bluuebunny/crossref_metadata_2025 huggingface dataset:
Talk title Bridging the gap between RNA editing and modification: A 10‐year solution to a 25‐year problem
Authors: Alfonzo Juan D.
Published: June 2021

All types of nucleic acids in cells undergo naturally occurring chemical modifications, including DNA, rRNA, mRNA, snRNA, and most prominently tRNA. Over 100 different modifications have been described and every position in the purine and pyrimidine bases has been seen modified; the sugar is often also modified. Despite recent progress, the mechanism for the synthesis of most modifications is not fully understood partly due to difficulty in reconstituting modification enzyme activity in vitro. Cytosine to uridine (C to U) editing of tRNAs in eukaryotes was first discovered over 20 years ago, yet due to the lack of an in vitro assay, the editing enzyme is still at large and the editing mechanism remains unsolved. In this work we show that cytosine 32 in the anticodon loop of tRNAThr is methylated to 3‐methylcytosine (m3C) as a pre‐requisite for C to U deamination. Formation of m3C in vitro requires the presence of both TbTRM140 (the m3C methyltransferase) and the TbADAT2/3 (the A to I editing deaminase). Once m3C is formed the same set of enzymes are required for further deamination from m3C to form 3‐methyluridine (m3U). In our previous work, we also showed that TbADAT2/3 was a highly mutagenic enzyme randomly deaminating DNA. Now we provide evidence that when co‐expressed with the methylase its mutagenic ability is kept in check via complex formation with TRM140. This helps explain how T. brucei escapes “wholesale deamination” of its genome while harboring both enzymes in its nuclear compartment. This observation has implications for the control of another mutagenic deaminase, human AID, and provides a rationale for its regulation.

Support or Funding Information

NIH GM084065

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Telomerase Expression in Myeloid Cells is Essential for Pulmonary Fibrosis
Authors: Liu Tianju , De Los Santos Francina Gonzalez , Zhao Yuyue , Wu Zhe , Phan Sem H.
Published: June 2021

Shortened telomeres and mutations in telomerase reverse transcriptase (TERT) or its RNA component (TERC) have been reported in some patients with fibrotic interstitial lung disease, such as idiopathic pulmonary fibrosis (IPF). However telomerase is induced in lung fibroblasts from such patients without evidence of telomere shortening, and selective TERT deficiency in mesenchymal cells significantly reduced pulmonary fibrosis in an animal model. Moreover TERT in the bone marrow (BM) is essential for fibrosis in the same model. However, the precise cellular source of TERT in the BM is unknown. The objective of this study was to elucidate further the identity and the role of the TERT‐expressing BM‐derived cells in the injured lung by evaluating the effects of selective TERT deficiency in myeloid (LysM+ cells) on bleomycin (BLM) induced pulmonary fibrosis. The myeloid cell specific TERT knockout mice (LysM‐TERT KO) were created by crossing floxed TERT mice with LysM‐Cre mice. The results showed that BLM induced significant TERT expression in BM‐derived CD45+ cells in lungs of control (WT) mice, which was not observed in the LysM‐TERT KO mice. This was also associated with the loss of TERT expression in whole lung tissue in LysM‐TERT KO mice while TERT expression in the BM was reduced by >80%. Interestingly, BLM‐induced TERT expression in lung CD45− cells in WT mice was also abolished by myeloid cell TERT deficiency in LysM‐TERT KO mice. The myeloid cell specific TERT deficiency caused significant reduction in pulmonary fibrosis along with diminished lung expression of pro‐fibrotic cytokines. Additionally, LysM‐TERT KO mice showed diminished numbers of c‐kit+ lung hematopoietic progenitor cells (LHPC) after BLM treatment in comparison with WT controls. Taken together these findings suggest a pro‐fibrotic role for the induction of TERT in myeloid cells.

Support or Funding Information

This study is supported by NIH grants HL052285 and HL112880.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Chronic Ethanol Intake on High‐Fat Diet Induced Metabolic Dysfunction in Mice
Authors: Coker Caitlin R. , Bingaman Sarah S. , Arnold Amy C. , Silberman Yuval
Published: June 2021

