bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evolution of differential 4‐1BB signaling in Human and Murine immune system
Authors: Bitra Aruna , Zajonc Dirk M 
Published: June 2021

4‐1BB (CD137) is an inducible tumor necrosis factor receptor superfamily (TNFRSF) member and is currently being targeted in cancer immunotherapy and autoimmunity. 4‐1BB interacts with its cognate ligand 4‐1BBL to promote T lymphocyte activation and proliferation, but the dissimilarities in mouse and human molecules might result in differential engagement of this pathway. To elucidate the molecular mechanism underlying this distinct behavior, we have determined the crystal structures of human and mouse 4‐1BB/4‐1BBL complexes. Collectively, these structures provide insights into the structural differences that drive the species‐specific receptor‐ligand interactions. The most significant feature is that the TNF homology domain of human (h)4‐1BBL form non‐covalent homo‐trimeric arrangements, whereas the mouse (m)4‐1BBL formed a novel disulfide linked homo‐dimer. In the human 4‐1BB/4‐1BBL complex, each receptor binds at the crevice formed between two protomers of h4‐1BBL resulting in the formation of a “hetero‐hexameric” functional signaling complex (1), whereas each m4‐1BB receptor binds exclusively to a single m4‐1BBL protomer leading to a “tetrameric” signaling unit (2). As multimerization and clustering is a prerequisite for TNFR intracellular signaling, appearance of differential functional signaling units in human and mouse supports a unique mechanism of 4‐1BB signaling in both of these species. Of note, we have discovered that h4‐1BB forms a disulfide‐linked dimer and this covalent receptor dimerization amplifies h4‐1BB signaling by cross‐linking individual ligand‐receptor complexes to form a 2D network. Since the m4‐1BBL dimer can only recruit two monomeric 4‐1BB receptors, cross‐linking of two hetero‐tetrameric signaling units via Galectin‐9 binding to the N‐linked glycans on m‐41BB is required for receptor oligomerization and strong signaling (3).While all characterized TNFR/TNF members exhibit interspecies cross reactivity between human and mouse molecules, 4‐1BB and 4‐1BBL lacks this distinguishing feature. Our structural studies together with comparison of human and mouse 4‐1BB/4‐1BBL complexes highlighted several differences between the ligand and receptor binding interfaces and provide an explanation for the absence of inter species cross‐reactivity between them (2). Furthermore, our biochemical and mutational analysis discloses more variations between mouse and human molecules that could affect signaling and thus suggest the generation of better animal models to study 4‐1BB biology in humans.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Exploring the Stability and Degradation of the Lipin‐1 Protein
Authors: Giebink Faye E , Harris Thurl E 
Published: June 2021

Lipin 1 plays a central role in the regulation of lipid metabolism, which is crucial to maintaining systemic energy homeostasis in the body. Lipin 1 functions as a phosphatidic acid phosphatase to convert phosphatidic acid (PA) to diacylglycerol (DAG) as part of the Kennedy pathway. Phosphorylation of lipin 1 at Ser483 and Ser487 creates a phosphodegron signal that is recognized by βTRCP and promotes lipin 1 ubiquitination and degradation by the proteasome. The polybasic domain (PBD) of lipin 1 is a stretch of nine contiguous Arg and Lys residues within the central domain that are essential for interaction with PA. Mutation of these residues to Ala greatly increases protein expression of lipin 1. We show that this increase in expression is due to an increase in protein half‐life and we suggest this is due to alterations in lipin 1 ubiquitination and proteasomal degradation. During proteomic analysis of lipin 1 we identified at least one Lys within the PBD as acetylated. Acetylated Lys cannot be ubiquitinated. We have previously demonstrated an interaction between lipin 1 and HDAC1 and 3 and lipin 1, and so we treated cells with Trichostatin A (TSA) to prevent the removal of acetyl groups and cause the accumulation of hyperacetylated lipin 1. We find that pre‐treatment of cells with TSA increases lipin 1 protein half‐life. We hypothesize that accumulation of acetylated Lys within the PBD may prevent lipin 1 ubiquitination and degradation. Alternatively, lipin 1 acetylation may affect stability through Lys residues outside of the PBD or through other mechanisms, such as altering phosphorylation. Understanding what regulates lipin 1 stability and its role as a magnesium‐dependent phosphatide phosphatase (PAP) enzyme in triacylglycerol synthesis is important to comprehend mechanisms involved in metabolic disorders that lead to obesity.Support or Funding InformationNIH DK101946 (TEH), UVA Harrison Undergraduate Research Award (FEG)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Platelet bioenergetics and its implications in disease
Authors: Prakhya Kanakanagavalli Shravani , Wang Qingjun , Whiteheart Sidney W 
Published: June 2021

