bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isolation and Characterization of the TATA binding protein (TBP) of Taenia solium
Authors: Rodríguez‐Lima Oscar , Jiménez Lucía , Zarain‐Herzberg Angel , Landa Abraham 
Published: June 2021

Taeniosis/cysticercosis are diseases caused by Taenia solium and mainly affects developing countries. Little it is known about the transcriptional system in cestodes, therefore we want study and found the differences with the human and pig (host) transcription system.The aim of this study was the identification of the elements present in the core promoter that regulates the formation of the pre initiation complex (PIC). We analyze the core promoter of different genes and identify (in silico) the cis‐elements presents. Moreover, we have cloned and characterized a TBP in T. solium. TBP is an important transcription factor to form the PIC. Besides, we identified the gene that encodes this TBP. The gen shows the presence of 4 introns in accordance with Chambon's rules and the recognition sites for splicing machinery. The amino acid sequence translated from cDNA reveal a 238‐residues protein with an expected weight of 26.7 kDa. Southern and northern blot analysis suggests the presence of one gene and two mRNA for TBP in T. solium. EMSA and supershift show a specific interaction between the TATA box of different promoters and nuclear extracts from T. solium. Our results indicate that we have isolated the gen encode a TBP and showed it interaction with TATA box from different genes.This work was supported by grant IN219711 and CONACYT 80134 to A.L., and IN204410 to A.Z.‐H.; O.R.‐L. is supported by a scholarship from CONACyT 240037.
Comparative analysis of transcriptional regulation by Ascl1 and Neurog2
Authors: Uhler Michael , Redmond Tanya , Huang Holly , Thompson Robert , Turner David 
Published: June 2021

The proneural basic‐helix‐loop‐helix (bHLH) transcription factors Ascl1 and Neurog2 direct the differentiation of specific populations of neurons by regulating the transcription of specific sets of genes during embryonic development. Here we describe the generation of stable P19 embryonic carcinoma cells expressing comparable levels of Ascl1 and Neurog2 mRNA in a tetracycline inducible system and the resulting neuronal differentiation of these cell lines in response to doxycycline.Microarray analysis of these cell lines showed that Ascl1 and Neurog2 overlap in their gene regulation but each bHLH protein also regulated distinct genes. We demonstrated that DNA methylation accompanied repression of the Oct4 gene by both Ascl1 and Neurog2, although DNA demethylation was not observed for any genes induced by either Ascl1 or Neurog2. While the ID1 gene showed preferential induction by Ascl1 over Neurog2, the induction of ID1 reporter constructs was dependent on distinct sets of E‐boxes within the ID1 proximal promoter. Finally, the induction of the Ppp1r14a gene was specifically induced by Neurog2 and this induction was at least partially due to preferences of Neurog2 for distinct canonical E‐box sequences within the Ppp1r14a promoter.Together, these results suggest that further complexities in the mechanisms by which Ascl1 and Neurog2 regulate gene expression during neuronal differentiation.
Functional variants of MARCO modulate susceptibility to respiratory syncytial virus (RSV)
Authors: Marzec Jacqui M , Ciencewicki Jon M , Kleeberger Steven R 
Published: June 2021

MARCO (macrophage associated receptor of collagenous structure) is a scavenger receptor that recognizes specific pathogens such as endotoxin and viral particles. We previously identified MARCO variants that associate with susceptibility to RSV disease, however the mechanism is unclear. We sought to identify functional variations in MARCO that modulate pathogenesis of RSV‐induced lower respiratory tract disease. The MARCO promoter was sequenced and aligned to identify variant single nucleotide polymorphisms (SNPs). Deletion constructs and MARCO polymorphic variants were amplified from genomic DNA and cloned into a pGL3 vector, transfected into A549 and Beas2B cells, and basal and RSV‐inducible MARCO activity assessed. We used gel shift analysis for binding efficiency and confirmation of transcription factor binding sites. One novel SNP within the MARCO promoter associated with RSV susceptibility in vitro, and another was located in a putative ARE site. We found a significant reduction in baseline transcriptional activity with the ‐ 1118A allele and the ‐156G allele. Similar results were found for transiently transfected cells exposed to RSV. Overall binding efficiency was reduced with the ‐156C allele, however NRF2:MARCO binding was enhanced. We conclude that functional SNPs in MARCO modulate RSV susceptibility.Support: NIEHS Division of Intramural Research.
Understanding the basis of pathogenicity in the R592W mutant of human mitochondrial alanyl‐tRNA synthetase
Authors: Borchardt Jennifer , France Katherine , Chihade Joseph 
Published: June 2021

