited by silent agonists. Taken together, our results suggest that nAChRs modulate pro‐inflammatory monocyte numbers and function via receptor desensitization rather than channel opening. nAChR silent agonists may thus be a novel source of anti‐inflammatory compounds that could be used for the treatment of inflammatory disorders.Support or Funding InformationThis work was supported by the MS Society of Canada, the New Brunswick Health Research Foundation and 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.
Biosynthesis of IAA and its role as signal molecule in the phytopathogenic bacterium Pseudomonas savastanoi
Authors: Calvillo Adrian Pintado , Pastor Victoria , Cerezo Miguel , Flors Victor , Ramos Cayo 
Published: June 2021

The strains of Pseudomonas savastanoi are responsible for the formation of tumors or excrescences in the aerial parts of woody plants, belong to the Pseudomonas syringaecomplex and are classified in the pathovars savastanoi (Psv, olive), nerii (Psn, oleander), retacarpa (Psr, Spanish broom) and fraxini (Psf, ash). The production of indole‐3‐acetic acid (IAA) in Psv and Psn is a factor of pathogenicity essential for the correct development of tumors in olive and oleander. In addition, IAA has been described as a signaling molecule capable of modifying gene expression of other virulence factors, such as the type III and type VI secretion systems. Most strains of P. savastanoi produce IAA from tryptophan through the indole‐3‐acetamide pathway. However, the iaaMHoperon is not present in Psf and it is not frequent in other pathovars of the P. syringae complex. Previously, our research group showed that a Psv ΔiaaMH mutant drastically reduces IAA levels and does not induce olive tumors. This mutant produces an IAA concentration in culture supernatant similar to produce by strains of Psf and other strains of P. syringae lacking these genes. These results suggest that the existence of an alternative route of IAA biosynthesis in Pseudomonas. In order to identify an alternative route, a comparative genomic analysis of P. savastanoi and other strains in the P. syringae complex was carried out. Currently, Psv mutants have been constructed in genes potentially involved in the biosynthesis of IAA. Additionally, a metabolic analysis of indolic compounds has been conducted to determine possible intermediates of this route. Finally, we carried out a comparative transcriptomic analysis (RNAseq) of the wild type Psv and its mutant Psv ΔiaaMHand was determined the impact of the addition of exogenous IAA to cultures of P. savastanoi. In summary, this work aims to identify new routes of IAA biosynthesis in P. savastanoi and to study regulatory roles of the phytohormone, IAA, on the expression of other virulence factors in Pseudomonas.Support or Funding InformationAGL2017‐82492‐C2‐1‐R (MINECO‐FEDER)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 Study of the Impact of Genetic Polymorphism of Leptin Gene G2548A on Obesity and Its Related Traits
Authors: Kamalakannan Theekshitha , Robert Roshan Sam , Haqh Mohammed Shadabul 
Published: June 2021

BackgroundLeptin is a neuroendocrine protein hormone with 167 amino acids mainly produced by adipose tissue that regulates food intake and energy expenditure to maintain relative stability of body weight and energy storage over long period of time. A single nucleotide polymorphism in leptin, consisting of G to A substitution at nucleotide −2548 upstream from ATG start site (LEP G‐2548A, rs7799039) has been found to influence leptin expression, possibly at the transcriptional level and was associated with increased leptin secretion in adipocytes. However, these high levels of leptin do not serve their function in controlling the obesity of these individuals possibly due to a condition known as ‘leptin resistance’.AimAssess the risk of the genetic polymorphism of leptin gene G‐2548A on obesity.Material and MethodA case‐control study which consisted of 66 individuals of different BMI (46 normal, 16 overweight and 4 obese). The different genotypes of leptin G‐2548A were determined by restriction fragment length polymorphism polymerase reaction after DNA extraction from saliva samples.ResultsA high risk of obesity was seen among carrier of AA genotype compared to other genotypes (GG&GA) (OR=2.6, 95% CI= 0.9–7.5). Furthermore AA genotype was found to be a risk factor for abdominal obesity (OR=1.3, 95%CI= 0.5–3.7).ConclusionCarrier of AA genotype of leptin G‐2548A polymorphism are at risk of obesity.Support or Funding InformationRak Medical and Health Sciences UniversityThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Adenomatous Polyposis Coli‐like protein (APCLP) Functions as a Novel Negative Regulator of NF‐kB Signaling in Colon Cancer Cells
Authors: Martin Matthew , Mundade Rasika , Lu Tao 
Published: June 2021