Bioenergetics drives energy production for mechanical processes that are critical to platelet functions, e.g., adhesion, spreading, aggregation, clot formation and contraction. Storage and pathological conditions e.g., stroke, sepsis, diabetes etc., alters platelet energetics and function; however, the specific mechanisms governing these alterations are not clear. It is not well understood which metabolic fuels can be used and which are required. We are investigating the roles of different metabolic pathways in the energy consuming process of clot retraction. Using a spectrum of techniques from in vitro platelet functional assays to biochemical methods, whole animal models, and measurement of bioenergetic functions of mitochondria we are elucidating platelet bioenergetics and its implications in disease. We developed a simple method for measuring clot contraction kinetics using time lapse imaging. Our system is efficient, high throughput and is favorable to study platelet bioenergetics of clot formation and retraction. We hypothesize that, upon activation with thrombin, fuel use can be highly plastic, depending on the fuels available. We found that glycolysis and oxidative phosphorylation can both contribute to provide energy for platelet function. Inhibitors of glycolysis and oxidative phosphorylation using 2‐Deoxyglucose and oligomycin inhibited clot retraction showing that both these processes are important.We have also begun to examine bioenergetics in platelets from diabetics. Platelets from type2 diabetes patients have been shown to have abnormal mitochondrial membrane potential, membrane fluidity, ATP content, and oxidative stress. However, the impact of diabetes on platelet bioenergetics is less clear. We have developed a mouse model to probe the role of mitochondria and oxidative phosphorylation in platelet function. TFAM (Mitochondrial Transcription factor A) regulates mitochondria DNA copy number and transcription of mitochondrial DNA encoded proteins. The regulation of mitochondrial DNA is necessary for mitochondrial biogenesis. We have selectively deleted TFAM in the megakaryocyte lineage using the Platelet factor 4 promoter driven Cre‐Lox system. This model will be very useful, and it supports our in vitro studies with metabolic inhibitors in understanding the role of mitochondria and oxidative phosphorylation in platelet function in platelet bioenergetics contributing to the thrombosisSupport or Funding InformationSupported by HL56652, HL138179 and VA Merit AwardThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Akt1 and Akt2 Isoforms Play Distinct Roles in the Development of Inflammation and Fibrosis during Alcoholic Liver Disease
Authors: Gordillo Karina Reyes , Shah Ruchi , Tuma Pamela L , Robledo Jaime Arellanes , Cheng Ying , Lakshman Raj M 
Published: June 2021

Akt isoforms (Akt1, Akt2, and Akt3), the key phosphorylating kinases, play vital and unique roles in all aspects of alcoholic liver disease (ALD). In vitro, cell specific silencing of Akt isoforms in acetaldehyde+LPS and/or ethanol+LPS treated human hepatic stellate cells (HSC), Kupffer cells and/or VL17A hepatocyte cultures; and in vivo, Akt1 and Akt2 silencing in a chronic ethanol/binge/lipopolysaccharide (EBL) induced mouse liver injury model using pharmacological inhibitors, were used to gain a better understanding of how each Aktisoform differentially regulates cellular functions during ALD. This study involves hepatocyte and Kupffer cells inflammation, and hepatic stellate cells proliferation, migration and fibrogenesis, which cumulatively lead to the pathogenesis of ALD. Herein, our in vitro results showed that siRNA‐directed silencing of Akt2, but not Akt1, significantly suppressed cell inflammatory markers in HSC and Kupffer cells. While Akt1 and Akt2 inhibited cell proliferation in HSC; Akt2 alone inhibited cell migration in HSC; and Akt1 and Akt2, but not Akt3 inhibited the fibrogenic markers in hepatocytes and HSC. Moreover, although Akt isoforms show extensive sequence similarity, they are localized in different subcellular compartments in HSC. On the other hand, our in vivo results showed that EBL treatment in wild‐type C57BL/6 mice stimulated all three isoforms of Akt with concomitant increases in phosphorylated species of phosphoinositide dependent kinase‐1, mammalian target‐o‐rapamycin complex 2, and phosphatidylinositol‐3‐kinase, resulting in increases in inflammatory, proliferative, and fibrogenic genes. In addition, pharmacological blocking of Akt2, but not Akt1, inhibited EBL‐induced inflammation; and of Akt1 and Akt2 inhibited the pro‐fibrogenic markers as well as fibrosis. Our finding suggest that Akt isoforms play unique roles against inflammation, cell proliferation, migration, and fibrogenesis during EBL‐induced liver injury. These in vitro and in vivo results also suggest that close attention must be paid when targeting all Akt isoforms as a therapeutic intervention.Support or Funding InformationNIAAA/NIH K01 award ‐ number: K01AA025140‐02This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In Vivo Knockdown of the IP3R1 Calcium Channel Disrupts ER and Actin Morphology, as well as TBC Bulb Maturation in the Apical Processes of Sertoli Cells in Rat Testes
Authors: Adams Arlo , Vogl Wayne 
Published: June 2021