High‐fat diet (HFD) induced obesity is associated with insulin resistance and is a major risk factor for numerous co‐morbidities such as type II diabetes. Conflicting clinical and preclinical evidence suggests that moderate alcohol consumption improves insulin sensitivity in obese subjects, and therefore may act to protect against obesity‐related metabolic comorbidities. However, the impact of free choice ethanol (EtOH) intake and HFD consumption on insulin action and glucose homeostasis in animal models remains unclear. Therefore, the overall goal of these studies was to determine if long‐term EtOH intake may alter insulin and glucose function in a mouse model of HFD induced obesity. Six‐week old male C57Bl/6J mice were given ad libitum access to HFD (n=10; 60% calories from fat) or control diet (Chow, n=10; 10% calories from fat) for two weeks. Mice were then given increasing volumes of EtOH (3%, 7%, and 10%) over one week as an EtOH‐ramp initiation period for subsequent 24‐hour access to a two‐bottle choice of water or 10% EtOH (vol/vol, in water). Body mass, EtOH and water intake, and EtOH preference was assessed every 24 hours for six weeks. HFD mice had significantly higher body mass compared to Chow mice (p<0.001). By the end of the study, Chow mice consumed significantly more EtOH than mice on HFD (p<0.001) and had higher preference towards EtOH (p<0.001). At the end of treatment, we measured 4‐hour fasting glucose levels and performed insulin tolerance tests (to examine the ability of intraperitoneal insulin to suppress blood glucose levels) in half of the mice in each group. These metabolic data were compared to weight and aged matched mice given HFD or Chow without EtOH access. Insulin sensitivity was significantly reduced in HFD mice (p=0.003; two‐way ANOVA), and EtOH consumption did not alter HFD‐induced insulin resistance. Furthermore, there appeared to be a trend for increased 4 hour fasting glucose levels in HFD (185.8±9.4), HFD+EtOH (176±25.9) and Chow+EtOH (177.8±21.6) groups compared to Chow alone (156.4±6.2). Together, these results suggest that moderate EtOH drinking in a two‐bottle choice model does not improve insulin sensitivity in HFD mice, but that higher levels of EtOH intake in control diet mice may cause impaired glucose homeostasis that may not be due to changes in insulin sensitivity.

Support or Funding Information

Penn State Dept of Neural and Behavioral Science, NIH grants AA022937 and HL122507

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Anticancer and pro‐apoptotic effects of Triphala extract in human breast cancer MCF‐7 cells
Authors: MURTHY MALLADI SATYANARAYANA
Published: June 2021

Triphala, an ayurvedic formulation, showed a substantial antiproliferative effect against various human cancer cells including breast cancer cells. In vitro, anticancer and pro‐apoptotic studies were carried out using MCF‐7 breast cancer cell line to demonstrate its anticancer activity. Cell proliferation by MTT on Human breast cancer cells (1×106/ml) Flow cytometric cell cycle analysis, ELISA based quantification of cytosolic cytochrome c, Bax and Bcl‐2 levels and colorimetric assays of caspases‐3 and ‐9 were performed using breast cancer cells to demonstrate the pro‐apoptotic activity of triphala.

Triphala showed substantial anti‐proliferative activity. In pro‐apoptotic studies, sub‐G0 cell phase population elevation, an increase of cytosolic cytochrome c and Bax levels, decrease of Bcl‐2 levels and enhanced caspases‐3 and ‐9 activities were observed in triphala‐treated MCF‐7 breast cancer cells. The results concluded that anticancer and apoptosis‐inducing potential of triphala in MCF‐7 breast cancer cells. Further detailed studies are in progress.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
3D Imaging and Visualization of Zebrafish and Giant Danio Hearts by Serial Block Face Scanning Electron Microscopy
Authors: Coffing Gabrielle , Hester Kamil , Kyaw Pyi T. , Lafontant Pascal J.
Published: June 2021

3D and 4D imaging of zebrafish heart can provide significant insights into the structural and dynamic nature of the development and growth of this important biomedical model. Transmission electron microcopy (TEM) and tomography allow high resolution ultrastructural imaging of subcellular structures, adding to our understanding of heart architecture in model organisms. Serial block face scanning electron microscopy (SBFSEM) is a novel approach to nanoscale volumetric imaging of tissues and cells. To test the usefulness of SBFSEM in investigations of structure‐function relationship in zebrafish and giant danio hearts, we acquired over 3000 serial block face scanning electron micrographs at 50 to 100 nm intervals, as well as transmission electron micrographs, of the hearts of these fish. We subsequently extracted anatomical and cellular structures of interest in larval and adult fish by semi‐automated segmentation, and generated 3D visualization of various heart regions. Our results show structural similarity in the integration of the compact and spongy hearts in these two species. Similar to the zebrafish, the adult giant danio possesses transitional cardiac myocytes and a network of cardiac fibroblasts in the junctional region. Ultrastructural analysis also reveals remodeling of the ventricular junctional space during repair and regeneration, as well as the presence of luminal lacunae and persistent intra‐myocardial space as late as 3 weeks post‐fertilization. Taken together, our studies demonstrate SBFSEM as a powerful approach to gather nanoscale volumetric structural data in developing, adult, and regenerating giant danio and zebrafish heart

Support or Funding Information

NIH R15 HD084262‐01

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Organ Regeneration and Anti‐Aging Strategies
Authors: Belmonte Juan Carlos Izpisua
Published: June 2021