We are exploring the effects of recently discovered (Götz, et al., (2011) Am. J. Hum. Genet. 88, 635) pathogenic mutations in human mitochondrial alanyl‐tRNA synthetase (mt AlaRS) on tRNA recognition and enzymatic function. These authors found an R592W mutation in multiple patients with severe infantile cardiomyopathy. R592 lies within the editing domain of mt AlaRS, but is distal to the aminoacyl editing active site. To characterize the editing function of mt AlaRS, we have constructed an editing‐defective C719A/Q622H mutant of the enzyme. While this editing defective mutant effectively mischarges human mt tRNAAla with serine, no accumulation of Ser‐tRNAAla is observed with wildtype or R592W mt AlaRS. We are currently investigating the hypothesis that, rather than disabling the enzyme's ability to hydrolyze misacylated tRNAs, the R592W mutation enables hydrolysis of the correctly acylated Ala‐tRNAAla product in the editing site of mt AlaRS.This work was supported by Carleton College and the Howard Hughes Medical Institute.
Degradation of the transcription factor Nrf2 in promyelocytic leukemia‐nuclear bodies
Authors: McIntosh Deneshia , Arinze Ifeanyi 
Published: June 2021

The cellular abundance of the redox‐sensitive transcription factor Nrf2 is controlled by Keap1, an inhibitory protein that functions as a substrate adaptor for ubiquitylation of Nrf2 by the Cullin3/RING box1 (Cul3‐Rbx1) E3 ubiquitin ligase complex, thereby targeting Nrf2 for proteasome‐dependent degradation in the cytoplasm. Nrf2 can also be degraded in the nucleus but mechanistic details are lacking. Here, we show that promyelocytic leukemia‐nuclear body (PML‐NB)‐enriched preparations of HepG2 cells treated with arsenic trioxide (ATO) contained modified (sumoylated) Nrf2 and the poly‐SUMO‐specific ubiquitin E3 ligase RNF4, but not Keap1. Overexpressing RNF4 decreased the steady‐state levels of Nrf2 whereas the catalytically inactive RNF4 did not. The proteasomal inhibitor MG‐132 interfered with this RNF4‐induced decrease. In MG‐132‐treated cells, the content of ubiquitylated Nrf2 was higher in cells transfected with wild‐type RNF4 than in cells transfected with catalytically inactive RNF4. RNF4 also decreased the half‐life of Nrf2, measured in PML‐NB‐enriched fractions prepared from ATO‐treated cells. Given that treatment with ATO enhances SUMO‐2/3 attachment to Nrf2, these data suggest that RNF4 induces poly‐ubiquitylation of poly‐sumoylated Nrf2, leading to its degradation by proteasomes in PML‐NBs.Supported by NIH grants #5T32HL007737, 2 SD1MD000104 and SC1CA143985
Carbon monoxide (CO)‐dependent modulation of polymorphonuclear leukocyte (PMN) adhesion to human cerebrovascular endothelial cells in an in vitro model of endotoxemia
Authors: Serizawa Fukashi , Patterson Eric , Potter Richard F , Fraser Duglas , Cepinskas Gediminas 
Published: June 2021

CO liberated from a water‐soluble CO‐releasing molecule (CORM‐3) suppresses sepsis‐induced inflammation. However, the exact mechanisms are poorly understood.In this study we examined the mechanisms of CORM‐3‐dependent modulation of PMN adhesion in an in vitro model of endotoxemia employing human‐derived cerebrovascular endothelial cells, hCMEC/D3 (provided by Dr. P. Couraud, INSERM, Paris). hCMEC/D3 were stimulated with LPS (1μg/mL) for 6 hrs. In some experiments hCMEC/D3 were: 1) pre‐treated, 2) co‐treated, or 3) treated with CORM‐3 or inactive CORM‐3 (iCORM‐3) (200μM) before/during/following stimulation with LPS, respectively.PMN rolling/adhesion to hCMEC/D3 was assessed under conditions of flow (shear stress 0.7dyn/cm2). In parallel, MAPK‐signaling (phosphorylation of p38, ERK1/2 and JNK1/2; Western Blot) and expression of adhesion molecules (E‐selectin, ICAM‐1 and VCAM‐1; RT‐PCR) were also assessed.CORM‐3 significantly reduced PMN adhesion to hCMEC/D3 with the most profound effects seen in “pre‐treatment” group. This was accompanied by decreased levels of JNK1/2 Tyr‐phosphorylation and suppressed expression of E‐selectin and VCAM‐1.The results indicate that CORM‐3 interferes with activation of JNK1/2 phosphorylation and expression of hCMEC/D3 pro‐adhesive phenotype, thus potentially could be used to reduce systemic inflammation (HSFO‐393, HSFO‐NA6914, IRF‐04–79)
VPA inhibits myo‐inositol phosphate synthase activity by inhibition of Gsk3
Authors: Yu Wenxi , Greenberg Miriam 
Published: June 2021

