bluuebunny/crossref_metadata_2025_split huggingface dataset:
A DNA Combinatorial Library of 258 Strand Combinations for DNA Computing
Authors: Waters Colin , Macula Anthony , Pogozelski Wendy 
Published: June 2021

DNA‐based computers use combinatorial libraries of DNA strands to store information while manipulating these strands to solve computationally hard problems in parallel. In order for DNA computing to be successful, these libraries must be constructed in an easy and reliable manner.We show the construction of two libraries of 2exp3 and 2exp5 strand combinations and then we link them to produce a library of 2exp8 or 256 combinations of 150‐bp DNA strands. We then verify that the expected strands are present. This library should be useful in solving mathematical satisfiability (SAT) problems.
Ubiquitination of the polyglutamine disease protein ataxin‐3 enhances its activity as a deubiquitinating enzyme
Authors: Todi Sokol V , Winborn Brett J , Williams Aislinn J , Paulson Henry L 
Published: June 2021

Protein ubiquitination regulates a broad spectrum of cellular functions. Diverse ubiquitin ligases and deubiquitinating enzymes (DUBs) control the ubiquitination state of, and types of ubiquitin chains attached to, various proteins. Ataxin‐3 (AT3) is a DUB with a polyglutamine tract which, when expanded, causes the neurodegenerative disorder Spinocerebellar Ataxia Type 3. Disease pathogenesis in expanded polyglutamine disorders is greatly influenced by the disease protein context and function, therefore understanding normal properties of AT3 is important. Previous data from our laboratory and others revealed that AT3 functions in protein quality control. Here we provide evidence that AT3 ubiquitination enhances its DUB activity. In cells, endogenous AT3 ubiquitination is increased by the presence of excess ubiquitin chains, suggesting that AT3 ubiquitination is linked to cellular ubiquitin levels. Ubiquitinated AT3 (AT3‐ub) cleaves lysine(K)‐63 hexaubiquitin chains more efficiently than does unmodified AT3. Similar to unmodified AT3, AT3‐ub preferentially cleaves K‐63 linked ubiquitin over K‐48 chains, and longer over shorter chains. Collectively our findings suggest that ubiquitination of AT3 and the corresponding enhancement of its DUB activity are important to cells in which protein homeostasis is impaired.This work was supported by: NRSA NS056609 (AJW), R01 NS038712 (HLP).
The Salmonella effector SopB affects phospholipid content of Salmonella‐induced membrane ruffles
Authors: Steele‐Mortimer Olivia 
Published: June 2021

Salmonella enterica serovars are pathogenic Gram‐negative bacteria that can cause disease in a variety of eukaryotic hosts. Central to pathogenesis is the ability to invade non‐phagocytic cells within the intestinal epithelium, a process that involves massive actin rearrangements and membrane ruffle formation on the plasma membrane. This is absolutely dependent on a type III secretion system (T3SS), which translocates a number of bacterial effector proteins directly into the host cell. One of these effectors, SopB/SigD, is an inositol phosphatase that is absolutely required for activation of Akt/PKB in infected epithelial cells. We have previously shown that the SopB/Akt pathway can protect infected cells against the onset of apoptosis. Here we have further delineated the mechanism of Akt activation in Salmonella‐infected cells. Unlike the canonical Akt pathway this process is Wortmannin‐insensitive and is prolonged, being detectable for at least 8 hrs. Using PH‐domain GFP fusions we have performed semi‐quantitative analysis of the phospholipids present in Salmonella induced ruffles and demonstrate a role for SopB in ruffle formation. This activity is absolutely dependent on the phosphatase activity of SopB since a catalytically inactive SopB mutant is unable to affect the phospholipid content of Salmonella‐induced ruffles.ConclusionsThe T3SS effector protein SopB modulates the phospholipid content of Salmonella‐induced membrane ruffles.
Compartment‐dependent modulation of Ras ubiquitination
Authors: Terrillon Sonia , Bar‐Sagi Dafna 
Published: June 2021