Aging can be defined as the progressive decline in the ability of a cell or organism to resist stress and disease. Recent advances in cellular reprog‐ ramming technologies have enabled detailed analyses of the aging process, often involving cell types derived from aged individuals, or patients with premature aging syndromes. In my talk I will discuss how cellular reprog‐ ramming allows the recapitulation of aging in a dish, describing novel experi‐mental approaches to investigate the aging process. Finally, I will explore the role of epigenetic dysregulation as a driver of aging, discussing how epige‐netic reprogramming may be harnessed to ameliorate aging hallmarks, both in vitro and in vivo. A better understanding of the reprogramming process may indeed assist the development of novel therapeutic strategies to extend a healthy lifespan.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Shark EvoDevOmics: Linking Teeth and Taste Buds to the Evolution of Tooth Regeneration
Authors: Fraser Gareth
Published: June 2021

The shark dentition is continuously regenerative, with teeth forming from a continuous and permanent epithelial dental lamina. We have identified an intriguing link between the regenerative potential of the shark dentition and the oral taste bud field, both of which form from a common epithelia. We performed RNAseq analyses on specific cell compartments within the continuous dental lamina and cells from the taste‐tooth (lamina) junction, associated with progenitor cells linked to tooth regeneration. We have identified a set of candidate genes involved in the initiation and maintenance of continuous dental regeneration. Expression analyses have verified these candidates and their relationship with specific compartments of the dental lamina associated with tooth regeneration. Our data provide a framework from which we can further investigate stem/progenitor candidates crucial for the process of tooth regeneration, in addition to providing new information on the timing and cyclicity of the shark tooth system. The dental lamina in sharks is an ideal regenerative organ in which to investigate perpetual tooth development, and to further our understanding of the origin of teeth. With these data we can start to construct gene regulatory networks as a tool to further investigate the functional role of these novel candidates. Our findings shed light on the developmental regulation of both the timing of continued dental initiation and the maintenance of stem progenitors for rapid tooth regeneration in an exciting and emerging EvoDevOmic model, the shark.

Support or Funding Information

This research was supported by the following research grants: Natural Environment Research Council (NERC) Standard Grant NE/K014595/1 (to G.J.F); and Leverhulme Trust Research Grant RPG‐211 (to G.J.F.).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Adapting the Chemistry and/or Biology of Proteostasis to Ameliorate Protein Aggregation Diseases
Authors: Kelly Jeffery W.
Published: June 2021

The cellular protein homeostasis, or proteostasis network, regulates proteome function by controlling ribosomal protein synthesis, chaperone and chaperonin mediated protein folding, protein trafficking, protein degradation and related processes. Stress responsive signaling pathways match proteostasis network capacity with demand in each subcellular compartment to maintain or alter cellular homeostasis. The beginning of the seminar will focus on how the proteostasis network can be adapted pharmacologically through unfolded protein response arm‐selective signaling to alleviate the gain‐of‐toxic‐function diseases, including light chain amyloidosis and the transthyretin amyloidosis, where excessive secretion of misfolding and aggregation of proteins leads to a degenerative phenotypes. This strategy will be contrasted with high affinity small molecule binding to the normally folded structural ensemble of an aggregation‐prone protein inside and/or outside of the cell to stabilize the native state, lowering the population of misfolded, misassembly competent states that lead to aggregates, including amyloid fibrils. Lastly, we will cover the progress made to discover autophagy activators. These drug candidates are envisioned to be generally useful for ameliorating multiple neurodegenerative diseases based on human genetic evidence.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cracking the Glycocode: Finding the Signal in the Noise
Authors: Mahal Lara K.
Published: June 2021

Glycans both encode cellular information, such as cell‐cell interactions and cell state (Signal), and must avoid being targeted by pathogens (Noise). This leads to a system in which the sugar code (i.e. the glycan motifs controlling function) is hidden within the noisy milleu of larger heterogenous glycan structures. This talk focuses on use of high‐throughput analytical methods, including our lectin microarray technology and newly developed miRNA‐proxy approach, in tandem with data integration, to decode structure‐function relationships in the glycome. Our work is identifying glycan drivers of biological function including those involved in melanoma metastasis and host‐response to pathogens (e.g. HIV‐1 and influenza), providing a host of new targets for small molecule intervention in these disease states.

Support or Funding Information

This work is supported by: NIH (1U01CA221229‐01, U01 AI111598) and the Melanoma Research Foundation.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Obesity, Insulin Resistance and Cancer: The PI3K connection
Authors: Cantley Lewis C.
Published: June 2021

Retrospective studies have revealed that cancer rates in certain organ sites increase with obesity and some studies have suggested that the subset of obese individuals who develop insulin resistance are more likely to develop cancers than those who are obese but remain insulin sensitive. The risk of developing uterine and breast cancers are particularly high in women who are obese and insulin resistant and these two cancers have the highest rates of activating mutations in PIK3CA, the gene that encodes phosphoinositide 3‐kinase 1 alpha (PI3K1alpha). PI3K1alpha mediates insulin dependent cell growth and the activating mutations in PIK3CA allow this enzyme to be more readily activated by insulin in tumors that express the insulin receptor. Normal endometrial and breast epithelial cells express the insulin receptor and tumors in these tissues often have elevated expression of the insulin receptor. These observations raise the possibility that the elevated serum insulin in patients who are insulin resistant may be contributing to the development of tumors, especially tumors in tissues with PI3K pathway mutations. Many PI3K inhibitors have entered clinical trials and some have been approved. However, since the inhibitors that target PI3K1alpha cause insulin resistance in liver, muscle and fat, they cause hyperglycemia and hyperinsulinemia. Our studies indicate that the elevated insulin in the serum can ultimately re‐activate PI3K1alpha in the tumor, compromising the effect of these drugs. Strategies for making PI3K inhibitors more effective by preventing this feedback reactivation of PI3K1alpha in tumors will be discussed.