Valproic acid (VPA) has been used for decades for the treatment of bipolar disorder. Yet, the therapeutic mechanism of VPA is not understood, hindering the development of more effective drugs. VPA causes inositol depletion in the human brain and in yeast cells. A similar degree of inositol depletion was observed in the yeast gsk3Δ mutant, in which the four glycogen synthase kinase 3 (GSK3) homologs were deleted. VPA was shown to inhibit Gsk3 in mammalian cells. These findings suggest that Gsk3 may play a role in inositol metabolism, and that VPA may cause inositol depletion by inhibiting Gsk3. The rate limiting enzyme in yeast inositol de novo synthesis is myo‐inositol phosphate synthase (MIPS). We hypothesize that Gsk3 is required for optimal MIPS activity. Consistent with this, the gsk3Δ mutant exhibited decreased MIPS activity. Although VPA inhibited MIPS activity in wild type cells, it did not inhibit MIPS in gsk3Δ cells, suggesting that VPA‐induced inositol depletion occurs by inhibition of Gsk3, leading to decreased MIPS activity. Of the four GSK3 homologs, we show that MCK1 is required for optimal inositol synthesis. mck1Δ exhibited multiple features of inositol depletion, including inositol‐dependent growth and decreased intracellular inositol. We conclude that inhibition of MIPS by VPA is dependent on Gsk3.Funding: NIH‐DK081367 & Enhancement Award for Doctoral Student Travel
Investigating the Molecular Basis of cPLA2α Membrane Bending
Authors: Ward Katherine E , Ropa James P , Stahelin Robert V 
Published: June 2021

Signal transduction mediates disease through key molecular targets in the cell. As protein‐lipid interactions have been examined in the literature, their role in cellular signaling has become more prevalent. Lipid‐binding proteins have become high impact drug targets in cancer, inflammation and viral egress. One such target, termed cytosolic phospholipase A2 α (cPLA2α), has been shown to play a key role in the production of inflammatory mediators. A novel function of the protein was recently discovered in our lab showing that the cPLA2α C2 domain bends zwitterionic membranes, a process that is mediated by cPLA2α's ability to deeply penetrate the lipid bilayer. Others in the literature have reported cPLA2α to participate in Fc mediated phagocytosis, intra‐Golgi trafficking and endosomal trafficking further supporting cPLA2α's ability to bend membranes in cellular processes. In addition, direct evidence has been reported in the literature using siRNA showing that cPLA2α induced vesiculation in cells. These results translate into our cellular system as cells transfected with EGFP‐cPLA2α form cytoplasmic vesicular structures. We have preliminary evidence showing cPLA2α membrane bending is mediated by oligomerization. The origin of oligomerization is currently under further investigation using both in vitro and cellular techniques.This research is funded by AHA 11PRE7640028 and NIH T32GM075762
The hZIP4 transporter has two binding affinities and transports multiple transition metal ions.
Authors: Antala Sagar , Dempski Robert E. 
Published: June 2021

Cells possess numerous membrane proteins to transport essential metal ions across cellular organelles. hZIP4 (human ZIP4) is one member of the ZIP (Zrt‐ and Irt‐like proteins) family of proteins that imports zinc to the cytoplasm. Autosomal mutations in hZIP4 have been associated with the zinc deficient genetic disorder Acrodermatitis Enteropathica. hZIP4 is expressed in small intestine, stomach, colon, cecum, kidney and pancreas. Recently, hZIP4 overexpression has also been associated with the pancreatic cancer. hZIP4 has 8 transmembrane domains with extracellular N‐ and C‐termini. Furthermore, the histidine rich non‐conserved N‐terminus and intracellular domain between transmembrane‐III and IV are believed to be important for Zn2+ coordination.We have established a radio‐isotope uptake experiment in Xenopus leavis oocytes to study the selectivity and kinetics of hZIP4 cation transport. An analysis of our experimental results demonstrate that hZIP4 transports Ni2+ and Cu2+ in addition to Zn2+. Our data also shows that hZIP4 has two binding affinities where one is in the μM range and the other one is in nM concentration. Subsequent experiments have also identified possible zinc coordination amino acid residues.
Structures and allosteric motions of Ryanodine Receptor Domains
Authors: Van Petegem Filip , Tung Ching‐Chieh , Kimlicka Lynn , Yuchi Michael , Lobo Paolo A , Lau Kelvin 
Published: June 2021