Spermatogenesis is the process by which sperm cells are produced, but at one phase of this process, late stage spermatids are temporarily attached to the apex of Sertoli cells in the seminiferous epithelium. Late spermatids are anchored by large adhesion like junction complexes termed ‘ectoplasmic specializations’ (ESs) that are proposed to be disassembled by novel endocytic structures only found in mammalian Sertoli cells, called ‘tubulobulbar complexes’ (TBCs). Both ESs and TBCs are heavily actin related, with parallel actin arrays and dendritic actin networks respectively being hallmarks of their identities. The endoplasmic reticulum (ER) in the Sertoli cell apical processes surrounding the late spermatids forms an extensive and continuous network. Cisternae of this network form part of the structure of the ESs, participate in a membrane contact site with the bulbs of TBCs, and have a parallel alignment with the surrounding Sertoli cell plasma membrane. Previously, we've localized the IP3R calcium channel to the TBC bulb‐ER contacts as well as other calcium exchange machinery to various parts of the ER in the apical process. Calcium release and exchange have been implicated in processes related to both actin dynamics as well as in fusion and acidification of endocytic components such as endosomes. We've proposed that calcium exchange may be a crucial mechanism in regulating both the actin networks of ESs and TBCs, as well as in facilitating the maturation and fusion of TBC bulbs as they develop into putative endosomes. If calcium exchange from the ER is responsible for regulating the local actin networks and maturation of endocytic components in the apical process, then knocking down the primary calcium release channel of the ER should alter local actin and TBC bulb morphology. To test this, we injected the testes of Sprague Dawley rats with siRNAs against IP3R1 (ITPR1) and collected tissues at 1 day, 2 day, and 3 days post injection and processed the tissues for either western blot, immunofluorescence, or electron microscopy. ITPR1 siRNA treated tissues showed significant morphological alterations to the ER, actin, and TBC bulbs in the apical processes of late spermatids compared to controls. This data supports the hypothesis that ER calcium exchange is an integral component to actin dynamics and TBC bulb maturation in the apical processes of Sertoli cells.Support or Funding InformationNatural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grant awarded to AWV; Alexander Graham Bell Scholarship (NSERC) awarded to AAThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
α‐Adducin is crucial for endothelial Tight Junction Integrity
Authors: Ponce Alexander García , Radeva Mariya Yosifova , Waschke Jens 
Published: June 2021