Ras GTPases are key regulators of cell growth, differentiation and transformation. Ubiquitination of Ras modulates its intracellular trafficking and signaling output. Yet, detailed knowledge about the in vivo dynamics of this process is limited. Here, we took advantage of the Bioluminescence Resonance Energy Transfer (BRET) technology to monitor HRas ubiquitination in living cells and to investigate its relationship to membrane compartmentalization. We demonstrate that HRas ubiquitination is dynamically altered upon growth factor stimulation. The growth factor‐dependent modulation of HRas ubiquitination is temporally coupled to changes in HRas microdomain localization as determined by BRET analysis of HRas/Caveolin1 interactions. Significantly, an HRas chimera that is unable to interact with Caveolin1 is refractory to ubiquitination, indicating that HRas ubiquitination requires an interaction with Caveolin1. Consistent with this observation, disruption of lipid microdomains by membrane cholesterol depletion inhibits HRas ubiquitination. Furthermore, Caveolin1 expression levels are tightly correlated with HRas ubiquitination. Taken together, our data demonstrate that HRas ubiquitination is dynamically regulated and requires the integrity of caveolae and their scaffolding protein Caveolin1.Supported by Human Frontier Science Program (S.T.) and NIH grant CA055360 (D.B.‐S.)
Novel probes of acetylcholinesterase inactivation
Authors: Guo Lilu , Suarez Alirica I. , George Katie , Thompson Charles M. 
Published: June 2021

Organophosphates (OPs) react with acetylcholinesterases (AChEs) to form a covalent bond at a specific serine residue in the active gorge, thereby providing a highly precise modification of this enzyme and offering unique explorations of protein structure, function and mechanism. The long‐range goal of this study is to show how covalent modification of AChEs by chromophore‐linked OPs can be used to probe the protein and its local environment. Click chemistry was used as a key transformation method to prepare an array of chromophores linked to a reactive fluorophosphonate (FP) head group. Successfully synthesized chromophore‐FPs varying in length and chromophore structure were docked in FlexX (Sybyl 7.0) to visualize and calculate positioning of the chromophore‐FPs relative to the protein active site/gorge/peripheral site. The inhibition rates of chromophore‐FPs against rMoAChE and EEAChE were determined via colorimetric assay. In addition, the binding effects of chromophore and FP part to protein were evaluated by measuring the inhibition rates of chromophores and FP parts, respectively. Results demonstrated that the chromophore structure and the ligand length mutually affect the inhibition potency. To differentiate AChE stereoselectivity, asymmetric analogs of the chromophore‐linked phosphonates will be synthesized and tested as anti‐AChEs.Supported by NIEHS UO1 ES016102 and NCRR COBRE RR15583
Three Tropical Plants with Hepatoprotective properties
Authors: Iroanya Onyekachi Ogbonnaya , Okpuzor Joy Elizabeth , Mbagwu Herbert O 
Published: June 2021

The use of medicinal plants is becoming increasingly popular among all social classes in our society. The lower socioeconomic class cannot afford the high cost of orthodox treatment; the only alternative to remain in good health is the use of medicinal plants which abounds in their surrounding to treat all types of diseases. The study aims to substantiate the claims made by the traditional healers on the use of a mixture of Gongronema latifolia, Ocimum gratissimum and Vernonia amygdalina in the treatment of liver problems.The mixture showed no toxicity when administered orally or intraperitoneally to Wistar albino rats. Aqueous extract of the mixture was evaluated against Acetaminophen induced acute hepatotoxicity in Wistar rats. The animals were divided into six groups with each group containing ten animals. Three doses of the extract (2000, 4000, 8000 mg\kg) were fed to the rats for four days at 24 hours interval.At a dose of 8000 mg\kg body weight there was a significant change in the level of Alkaline Phosphatase (21.3± 3.8), Aspartate Aminotransferase (23.3±2.3) and Alanine Aminotransferase (18 ±1.1) when compared to positive (19.7 ± 0.3, 23.7 ± 1.2 and 23.7 ± 1.2) and normal (25 ± 2.1, 22.3 ± 1.9 and 23.7 ± 1.2) control values. These results indicate a significant hepatoprotective effect against liver damage induced by Acetaminophen.We therefore, provide a scientific rationale for the traditional use of the mixture in the management of liver diseases.
Modulation of Dendritic cell response by human CD8 T cells in the presence of a microbial stimulus
Authors: Chong Shu Zhen 
Published: June 2021