Support or Funding Information

Supported by the NCI, the Breast Cancer Research Foundation, the Gray Family Foundation and the Hertog family.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Metalloenzyme‐catalyzed carbon‐carbon bond formation in natural product biosynthesis
Authors: Seyedsayamdost Mohammad R.
Published: June 2021

Microbial secondary metabolites continue to inspire chemists and biologists alike with their fascinating structures and potent biological activities. Investigations into their biosynthetic pathways not only provides possible means of creating these molecules in the laboratory but can also reveal new enzymatic reactions and their underlying mechanisms. In this talk, I will present new transformations that my group has discovered in the biosynthesis of complex natural products, with a focus on carbon‐carbon bond formation catalyzed by metalloenzymes. Our efforts are expanding the scope of reactions that microorganisms have at their disposal in concocting bioactive small molecules.

Support or Funding Information

This work was supported by the National Institutes of Health (DP2‐AI‐124786) and the Burroughs Wellcome Fund.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
IL‐36γ Promotes Alveolar Macrophage Survival During Influenza Infection, Limiting Morbidity and Mortality
Authors: Wein Alexander N. , Dunbar Paul , McMaster Sean R. , Denning Timothy L. , Kohlmeier Jacob E.
Published: June 2021

Influenza viral illness has been a high priority research topic in public health and immune pathogenesis, but the roles of individual cytokines in controlling the infection remains unclear. The IL‐36 family of proteins is made up of novel IL‐1 family members that have shown key roles in lung immunity to bacterial infections, but the role of IL‐36γ in influenza infection is unknown. We investigated the function of IL‐36γ during infection with low and high pathogenesis stains of influenza Il36g mRNA is upregulated in the lung following influenza infection with both H1N1 and H3N2 influenza. Genetic deletion of Il36g results in greatly increased morbidity, mortality, and viral titers compared to mice replete for the gene. This difference correlates with increased pro‐inflammatory cytokines in the airways of mice early during infection. Flow cytometric analysis of the immune compartment revealed a dramatic loss of alveolar macrophages by three days post‐infection. Previous studies have shown that alveolar macrophages have a key role in limiting early viral replication and preventing death during lethal influenza infection. We observed that the alveolar macrophages in Il36g−/− mice have an M2‐like phenotype at baseline, possibly contributing to their rapid depletion. Transfer of WT alveolar macrophages to Il36g−/− mice prior to influenza infection leads to increased survival compared to mock‐transferred Il36g−/− mice. These data indicate that IL‐36γ is a key protein in driving alveolar macrophage survival during influenza infection and protecting against severe disease.

Support or Funding Information

Research Support: R01 HL122559

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
PAF Expression in Skin Tumors of Smokers and Never Smokers: a Potential Role in Skin Cancer Development
Authors: McHowat Jane , Lozano Juan Carlos , Missall Patricia
Published: June 2021

Significant progress has been made in the management and understanding of cancer, but there remains a large disparity in cancer health within our population. One of the most well‐known cancer risk factors, cigarette smoking, maintains a disproportionate prevalence in populations distinguished by socioeconomic status, race, and gender. Previous studies have demonstrated that cigarette smoking is associated with changes in phospholipase A2 (PLA2)‐mediated pathways that are implicated in cancer development and progression. More specifically, PLA2 activation leads to increased platelet‐activating factor (PAF) synthesis, a metabolite that is increased in cancer cells. PAF accumulation is further enhanced with cigarette smoke via inhibition of PAF‐acetylhydrolase, the enzyme responsible for PAF hydrolysis and inactivation. Much remains unknown about the biochemical effects of smoking on the development of skin cancers including squamous cell carcinoma (SCC) and basal cell carcinoma (BCC). However, a recent meta‐analysis study demonstrated that cigarette smoking is associated with a 52% increase in the risk for SCC, but does not appear to modify the risk for BCC. In order to assess the effects of cigarette smoking in these two skin cancers, we analyzed PAF expression in skin biopsies from six smokers and non‐smokers with SCC, and six smokers and non‐smokers with BCC. The biopsy samples were immunohistochemically stained with antibodies specific to PAF and PAF‐receptor. Immunohistochemistry revealed positive PAF and PAF‐R expression with nuclear localization in all patient samples, including SCC and adjacent normal sun exposed skin. There was no significant difference between the PAF or PAF‐R expression in samples of smokers when compared to nonsmokers. However, immunohistochemical analysis demonstrated a significant difference in PAF expression between SCC and BCC cells. SCC cells expressed nuclear PAF, but PAF was not detectable in BCC samples. PAF signal was also not detectable in the basal cell epidermal layer in all patient samples. These findings support a biochemical difference in SCC and BCC cells in association with PAF, which is traditionally upregulated with cigarette smoking.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chlorinated lipids mediate small airway epithelial dysfunction
Authors: McHowat Jane , Albert Carolyn , Hartman Celine , Pike Daniel , Ford David
Published: June 2021