Adducin (Add) is a membrane and cytoskeleton‐associated protein participating in the assembly of spectrin‐actin networks, which in turn regulate adherens junction (AJ) and tight junction (TJ) integrity. This process is important for endothelial barrier homeostasis. However, the mechanisms by which Add participates in endothelial barrier regulation are not well understood. Therefore, we generated immortalized Add knock‐out (KO) endothelial cell lines and investigated barrier dynamics. In absence of Add, transendothelial electrical resistance (TER) measurements showed impaired barrier function under basal conditions. Moreover, cAMP‐mediated barrier stabilization induced by treatment with Forskolin (F) and Rolipram (R) was compromised. Additionally, cells lacking Add showed higher intracellular cAMP concentration although the cAMP‐degrading enzyme phosphodiesterase 4D was overexpressed. Strikingly, ablation of Add led to a complete absence of TJ protein claudin‐5. Furthermore, in contrast to WT cells, Add KO cells displayed reduced membrane staining of other TJ molecule such as ZO‐1, both under resting conditions and after F/R treatment. However, the cellular distribution of this protein remained similar. In addition, localization of AJ protein VE‐cadherin at cell‐cell contacts was altered in Add KO cells, which was paralleled by significantly reduced protein levels. Taken together, in this study we show that Add is involved in endothelial barrier dynamics by regulating AJ and TJ integrity as well as cAMP‐mediated‐signallingThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Coq2p function in the Saccharomyces cerevisiae coenzyme Q biosynthetic pathway
Authors: Marbois Beth Noelle , Xie Letian , Shi Yuchen , Hyman Kyle , Clarke Catherine F. 
Published: June 2021

Respiratory electron transport in the plasma membrane of prokaryotes and in the inner mitochondrial membrane of eukaryotes is supported by the crucial lipid component, ubiquinone (coenzyme Q or Q). Here we further characterize the Coq2 polypeptide, a key resident polypeptide of a complex of Coq polypeptides that is necessary to manufacture Q. One mutant allele of Coq2p and a hemaglutanin tagged Coq2p are described and compared to the wild‐type polypeptide in S. cerevisiae via Western blotting, growth curves and lipid analyses. Although stable levels of Coq2 polypeptide are evident in mitochondria isolated from the coq2‐1 mutant, the strain is incapable of aerobic respiration. We also present biochemical evidence that Coq2p is expressed in two forms, one at 30 kDa and another at 41kDa. Growth on non‐fermentable carbon sources invokes the 41 kDa isoform. We speculate that the 41kDa form is necessary for Q synthesis since it is lacking in the coq2‐1 mutant, and not highly expressed in cell extracts of cells grown in media containing fermentable sugars. We describe the coenzyme Q intermediates present in lipid extracts of the coq2‐1mutant. Our data demonstrate that Coq2p is essential for Q synthesis.This work was supported by NIH grants GM 45952 and NIA AG 19777.
Dendritic cell differentiation in the presence of PGE2 primes dendritic cells for a proinflammatory response, induces IL‐23, and promotes Th17 differentiation
Authors: Khayrullina Tanzilya R , Yen Jui‐Hung , Ganea Doina 
Published: June 2021

Dendritic cells (DC) play an important role in the induction of the immune responses. The DC phenotype and function is modified by different factors. PGE2 plays an important role in inflammation and autoimmunity through changes in the proinflammatory cytokine profile. The proinflammatory cytokine IL‐23 is associated with autoimmune disorders. In the periphery, macrophages and DC are major producers of IL‐23, whereas microglia play a similar role in the CNS. IL12 and IL‐23 share the p40 subunit, but have specific p35 (IL‐12) and p19 (IL‐23) subunits. The main goal of this study was to define the role of PGE2 in DC differentiation and production of bioactive IL‐23. DC were generated in GM‐CSF (DC) or GM‐CSF+PGE2 (DCp) and characterized following stimulation with LPS. PGE2 functions as an enhancer of DC differentiation and influence their Th‐cells lineage driving ability, by providing factors required for Th17 differentiation and survival. Activated DCp have higher levels of proinflammatory cytokines, such as IL‐6, IL‐1β and TNF, as a result of higher levels of TLR expression and of NFkBp65 phosphorylation and nuclear translocation. In addition, PGE2 shifts the IL‐23/IL‐12 balance in favor of IL‐23 in activated DC. These observations may help to understand the distinct role of PGE2 in the induction of pathogenic Th17‐cells.This work is support by NIH/NIAID R01AI052306
Lymphoma cell surface expression of CD200 modulates anti‐tumor immunity
Authors: Wong Karrie Ka Wai , Gorczynski Reg , Spaner David 
Published: June 2021