Dendritic cells (DCs) play a central role in the induction of immune responses by priming CD8 T cells for effector and cytotoxic functions. However, little is still known about the priming effects of CD8 T cells on DCs in the event of a microbial infection. Here, we show that human CD8 T cells are capable of modulating DC cytokine production early during the priming process. We showed that activated CD8 T cells were consistently better in activating DCs for cytokine production. Activated CD8 T cells enhanced the cytokine production of IL‐12p40, TNF‐α and GM‐CSF by DCs in the presence of a microbial stimulus. Notably, IL‐1 and IL‐12p70 required the absolute presence of both CD8 T cells and LPS. Interestingly, IL‐1 is produced by DCs with CD8 T cells but not with IFN‐g, indicating the involvement of other signals. CD8 T cells were also able to prime DCs for the production of chemokines such as RANTES and IP‐10. This study demonstrates the helper role of CD8 T cells in the enhancement of DC activity and its possible implications in immune modulation.This work is funded by the Immunology Programme (Grant no. C182‐002‐002‐001). Chong SZ is supported by the NUS Graduate School for Integrative Science and Engineering Scholarship
The evolutionary conserved HoxC4 homeodomain protein induces AID expression and regulates immunoglobulin class switch DNA recombination and somatic hypermutation
Authors: Park Seok‐Rae , Pal Zsuzsanna , Al‐Qahtani Ahmed , Mai Thach , Xu Zhenming , Zan Hong , Casali Paolo 
Published: June 2021

The regulation of activation induced cytidine deaminase (AID) gene (aicda) expression and the modulation of immunoglobulin gene class switch DNA recombination (CSR) and somatic hypermutation (SHM) remain to be defined. We found that the HoxC4 homeodomain protein is preferentially expressed in germinal center B cells and upregulated by stimuli that induce AID expression and CSR. In HoxC4‐deficient mice, the mutation frequency in the intronic JH‐iEμ region of IgH DNA and the serum IgG1 levels were decreased by about 60%. Accordingly, the high affinity anti‐NP response was significantly compromised in hoxC4−/− mice, and CSR to IgG1, IgG2a, IgG2b, IgG3 and IgA in in vitro stimulated hoxC4−/− B cells was impaired. This was not due to altered hoxC4−/− B cell proliferation or germinal center formation in hoxC4−/− mice. Rather, it resulted from a defective induction of AID expression. Indeed, by performing in vitro promoter bashing experiments using luciferase gene reporter assays and ChIP, we showed that HoxC4 activates aicda by binding to an evolutionarily conserved HoxC4/Oct‐binding ATTT(GCAT) motif in the promoter for this gene. Enforced expression of AID in HoxC4‐deficient B cells restored CSR. Thus, these findings show that by regulating AID expression through direct activation of the aicda promoter, HoxC4 plays a critical role in modulating CSR and SHM.Supported by NIH grants, AI 45011, AI 60573 and AR 40908.
Hsp90 associates with N‐terminal extended peptides and is required for direct antigen presentation
Authors: Garg Manish , Callahan Margaret K , Srivastava Pramod K 
Published: June 2021