Neutrophil myeloperoxidase is a major mediator of microbicidal activity, catalyzing the conversion of hydrogen peroxide to hypochlorous acid (HOCl), a potent oxidant that reacts with both microbial and host molecular targets. HOCl targets the vinyl ether bond of plasmalogen lipids, resulting in the production of 2‐chlorofatty aldehyde, which is in turn metabolized to 2‐chlorofatty acid (2‐ClFA). Recently, we have shown that free 2‐ClFA levels are significantly associated with acute respiratory distress syndrome (ARDS) in sepsis patients. Lung endothelial cells treated with 2‐ClFA showed increased permeability, surface expression of adhesion molecules, and neutrophil and platelet adherence. These data indicate that plasma 2‐ClFA is a predictor for ARDS, likely through effects on the lung microvasculature. In this study, we determined the effect of 2‐ClFA on human small airways epithelial cell (SAEC) function. Incubation of SAEC with 2‐ClFA (100 nM‐10 μM, up to 24 hours) resulted in a concentration‐ and time‐dependent decrease in electrical resistance, with no change in resistance noted when SAEC were incubated with non‐chlorinated fatty acid (FA). Our recently published studies published recently by us show that 2‐ClFA (1 μM) treatment of lung microvascular endothelial cells resulted in a 2‐fold increase in cell surface expression of intercellular adhesion molecule 1 (ICAM‐1) and a 1.7‐fold increase in vascular cell adhesion molecule 1 (VCAM‐1) after a 4‐hour incubation. We now show that incubation with 2‐ClFA for 4 hours resulted in a 3.3‐fold increase in cell surface expression of ICAM‐1 and a 4.4‐fold increase of VCAM‐1 in SAEC. No increase in adhesion molecules was observed with FA incubation. 2‐ClFA treatment resulted in neutrophil adherence to SAEC and enhanced transmigration across the SAEC monolayer after 2‐ClFA treatment. Neutrophil transmigration across SAEC was 3‐fold greater in a basolateral‐to‐apical direction when compared to apical‐to‐basolateral. Taken together, these data suggest that 2‐ClFA mediates small airways epithelial dysfunction and neutrophil transmigration that may contribute to fluid accumulation and inflammation observed in ARDS patients.

Support or Funding Information

NIH R01 GM115553 (to D.A.F. and J.M.)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pak2 promotes ER‐dependent cardioprotection
Authors: Binder Pablo , Wang Shunyao , Zi Min , Collins Lucy , Liu Wei , Wang Xin
Published: June 2021

Heart failure is a devastating condition affecting millions of people globally with a rising prevalence due to an expansion of the ageing population and epidemics of obesity, diabetes and hypertension. Despite numerous causative factors, the majority of heart failure cases are the consequence of massive cardiomyocyte loss. As cardiomyocytes hold very little potential in replication, exquisite regulation of cellular homeostasis and integrity reliant on protein quality control is crucial for cardiomyocyte survival and function, particularly under stress. A major site of protein quality control is the endoplasmic reticulum (ER). The endoplasmic reticulum is the site for synthesis and folding of secreted and membrane‐bound proteins, which account for 1/3 of total proteins. Loss of ER homeostasis and function culminates in a greater risk of cardiomyocyte death and it has been shown that detrimental ER stress underlies the pathogenesis of many forms of heart disease.

With the aim to uncover novel ER stress regulation mechanisms, we examined the expression pattern of p21activated kinase 2 (Pak2) in cardiomyocytes and myocardium across species. We investigated its role in cardioprotection using Pak2 cardiac deleted mice (Pak2‐CKO) under ER stress conditions. Molecular elucidation unveiled a new mechanism by which Pak2 promoted the protective ER stress response. The translational relevance of this protective mechanism was evaluated in human induced pluripotent stem cells‐derived cardiomyocytes (iPSC‐CMs), and the therapeutic potential was explored by inducing Pak2 activation using a genetic approach or AAV9‐based gene delivery.

We found that Pak2 was an ER‐resident stress‐responsive kinase in cardiomyocytes, and showed that Pak2‐CKO mice under stress manifested defective ER response, cardiac dysfunction and profound cell death. Pak2 loss‐induced cardiac damage was an ER‐dependent event. Mechanistic studies in rat cardiomyocytes and human iPSC‐CMs, revealed that Pak2 regulation of protective ER function was via the IRE1/XBP1s‐dependent pathway. Therapeutically, Pak2 activation was capable of strengthening ER function, improving cardiac performance and diminishing apoptosis, thus protecting the heart from failure. Our findings reveal a novel ER‐centered cardioprotective mechanism, which promotes the protective ER stress response via modulation of Pak2; this novel therapeutic strategy may be used to treat cardiac disease and heart failure.