CD200 is a transmembrane protein broadly expressed on a variety of cell types and delivers immunoregulatory signals through binding to receptor (CD200Rs) expressed on monocyte/myeloid cells and T lymphocytes.We showed in earlier reports that infusion of a soluble form of CD200, CD200Fc, into EL4 thymoma‐bearing C57/B6 mice enhanced tumor progression (Clin. Exp. Immunol. 2001). More recently, independent groups have reported CD200 overexpression associated with multiple myeloma, AML, and CLL.In this study we investigated the in vitro effect of blockade of CD200 expression, using both anti‐CD200 mAbs and CD200‐specific siRNAs, on CTL induction in human PBL using a CD200+ lymphoma cell line (ly5). We found that blockade of CD200 on ly5 cells significantly enhanced anti‐tumor CTL responses in vitro. Production of the inflammatory cytokines TNFa and IFNg were similarly enhanced. Cell depletion studies supported an important role for antigen presenting cells in CD200 regulation of anti‐tumor responses. We also developed a CD200 ELISA assay to measure serum CD200 levels in healthy donor controls and cancer patients, and in preliminary analyses found elevated levels of sCD200 in the plasma of patients with CLL.Our data provide novel insights into the mechanisms of CD200 mediated suppression of tumor immunity, the understanding of which may open new avenues to development of different cancer therapies.
Food group intake patterns differ between low‐income Puerto Ricans and Non‐Puerto Rican pregnant Latinas
Authors: Bermúdez‐Millán Angela , Hromi‐Fiedler Amber , Damio Grace , Segura‐Pérez Sofia , Pérez‐Escamilla Rafael 
Published: June 2021

Latinos are the largest ethnic minority group in the United States. They are a heterogeneous group, represented by people from Mexico, Central and South America, and the Caribbean Islands. A longitudinal study (n=234) was conducted to examine differences in food group intake between low income Puerto Ricans and Non‐Puerto Rican pregnant Latinas. A single 24‐hour dietary recall was collected to assess food group intake. At baseline 94% of the participants were in their second to third pregnancy trimester. Mean age was 24 ± 5y. Puerto Ricans comprised 69% of the sample. Mean length stay in the US was 15 ± 9y for Puerto Ricans and 5 ± 5y for Non‐Puerto Ricans. Puerto Ricans were more likely to be bilingual, have a larger household composition, participate in WIC and the food stamp program and less likely to have a partner. Non‐Puerto Ricans consumed significantly (p<0.05) more servings of dark green‐leafy vegetables, tomatoes, other vegetables, fruits, beans, tortillas and candy/sugars. Puerto Ricans consumed more servings of breakfast cereals, crackers, processed meat, cheese, dairy desserts, soft drinks and sweetened fruit drinks. Acculturation may be playing an important role as Non‐Puerto Ricans had been for a shorter period of time in the US than their Puerto Rican counterparts.Funded by the Connecticut NIH EXPORT Center of Excellence for Eliminating Health Disparities among Latinos (NIH‐NCMHD Grant # P20MD001765).
Comparison of BCFA types in vernix and meconium of healthy neonates
Authors: Ran‐Ressler Rinat , Devapatla Srisatish , Lawrence Peter , Brenna J. Thomas 
Published: June 2021

Background. Branched Chain Fatty Acids (BCFA) are rare in mammalian lipids but are known to be synthesized in the sebaceous gland. They are richest in vernix caseosa, the substance that covers the fetus in late gestation. During pregnancy, the fetus swallows amniotic fluid and with it sloughed vernix particles. We recently reported the presence of BCFA in meconium, the first pass of the neonate. Since vernix and meconium are the input and output, respectively, of the GI tract, we hypothesized that the BCFA profile of the two would be related in some systematic way.Methods. Vernix was skimmed off newborns in the delivery room and meconium was collected into diapers and saved for analysis. Samples were processed by a modified Bligh & Dyer extraction, and FAME were analyzed by GC and GC/MS. The BCFA profiles for vernix and meconium was compared for individual neonates and on average.Results. Vernix BCFA contain iso‐BCFA, anteiso‐BCFA, dimethyl BCFA and middle‐chain monomethyl BCFA and range from 12 to 26 carbons. In contrast, meconium BCFA are mainly iso‐BCFA and range from 16 to 26 carbons. Major differences in profiles were found from infant to infant.Conclusions. BCFA are a normal component of the term newborn gut throughout its entire length. BCFA with <16 carbons and with branching other than iso disappear in the sterile fetal gut in late gestation. The mechanism is unknown. Funding: NIH GM071534.