Antigen presentation by MHC I molecules requires the generation of peptides in the cytosol, ferrying of peptides from cytosol to endoplasmic reticulum, loading of MHC I with peptides, and transport of MHC I‐peptide complexes to the cell surface. Hsp90 is a cytosolic chaperone known to associate with peptide and peptide precursors of MHC I epitopes. We report here that treatment of a variety of cells with hsp90 inhibitors leads to generation of “empty” MHC I molecules on the cell surface, due to inhibited loading of MHC I with peptides. Inhibition of hsp90 does not inhibit protein synthesis in general, nor synthesis of MHC I in particular, nor does it affect the activity of proteasomes. Hsp90‐inhibited cells, like lactacystin‐treated cells, are poor stimulators of T lymphocytes. The role of hsp90 in presentation of an ovalbumin epitope is shown to be at a post‐proteasomal step: hsp90 associates with N‐terminally extended precursors of the SIINFEKL epitope, and such peptides are depleted from hsp90 preparations in inhibitor‐treated cells. These results argue for a substantial role for hsp90 in chaperoning of antigenic peptides in direct presentation.This work was supported by NIH grant CA 084479 to P.K.S.
Maturation of bone marrow‐derived dendritic cells stimulated with collagen is induced via DDR2‐ mediated NF‐kB activation
Authors: Lee Ji‐Eun , Lee Sun‐Hee , Lee Young‐Mi , Kim Bum‐Tae , Moon Woo‐Sung , Kim Dae‐Ki 
Published: June 2021

Dendritic cells (DC) are critical immune sentinels with the unique ability to initiate both innate and adaptive immunity. The ability of DC to activate naïve T cells is tightly linked to their maturation state. The mechanism by which a collagen receptor discoidin domain receptor‐2 (DDR2) controls the maturation of DC remains undefined. In this study, we investigated a signaling mechanism of DDR2, which is one of collagen receptors, to induce the functional maturation of murine bone marrow‐derived DC. The expression of surface molecules, including MHC class II and CD86, on DC were upregulated by DDR2 engagement to collagen, in comparison with unstimulated DC. DDR2 activation significantly increases the production of IL‐12 by DC, as well as proliferation of allogeneic T cells. Moreover, IL‐12 production of DC by collagen was increased through NF‐kB activation to induce and suppressed by pretreatment of NF‐kB and PKC inhibitors. Also, the pretreatment of DC with inhibitors of NF‐k B and JNK signal pathways inhibited collagen‐induced up‐regulation of surface molecules, such as MHC class II and CD86. Our results demonstrated that collagen stimulation could induce the functional maturation of DC via DDR2 mediated‐NF‐kB, PKC and JNK signal pathways.Supported by KSEF Basic Research Program [R01‐2005‐000‐10942‐0]
Neutrophil adhesion under flow conditions induces zyxin nuclear translocation in endothelial cells
Authors: Bowley Erin Nicole , Patel Kamala , Colarusso Pina 
Published: June 2021

High shear stress causes zyxin translocation from focal adhesions (FA) to stress fibers. We previously showed that leukocyte adhesion to stimulated endothelial cells (EC) activated mechanosensitive signaling pathways in ECs at low shear stress, which in turn modulated leukocyte transmigration. In this study, we examined the remodeling of the FA protein zyxin following neutrophil (PMN) adhesion under shear stress and determined if downregulating this protein altered PMN transmigration. Zyxin localized to FAs and stress fibers under static conditions. Buffer perfusion across TNFα‐stimulated ECs did not change zyxin localization at lower shear stresses, but resulted in modest nuclear translocation of zyxin at a shear stress of 4 dyne/cm2. In contrast, PMN recruitment to TNFα‐stimulated ECs resulted in a loss of zyxin from FAs and translocation of zyxin to the nucleus at shear stresses as low as 1 dyne/cm2. Translocation specifically occured in ECs with adherent PMNs and did not occur in ECs distant from regions containing adherent PMNs. Perfusion of fixed PMNs, which roll but to do not adhere or transmigrate, did not induce changes in zyxin localization. Finally, zyxin downregulation in ECs attenuated PMN transmigration. These data suggest that PMN trafficking across stimulated ECs leads to focal adhesion remodeling and that EC zyxin nuclear translocation during this process may play a potential role in gene transcription.Funding: Canadian Institutes of Health Research, Alberta Heritage Foundation for Medical Research.
Endothelial cell paxillin is remodeled during leukocyte adhesion and is essential for transmigration
Authors: Roccamatisi Dawn Louise , Zhang Hong , Colarusso Pina , Patel Kamala 
Published: June 2021