Colon cancer (CRC) is the second leading cause of cancer related deaths in the United States. CRC is marked by aberrantly activated signaling of the nuclear factor of kappaB (NF‐kB), a family of transcription factors which regulate wide varieties of cellular processes. Despite recent advances in comprehension of players of NF‐kB signaling, deeper understanding of its regulation is imperative for the development of novel cancer therapeutics. Using the powerful validation‐based insertional mutagenesis (VBIM) technique, we recently discovered APCLP as a novel negative regulator of NF‐kB. The objective of this study is to elucidate the role APCLP in regulating NF‐kB signaling in CRC at the molecular and biological levels and to understand the mechanism by which this regulation occurs. To determine the biological effect of APCLP on NF‐kB signaling, we used lentiviral vectors to either overexpress or knockdown (shRNA) APCLP in human CRC cell lines (HT‐29, HCT116, DLD‐1). We show that overexpression of APCLP decreased the NF‐kB activity, reduced cellular proliferation, migratory ability, as well as anchorage‐independent growth of cells while knockdown of APCLP had an inverse effect. Furthermore, in vivo experiments in a xenograft mouse model confirmed that APCLP overexpression impeded whereas shRNA knockdown promoted tumor growth. To study the mechanism by which APCLP regulated NF‐kB signaling, we conducted co‐Immunoprecipitation experiments and confirmed that APCLP and the major subunit of NF‐kB, p65, may complex or bind directly to each other. Studies are ongoing regarding the mechanism of interaction between APCLP and p65. In summary, discovery of APCLP and understanding of its molecular mechanism and biological function are significant because the knowledge acquired from this study could lead to utilization of APCLP as a potential biomarker and therapeutic target in CRC as well as other cancers that are driven by hyperactivated NF‐kB.Support or Funding InformationThis work is supported by NIH‐NIGMS Grant (# 1R01GM120156‐01A1 to TL), NIH‐NCI Grant (# 1R03 CA223906‐01 to TL), V foundation Kay Yow Cancer Fund (Grant 4486242 to TL), and 100 VOH Grant (# 2987613 to TL).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Celecoxib Impairs Differentiation of Primary Human Skeletal Myoblasts
Authors: Geddis Alyssa , Kolb Alexander , Matheny Ronald 
Published: June 2021