Support or Funding Information

This research project was supported by the British Heart Foundation

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Policing Cell‐Cell Connections: A Novel Role for the COP9 Signalosome in Mechanisms Underlying Arrhythmogenic Heart Disease and Sudden Death
Authors: Sheikh Farah , Liang Yan , Lyon Robert , Pellman Jason , Mezzano Valeria , Gu Yusu , Dalton Nancy , Lee Mong‐Hong , Iwakuma Tomoo , Nigam Vishal , Peterson Kirk
Published: June 2021

Arrhythmogenic right ventricular cardiomyopathy is a genetic based cardiomyopathy, characterized by ventricular dysfunction, fibrofatty replacement of the ventricle and ventricular arrhythmias leading to sudden cardiac death in young people. ARVC is termed a “disease of the desmosome”; however, limited information exists on how desmosomal protein dysregulation/loss triggers ARVC. To identify novel protein targets that impact desmosomal biology and the desmosomal disease, ARVC, we performed a yeast two hybrid screen using the desmosomal gene, desmoplakin (DSP) as bait to screen an adult human heart cDNA library. We identified CSN6, subunit 6 of the COP9 signalosome, as a novel cardiac desmosomal interacting protein (via desmoplakin, DSP). Traditional functions of the CSN complex (composed of subunits 1–8) are to inhibit protein degradation by “turning off” ubiquitinatin‐mediated protein degradation via deneddlylation; however, the role of CSN6 in the heart is undefined. We show that CSN6 uniquely protects the cardiac desmosome and its loss accelerates desmosome destruction as (i) CSN6 co‐localizes to desmosomal junctions, (ii) CSN6 co‐immunoprecipitates with desmosomal proteins and (iii) hearts from novel cardiac‐specific CSN6 knockout (CSN6‐cKO) mice display selective loss of desmosomal protein levels (and its primary target, connexin43). CSN6 loss is a trigger for ARVC as CSN6‐cKO mice exhibit sudden death and cardiac disease features associated with a biventricular form of ARVC, similar to our cardiac‐specific DSP knockout mice (DSP‐cKO) that is a classic biventricular model of ARVC. CSN6 and DSP‐cKO hearts both selectively display underlying hyper‐accumulation of protein degradation machinery at the cell junction, specifically linking CSN6 pathways to the desmosome. CSN6 pathways are relevant to human ARVC as (i) CSN6 levels are down‐regulated in ARVC hiPSC‐derived cardiac cells that exhibit striking desmosomal defects and arrhythmogenic behavior and (ii) CSN6 localization is lost from cell junctions in a cardiac biopsy from an ARVC patient harboring desmosomal (DSP (R315C) and plakophilin‐2 (PKP2 IVS10‐1 G>C) mutations. Yeast and in silico modeling assays reveal that this DSP R315C mutation is sufficient to abrogate DSP binding to CSN6. We highlight the importance of CSN6 at the cardiac desmosome in maintaining desmosomal protein turnover (“desmophagy”), and suggest that loss of CSN6 and desmosomal protein turnover as new underlying mechanisms driving ARVC and sudden death.

Support or Funding Information

F.S. is supported by grants from National Institutes of Health (NHLBI), American Heart Association and Tobacco Related Disease Research Program.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Role of Neuropeptides in Peripheral Taste Signaling
Authors: Huang Anthony Y. , Wu Sandy Y.
Published: June 2021

Chemesthesis, the general chemical sensitivity of the skin and mucus membranes in the oronasal cavities and being perceived as pungency, irritation or heat, impacts taste responsiveness. The modulation of taste nerve responses is thought to be produced indirectly, via stimulation of peripheral sensory nerve terminals that have ability to release stored transmitters, e.g., calcitonin gene‐related peptide (CGRP) and substance P (SP), onto taste cells implying playing a role in gustatory signaling, by the functions of somatosensory neurons. The underlying assumption has been that neuropeptides exert their effects on taste transmitter secretion in taste buds of mice, a well‐accepted model for studying taste bud functions, as theoretically may relate to the processing of the gustatory information. Indeed, CGRP shapes peripheral taste signals via serotonergic signaling. In this study, using a combination of Ca2+ imaging with cellular biosensors – genetically‐engineered Chinese Hamster Ovary (CHO) cells, transgenic mice expressing green fluorescent protein (GFP) in the specific taste cell type and immunostaining, we showed that the net effect of SP reduced taste‐evoked ATP secretion from mouse taste buds. Specifically, SP elicited phospholipase C (PLC) activation‐dependent intracellular Ca2+ transients in taste cells via neurokinin 1 (NK1) receptors, most likely on glutamate‐aspartate transporter (GLAST)‐expressing Type I cells. Recently, γ‐aminobutyric acid (GABA) as well as the associated synthetic enzyme have been identified in Type I cells. Consequently, Type I cells secrete GABA in respond to SP. Combined with the recent findings that GABA provides negative paracrine feedback onto Receptor (Type II) cells by activating GABAA and/or GABAB receptors and reducing taste‐evoked ATP secretion, our results showed that taste‐evoked ATP secretion from intact taste buds, which remain cell‐cell communication intact, was reduced when SP was co‐applied with taste stimuli in the bath medium. We conclude that SP plays a role as an inhibitory transmitter that shapes peripheral taste signals via GABAergic signaling during processing gustatory information in taste buds. This seemingly tangled complex represents a new concept of taste signaling and that neuropeptides released from afferent sensory terminals play a regulatory role in processing taste signals.