The focal adhesion protein paxillin plays a critical role in focal adhesion remodeling and signaling and as such was examined within endothelial cells (EC) during in vitro neutrophil transmigration. We perfused isolated human neutrophils across TNF‐treated EC in a parallel plate flow chamber and investigated paxillin remodeling by immunostaining. Immunofluorescence data showed that there was a localized loss in EC paxillin staining following neutrophil adhesion under flow conditions. Imaging exogenous paxillin‐YFP in EC, we observed endothelial paxillin loss and remodeling in real‐time. In flow experiments in which we perfused fixed neutrophils or a neutrophilic cell line, both of which adhere but do not transmigrate, we found that paxillin loss was transmigration‐dependant. A functional role for EC paxillin was examined by utilizing siRNA. Using flow cytometry and western blotting, we showed more than ninety percent transfection efficiency and down regulation of paxillin. Downregulation of EC paxillin had no effect on the total number of neutrophils recruited to TNF‐stimulated EC; however, transmigration was blocked by more than 40%. This was not due to changes in expression of E‐selectin, IL‐8, or ICAM‐1 in paxillin downregulated EC. These data suggest that focal adhesion remodeling occurs during leukocyte recruitment and is essential for maximal transmigration.Research funded by The Canadian Institutes of Health Research.
Short‐term depletion of regulatory T cells converts self‐limiting arthritis into a chronic disease
Authors: Kamradt Thomas , Bruns Lisa , Frey Oliver , Reichel Andreas , Morawietz Lars 
Published: June 2021

Background: we have established a new arthritis model, induced by immunization of normal mice with the glycolytic enzyme glucose‐6‐phosphate isomerase (G6PI) (Schubert et al. J. Immunol 2004;172:4503). Arthritis occurs in > 90% of the animals from d9 after the immunization and is usually self‐limiting within 30–40 days. We examined if regulatory T cells (Treg) were involved in the resolution of arthritis.Results: Injection of anti‐CD25 antibody on d‐11 and –8 before immunization resulted in a massive yet transient reduction of CD25+FoxP3+ Treg. The percentage of Treg was normal at d12 after immunization i.e. early during the course of arthritis. This transient depletion affected the disease course profoundly, resulting in chronic‐destructive arthritis with histologically active inflammation over the whole follow‐up period (80d). Exacerbated arthritis was accompanied by increased T‐cell proliferation and increased numbers of IL‐17 producing T cells.Conclusions: CD25+FoxP3+ Treg may not be effective in the resolution of ongoing arthritis. Rather, they seem to act in the induction phase of the disease by controlling the extent of T cell activation and differentiation.
Role of PKA‐activating Gs‐coupled GPCR ligands in the regulation of SLE T cell responses
Authors: Khan Islam U. , Das Maitreyee , Hudgens Joseph M. , Loza Matthew J. 
Published: June 2021