Non‐steroidal anti‐inflammatory drugs (NSAIDs) are medications commonly consumed for their anti‐inflammatory, analgesic, and antipyretic effects. NSAIDs act by inhibiting cyclooxygenase (COX) enzymes, which mediate the synthesis of pro‐inflammatory prostaglandins from substrates such as arachidonic acid. Despite the widespread use of NSAIDs, the risks of taking NSAIDs are only beginning to be defined; in particular, their effects on highly metabolic tissues such as skeletal muscle. We therefore sought to determine whether celecoxib, an NSAID selective for COX2, impaired the differentiation and hypertrophy of cultured primary human skeletal myoblasts. Myoblasts were exposed to celecoxib at concentrations from 50 μM – 0.08 μM and allowed to differentiate for 72 hours. Vehicle‐treated myotubes showed a robust fusion index and myotube area as determined by immunocytochemistry and confocal microscopy; however, the fusion indices and areas of myotubes exposed to 50 μM and 25 μM celecoxib were significantly reduced (90% reduced fusion index and 89% reduced myotube area in cells exposed to 50 μM celecoxib; P<0.001, and 33% reduced fusion index and 25% reduced myotube area in cells exposed to 25 μM celecoxib; P<0.001). The total nuclei present across all treatments did not differ. Myoblasts treated with NS‐398 (a COX2‐selective pharmacological inhibitor) showed no impaired differentiation at any concentration. These observations were associated with dose‐dependent decreases in expression of myosin heavy chain mRNA transcripts of slow (Myh7) and developmental (Myh3 and Myh8) myosins, and exposure to 50 μM celecoxib virtually eliminated expression of all myosin heavy chain transcripts (P<0.001 for all). Phosphorylation of AKT (S473), p70S6K (T389), and S6 (S235/236) were dose‐dependently reduced in celecoxib‐treated myotubes (P<0.05), whereas administration of NS‐398 had no effect on phosphorylation of any of these molecules at any concentration. These data reveal that celecoxib inhibits differentiation of primary human skeletal myoblasts, possibly through a mechanism that regulates the activation of molecules involved in protein synthesis. Celecoxib‐mediated impairment of human myoblast differentiation may be independent of its actions on COX2, because treatment with NS‐398 did not alter differentiation. The exact mechanism through which celecoxib acts to alter muscle cell differentiation and physiology remains to be established.The views expressed in this abstract are those of the authors and do not reflect the official policy of the Department of Army, Department of Defense, or the U.S. Government. This abstract has been approved for public release with unlimited distribution.Support or Funding InformationUS Army Military operational Medicine Research ProgramThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Large Tumor Suppressor (LATS) and 14‐3‐3 Regulate Mixed Lineage Kinase 3 (MLK3) Subcellular Localization
Authors: Kasturirangan Srimathi , Chadee Deborah N 
Published: June 2021

Mixed Lineage Kinase 3 (MLK3) is a mitogen‐activated protein (MAP) kinase kinase kinase (MAP3K) that activates multiple MAPK signaling pathways in response to cytokines and stress stimuli. MLK3 is important for ovarian cancer cell proliferation and invasion. However, few upstream regulators of MLK3 are known, and MLK3 subcellular localization in ovarian cells is not well understood. LATS (Large Tumor Suppressor), a Ser/Thr kinase, is activated by cell‐cell contact to inhibit proliferation, and also regulates apoptosis and mitotic exit. We observed that confluent ovarian epithelial cells had high levels of phosphorylated, activated LATS (p‐LATS). Furthermore, ovarian cancer cells had substantially lower amounts of p‐LATS in comparison to ovarian epithelial cells. Using confocal microscopy and cell fractionation, we found that MLK3 is both cytoplasmic and nuclear in ovarian cells. Co‐immunoprecipitation assays indicated that LATS interacts with MLK3 and promotes MLK3/14‐3‐3 association and MLK3 cytoplasmic localization; thereby linking the MAPK and Hippo signaling pathways. In conclusion, our results identify LATS as a novel regulator of MLK3 that affects MLK3 subcellular localization in ovarian epithelial cells.Support or Funding InformationR15‐CA199164This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigation of Differential Post Transcriptional Regulation of 4E‐BP1 and 4E‐BP2 in Eukaryotic Cells Under Stress
Authors: Gobbooru Shruthi , Low Woon‐Kai , Kommaraju Sai Shilpa 
Published: June 2021