Support or Funding Information

AAA Fellows Grant Award Program Research Seed Grant to AYH (AAA‐3419).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hair Follicle‐derived Neural Crest Stem Cells (HF‐NCSCs) as a Cell Source of Enteric Neurons
Authors: Wang Zitian , ZHAO Hui , LEE Tin Lap , RUDD John , CHAN Wood Yee
Published: June 2021

The enteric nervous system (ENS), which regulates most of the gastrointestinal functions, is derived from neural crest cells during early embryonic development. Abnormal migration and differentiation of neural crest cells may cause defective formation of the ENS, thus leading to enteric aganglionosis (absence of ganglia consisting of neurons and glial cells) in the distal colon. In the present study, we aimed to regenerate enteric neurons by transplanting hair follicle‐derived neural crest stem cells (HF‐NCSCs) to the aganglionic distal colon. We first isolated hair follicle bulges from the vibrissa whisker pad of mice, and cultured the bulges for 3 to 4 days. After HF‐NCSCs migrated onto the culture surface, the bulges were removed, and HF‐NCSCs were cultured in an expansion medium for another week. HF‐NCSCs were found to be able to differentiate into myocytes, neurons and glial cells in different differentiation media. Ten days after they were transplanted to the hindgut explants isolated from E13.5 mouse embryos, the transplanted HF‐NCSCs spread along the gut explant cultured ex vivo and differentiated into neurons. Immunostaining of the serial cross sections indicated most of the transplanted cells were located in the myenteric plexus between two muscle layers. HF‐NCSCs were also transplanted to the descending colon of an adult mouse in vivo after laparotomy. One week later, the cells spread along the colon and differentiate into Tuj1 and PGP9.5 immunoreactive cells. Ten days after transplantation in vivo, subtype inhibitory neurons immunoreactive to nNOS were found among the transplanted cells, and a small number of transplanted cells differentiated into excitatory neurons immunoreactive to ChAT. Additionally, almost all the transplanted cells were found in the myenteric plexus which is also the normal location of endogenous enteric neurons between the longitudinal muscle and circular muscle layers 6 days after transplantation. These results showed that (1) enough HF‐NCSCs were obtained for cell transplantation from a small number of skin hair follicles, and (2) HF‐NCSCs from hair follicles could serve as a cell source for stem cell‐based therapy. This work also gives insights into the feasibility of using autologous cell transplantation with a less invasive hair follicle isolation method on patients with ENS defects in the colon.

Support or Funding Information

The work was supported by a grant from the Research Grants Council of the Hong Kong Special Administrative Region, China (Project No.: 14175017).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Initiation, targeting and sculpting of the phagophore
Authors: Hurley James H.
Published: June 2021

Autophagy is a conserved mechanism that is essential for cell survival in starvation and for cellular homeostasis. The autophagosome and the proteasome comprise the two main pathways for the degradation of proteins. Autophagy proceeds by the engulfment of bulk cytosol and organelles by a cup‐shaped double membrane sheet known as the phagophore, which matures into the autophagosome. Two protein complexes control the initiation of the phagophore: the Atg1/ULK1 kinase complex and the class III phosphatidylinositol 3‐kinase complex I (PI3KC3‐C1). Our laboratory has been applying crystallography, electron microscopy, mass spectrometry, and allied biophysical techniques, in conjunction with cell biology approaches, to understand how these complexes are organized in three dimensional space and time, and how their architectures underpin their regulation and activity.

Support or Funding Information

NIGMS R01 111730 and P01 051487

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Discovery and Engineering of Plant Chemistry for Plant and Human Health
Authors: Sattely Elizabeth
Published: June 2021

Plants are some of the best chemists on the planet and produce an impressive array of small molecules. We are inspired by the fact that humans have become extraordinarily reliant on plant‐derived molecules for food, medicine, and energy. However, remarkably little is known about how plants make these molecules, limiting our ability to engineer and optimize plant metabolic pathways. New plant genome sequences and synthetic biology tools have opened the door to three transformative research areas under investigation in my lab: 1) Identifying the minimum set of enzymes required to make known plant‐derived molecules and non‐natural derivatives through metabolic engineering, and 2) discovering new molecules from plants, and 3) developing new strategies to enhance plant fitness. In this talk, I will describe some of our recent efforts to accelerate the discovery of complete plant pathways for known and novel molecules, not only in the model plant Arabidopsis but also in non‐model plants.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Structure and Signaling Mechanisms of G Protein‐Coupled and b‐Arrestin‐Biased Chemokine Receptors
Authors: Handel Tracy , Gustavsson Martin , Zheng Yi , Stephens Bryan , Baker Genevieve , Ngo Tony , Holden Lauren , Stevens Ray , Cherezov Vadim , Abagyan Ruben , Kufareva Irina
Published: June 2021