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by generalized T cell dysfunction. A subpopulation of SLE subjects demonstrate a T cell‐specific defect in type I PKA (PKA‐I). We hypothesize that T‐cell specific defects in PKA‐I contribute to reduced inhibition of T cell responses mediated by Treg cells and GS‐coupled GPCR ligands. Tregs (CD4+ CD25+hi) and responder T cells (CD4+ CD25−) were purified by cell sorting. The responder T cells were stimulated with CD3+CD28 mAb alone or with added Tregs, without or with added XAC (pan‐adenosine receptor antagonist) or indomethacin (pan‐cyclooxygenase inhibitor). Addition of Tregs suppressed proliferation of responder cells, with the suppression significantly greater in control subjects than in SLE subjects with a PKA‐I defect. Blocking adenosine receptors with XAC and inhibition of prostaglandin production with indomethacin significantly reversed suppression of responder cell proliferation for control but not SLE. These results demonstrate the role of GS‐coupled GPCR ligands in Treg‐mediated suppression, and provide a mechanism by which Tregs from SLE have reduced capacity to inhibit T cell responses.Supported by NIH RO1‐AR39501.
Blockade of CD70 Exacerbates Experimental Autoimmune Myocarditis by Suppressing Regulatory T cells
Authors: Li Haiyan S. , Ligons Davinna L. , Guler Mehmet L. , Talor Monica V. , Cihakova Daniela , Yagita Hideo , Rose Noel R. 
Published: June 2021

Our previous studies of genes determining susceptibility to experimental autoimmune myocarditis (EAM) revealed Eam2 locus on chr.6. In searching for differentially regulated transcripts during EAM induction in susceptible A.SW vs resistant B10.S mice by microarray analysis, CD27 was identified in Eam2. CD70/CD27 are cell surface molecules belonging to the TNF/TNF‐receptor superfamilies and provide costimulatory signals for T cell activation and survival. In this study, administration of CD70‐blocking antibody FR70 exacerbated EAM incidence, severity and mortality in A.SW but not B10.S mice, accompanied with enhanced T‐ and B‐ cell responses to cardiac myosin. We observed a significant increase in IL‐17 in the spleens of FR70‐treated group, whereas levels of IL‐2, IL‐4, IL‐10 TNFα and IFNγ were reduced. Surface staining of CD27/CD70 showed that CD27 was constitutively expressed on naïve T cells and was downregulated after activation, whereas CD70 was largely restricted to a regulatory T cell (Treg) subset before activation and was rarely upregulated in activated T, B and dendritic cells. Addition of FR70 to in vitro cultures of cardiac myosin‐ primed lymphocytes enhanced their proliferation and IL‐17 production. These results show that blockade of CD70 impairs Treg functions, thus promoting EAM. Our findings suggest a novel molecular mechanism by which Treg may exert suppressive functions.Supported by NIH Grants R01 HL‐077611 and R01 HL‐067290
Anti‐inflammatory properties of cannabinoid receptor‐2‐selective agonists in experimental autoimmune encephalomyelitis
Authors: Kong Weimin , Adhikary Sabina , Zhang Ming , Tuma Ronald , Ganea Doina 
Published: June 2021

Previous studies showed that cannabinoids have anti‐inflammatory effects in experimental autoimmune encephalomyelitis (EAE), but the mechanism is elusive. Cannabinoids act through two cloned receptors, CB1 and CB2. The CB1 receptor is expressed primarily in the CNS, whereas the CB2 receptor is highly expressed in immune cells in the periphery. Previous results from our laboratories indicated that the CB2 agonist O‐1966 has both a protective and a therapeutic effect in models of chronic EAE and of remitting‐relapsing EAE. Here we investigated the effect of the CB2 agonist O‐1966 on dendritic cells and T cells. We report that the CB2 agonist inhibits antigen‐induced T cell proliferation in co‐cultures with syngeneic dendritic cells. The CB2 agonist decreased production of dendritic cell‐derived inflammatory cytokines, i.e., IL‐12p70, IL‐6 and IL‐23, and increased production of the anti‐inflammatory cytokine IL‐10. The CB2 agonist also inhibited IFNγ production by T cells. We also found that the CB2 agonist induced Foxp3 expression in T cells, as determined by quantitative RT‐PCR. We conclude that the anti‐inflammatory role of CB2 agonists could be mediated through effects on dendritic cells and subsequent T cell differentiation, and through the induction of regulatory T cells.This work is supported by NIH/NIAID R01AI052306.