eIF4E‐binding proteins (eIF4E‐BPs or 4E‐BPs) are a family of proteins that inhibit the initiation of eukaryotic translation. They bind to eIF4E and inhibit the formation of eIF4F complex which is crucial for cap‐dependent translation initiation. There are 3 isotypes of 4E‐BPs, 4E‐BP1, 4E‐BP2 and 4E‐BP3. They are nearly 60% identical at the amino acid level. According to the NCBI reference sequences, 4E‐BP2 has a long 3′ UTR (6.8kb) whereas 4E‐BP1 (428 bp) and 4E‐BP3 (315 bp) do not. The presence of this relatively longer 3′ UTR could indicate the presence of cis‐acting regulatory elements (CREs) that are specific to 4E‐BP2. Under various cellular stress conditions, global translation is shut down to conserve energy. To examine the role of 3′ UTRs in promoting the availability of these 4E‐BPs, during stress, to decrease the cap‐dependent translation, 4E‐BP1 and 4E‐BP2 were both studied in relation to their gene expression (transcription and translation) under various stress conditions (ER stress, oxidative stress), cell cycle arrest and mTOR inhibition (conditions of growth arrest and inhibition, respectively). Under conditions of acute ER stress, 4E‐BP1 showed nearly 3‐fold increased expression than in control treatment (DMSO) whereas 4E‐BP2c and 4E‐BP3 did not show significant increase using qPCR. No changes in 4E‐BP1 protein levels (controls vs various conditions of stress) were detected whereas different 4E‐BP2 protein levels were seen under various conditions of cellular stress. Future studies will include investigation of promoter activity of these three gene products. The data suggests possible differential regulation of 4E‐BPs under cell stress condition.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
FGFR2 and TRPA1 Interaction in Lung Cancer
Authors: Kyriakopoulou Eleni , Gamper Nikita 
Published: June 2021

Lung cancer is the leading cause of cancer‐related mortality worldwide. The most common type of lung cancer is lung adenocarcinoma (LUAD) accounting for approximately 40% of all cases. Recent studies unravelled the oncogenic role of the Fibroblast Growth Factor Receptor‐2 (FGFR2) in LUAD. Moreover, it has been shown that the Transient Receptor Potential Ankyrin‐1 (TRPA1), a non‐selective cation channel commonly recognised for its role in somatosensation, is involved in lung malignancies, yet the exact mechanism of this involvement remains unknown. Thus, the aim of this study was to test for potential interaction between FGFR2 and TRPA1 in transfected HEK‐293T cells and LUAD cells as well as to map the binding sites on both proteins. Protein interaction techniques, including immunoprecipitation and Proximity Ligation Assay (PLA) in HEK‐293T cells overexpressing FGFR2 and TRPA1, revealed complex formation between the two proteins. Microscale Thermophoresis (MST) performed on purified proteins indicated direct interaction (Kd = 122.8 ± 23.4 nM) and involvement of the N‐terminal Ankyrin repeat (AR) domain of TPRA1 in the interaction with FGFR2. This result was verified by immunoprecipitation experiments performed on HEK‐293T cells overexpressing FGFR2 and truncated versions of TRPA1. On the other hand, mutations on the C‐terminal proline‐rich motif of FGFR2 pulled down TRPA1 less effectively compared to the wild type C‐terminal domain of FGFR2. These results suggest the direct binding through the AR domain of TRPA1 and the Proline‐rich motif of FGFR2. Complex formation was also shown in a LUAD cell line (HCC‐515) that endogenously expresses both proteins. In conclusion, these results describe for the first time the novel interaction between a tyrosine kinase receptor and an ion channel that could have an effect in signal transduction and subsequently in cancer progression.Support or Funding InformationUniversity of Leeds, BBSRCThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Does DMD‐10 independently affect levels of GLR‐1?
Authors: Nammour Josette , McGehee Annette 
Published: June 2021

We aim to study the mechanisms that affect the Caenorhabditis elegans glutamate receptor GLR‐1. The roles of genes that may be involved in the regulation of GLR‐1 levels are assessed using GLR‐1‐dependent behavioral tests such as spontaneous reversals assays and nose touch assays. Results from our laboratory have shown that dmd‐10 mutants exhibit a low rate of spontaneous reversals when compared to the wildtype strain. We generated rescue strains by cloning DMD‐10 and microinjecting it into dmd‐10 mutant worms to assess the effect of wildtype DMD‐10. The constructs developed for microinjection contained the dmd‐10 gene being driven by either the glr‐1 promoter (Pglr‐1::dmd‐10) or its own promoter (Pdmd‐10::dmd‐10). Pglr‐1 rescue strains, which consist of the dmd‐10 mutant and wildtype DMD‐10 expressed in cells that express GLR‐1, were compared to the dmd‐10 mutant strain and wildtype using behavioral assays. Pglr‐1 rescue strains did not exhibit behavioral differences from dmd‐10 mutant worms. Pdmd‐10 rescue strains have been generated and are currently being tested for behavioral rescue. Additionally, we are developing constructs that will allow us to visualize the location of dmd‐10 neurons in the wildtype worms using the red fluorescent protein, mCherry.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 potential role of IDA‐1 and neuromodulatory signaling in GLR‐1 regulation
Authors: McLaughlin Shannon 
Published: June 2021