Most chemokine receptors are G Protein‐Coupled Receptors (GPCRs), and are best known for their role in controlling cell migration in the context of immune system function. They are also implicated in many diseases particularly inflammatory diseases as well as cancer and HIV, making them important therapeutic targets. Recent structurally and biophysical studies have revealed that the recognition interface between chemokines and receptors is very large and characterized by more complex epitopes than previously believed. In this presentation, our current understanding of how chemokines bind and activate their receptors will be described, as well mechanisms for inhibiting chemokine receptor signaling with orthosteric and allosteric ligands. Common and unique features of G protein‐coupled versus b‐arrestin‐biased receptors will also be described, focusing on CXCR4 (a GPCR) and the b‐arrestin‐biased, atypical chemokine receptor 3 (ACKR3) both of which respond to the chemokine CXCL12.

Support or Funding Information

Funded by NIH grants R01 GM117424 and R01 AI118985

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The PAX‐5 transcript is submitted to 3'UTR editing which regulates its oncogenic expression in cancer cells
Authors: Robichaud Gilles A. , Beauregard Annie‐Pier , Guerrette Roxann , Crapoulet Nicolas , Finn Nicolas , St‐Hilaire Ève
Published: June 2021

The Pax‐5 oncogene is a major factor in various cancer lesions such as hematopoietic cancers. Recently, we have also shown that Pax‐5 regulates breast cancer phenotypic transitioning processes (EMT‐MET) leading to the regulation of cancer cell malignancy. To date, the aberrant expression of Pax‐5 in cancer cells is still unknown. Our team has thus turned its attention to the post‐transcriptional regulation of Pax‐5 expression by important regulatory elements such as miRNAs and alternative polyadenylation (APA). Upon our analysis of the Pax‐5 transcript in Pax‐5 bearing cells (breast epithelia and B lymphocytes), we found that not only is the 3'UTR submitted to APA; but also, alternative splicing of the Pax‐5 mRNA 3'UTR. Interestingly, upon rapid amplification of cDNA ends (3'RACE) from polysomal fractions, we found that Pax‐5 3'UTR shortening correlates with increased translation frequency. These observations were also validated using reporter gene assays where various truncated 3'UTRs cloned downstream of a luciferase gene resulted in differentially translational expression. More importantly, we also found that Pax‐5 transcripts characterized with shorter 3'UTRs were also associated with advanced staging of hematopoietic cancer lesions. Our findings identify novel molecular mechanisms, which account for Pax‐5 aberrant expression and function in cancer cells. These findings will further elucidate Pax‐5‐mediated cancer processes and may provide new avenues for therapeutic intervention.

Support or Funding Information

This work was supported by grants from the New Brunswick (NB) Innovation Foundation, the Canadian Breast Cancer Foundation, the Canadian Breast Cancer Society/QEII Foundation, the NB Health Research Foundation and by the Beatrice Hunter Cancer Research Institute

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Transcriptomic hallmarks of tumor‐stromal interactions in brain metastasis
Authors: Nguyen Don
Published: June 2021

The central nervous system (CNS) is a major site of relapse for several types of treatment refractory cancers including tumors from the lung, skin, and breast. The incidence of brain metastasis associated deaths is rising, because of the limited delivery of drugs across the blood brain barrier (BBB) and morbidity of metastasis once spread in the CNS. One of the most rate limiting steps of CNS metastasis is the ability of disseminated tumor cells (DTC) to outgrow after extravasating across the BBB. Deciphering the molecular interactions which dictate this step in vivo remains a challenge due to the heterogeneous patterns of tumor outgrowth in the CNS and the unique nature of this tumor microenvironment (TME).

To address this challenge, we optimized a novel method for RNA sequencing analysis which leverages xenograft models and overcomes the confounding analytic effects incurred during the isolation of cells ex‐vivo. Using this method, we could decipher molecular alterations that are specific to the tumor cells or the stroma in situ with increased accuracy and sensitivity, revealing striking new insights into brain‐specific transcriptional co‐adaptation of malignant cells and their neuroinflammatory milieu. These gene expression programs are linked to specific patterns of metastatic outgrowth, anatomical location in the brain, and interaction with neuronal cell types. Our study provides a molecular framework to understand the heterogeneous biological manifestation of brain metastasis while revealing novel epigenetic and phenotypic state(s) utilized by metastatic cells to persist and outgrow in the CNS.

Support or Funding Information

This study was funded by grants from the National Cancer Institute (R01CA166376 and R01CA191489).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.