In organisms, behavior can be a reflection of environment and nervous system activity. Previous studies have shown that the DAF‐7 signaling pathway in C. elegans is affected by the environment, such as inadequate food supply or high temperature. The DAF‐7 signaling pathway also regulates the glutamate receptor GLR‐1, and increased amounts of GLR‐1 occur in C. elegans strains with mutations in the genes of the DAF‐7 signaling pathway. Additionally, spontaneous reversal behavior depends on GLR‐1 and increased reversals are evident in the daf‐7 mutants. The mechanism by which the DAF‐7 pathway affects GLR‐1 levels is currently unknown. Our research focuses on understanding this mechanism by investigating the role of the candidate gene ida‐1. IDA‐1 was identified because of its involvement in regulating presynaptic neurotransmission and its presence in neurons along the ventral nerve cord (VNC) that express GLR‐1. To test if IDA‐1 was involved in the DAF‐7 pathway regulation of GLR‐1 an assessment of spontaneous reversals in single and double mutants was performed along with fluorescence imaging. Results supported the idea that IDA‐1 plays a role in the DAF‐7 pathway regulation of GLR‐1. To follow up on IDA‐1's potential regulation of GLR‐1 levels, the involvement of IDA‐1 in neuromodulatory signaling was examined. IDA‐1 is normally found on the surface of dense core vesicles (DCV's) and is involved in normal neuropeptide and DCV signaling. To investigate if IDA‐1 is working through the DCV pathway to regulate GLR‐1 levels several strains will be obtained with mutations in various places in the DCV pathway. Future directions will include analysis of these mutants using fluorescence imaging as well as direct protein level analysis through immunoblotting.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Arl4A‐PAK1 complex establishes a positive‐feedback loop contributing to PAK1 activation for cell migration
Authors: Lee Fang‐Jen Scott , Chen Kuan‐Jung , Chiang Tsai‐Chen , Yu Chia‐Jung 
Published: June 2021

Cell migration requires coordination of multiple signaling pathways involving in membrane dynamic and cytoskeleton rearrangement. Arf‐like small GTPase Arl4A has been shown to modulate the Golgi organization and actin cytoskeleton remodeling. However, the evidences for Arl4A function in cell migration are insufficient. Here, we report that Arl4A acts with a serine/threonine protein kinase PAK1 to modulate cell migration. We first observe that Arl4A directly interacts with PAK1, and recruits PAK1 to the plasma membrane in a GTP‐dependent manner. Unexpectedly, we found that PAK1 can recruit a cytoplasmic myristoylation‐deficient Arl4A‐G2A mutant, but not GTP‐binding defective Arl4A‐T34N mutant, to the plasma membrane. Thus, these results suggest that there is a positive feedback loop between Arl4A and PAK1, which mutually recruit each other to the plasma membrane. Furthermore, we found that Arl4A promotes cell migration in a GTP‐dependent manner and Arl4A‐PAK1 interaction is critical for Arl4A‐induced cell migration, which is independent of Rac‐PAK1 interaction. Overall, this study shows that Arl4A plays important role for cell migration through direct interaction of PAK1 without effecting Rac1 activation. Our data presented here provide evidence for a novel mechanism for PAK1 regulation and activation.Support or Funding InformationThis work was supported by grants from the NHRI in Taiwan (NHRI‐EX106‐10601B1) to F.‐J. S. LeeThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.