Ryanodine Receptors (RyRs) form large (>;2.2 MDa) ion channels that release Ca2+ from the endoplasmic and sarcoplasmic reticulum. They are thus involved in many calcium‐dependent events, including the contraction of skeletal and cardiac muscle. Mutation in the ryr genes can cause severe skeletal muscle disorders and cardiac arrhythmias. Here we present our recent insights in the structure‐function of these membrane protein giants. We have solved crystal structures of multiple domains in the N‐terminal and central regions and mapped the positions of ~100 disease‐causing mutations. By creating pseudo‐atomic models, we analyzed the motions these domains undergo during opening and closing of the pore region.Channel opening includes major motions in the central rim. Incidentally, this area contains a hot spot for disease mutations (>;80 total). In particular, channel opening disrupts an intersubunit interface that contains the largest cluster of mutations. By solving structures of several mutant forms of the domains, we observe that mutations can alter relative domain orientations. They thus mimic the effect of binding of ligands to the related IP3 receptor. Overall, we present a model whereby the N‐terminal region of the RyR serves as a gating ring that forms a hub for allosteric control.
Involvement of Phospholipase C‐related but catalytically inactive protein in regulated exocytosis through the pleckstrin homology and the C2 domains
Authors: Takeuchi Hiroshi , Zhang Zhao , Gao Jing , Wang DaGuang , Hirata Masato 
Published: June 2021

Phospholipase C‐related but catalytically inactive protein (PRIP) is a scaffolding protein identified in our laboratory. We have recently found that the secretion of various peptide hormones from multiple organs were upregulated in PRIP knock‐out mice, indicating that PRIP regulates the exocytic event common to dense‐core vesicles in a negative manner. In this study, we investigated the molecular mechanism underlying the inhibitory role of PRIP in exocytosis.We examined the effect of exogenously expressed various mutants of PRIP on noradrenaline (NA) secretion using rat pheochromocytoma cell line, PC12. Expression of the wild‐type PRIP suppressed High‐K+ induced NA release from PC12 cells, while the mutant of which Arg‐134 in the pleckstrin homology (PH) domain was replaced with Glu (R134Q) to loose the binding to membrane phospholipid PtdIns(4,5)P2, little suppressed NA release. The mutant lacking the C2 domain was also less effective than the wild‐type PRIP to suppress exocytosis. We found that the C2 domain of PRIP interacts with the components of SNARE complex including syntaxin 1A and synaptosomal‐associated protein of 25 kDa (SNAP‐25) and inhibited the formation of stable SDS‐resistant SNARE complexes. The results suggest that the direct bindings of the PH domain to PtdIns(4,5)P2 and the C2 domain to SNARE proteins are both required for PRIP to negatively regulate exocytosis.
Enzymatic Production of Heparin Anticoagulants for Human Use
Authors: Joice April Coley , Mencio Caitlin , Raman Karthik , Painter Chelsea , Tran Vy , Kalita Mausam , Kuberan Balagurunathan 
Published: June 2021

Currently, the pharmaceutical heparin being used as an anticoagulant is isolated from bovine lungs or porcine intestines. The primary problem resulting from heparin isolation from animal sources is that this process leaves the heparin open to contamination from animal pathogens that may also be infectious to humans. Enzymatic production of heparin anticoagulant is an attractive alternative to animal derived heparin as it removes the potential for contamination with animal pathogens and also potentially reduces molecular heterogeneity. Heparosan, produced in bacteria, has a structure similar to the backbone structure of heparin and therefore is used as a precursor in the enzymatic production of heparin anticoagulant using various recombinant sulfotransferases. After production, the synthetic heparin was assessed for anticoagulant activity. This approach offers a great promise for human use in the coming years.Supported by NIHLBI grant HL107152.
The A2B adenosine receptor controls a fork in the road of mesenchymal stem cell differentiation
Authors: Carroll Shannon H , Ravid Katya 
Published: June 2021

The A2B adenosine receptor (A2BAR) is a G‐protein coupled receptor that signals by increasing cAMP and/or activating PKC signaling. The A2BAR is expressed on mesenchymal stem cells (MSCs), which give rise to osteoblast, chondrocyte and adipocyte lineages. We recently reported that activation of the A2BAR upregulates RUNX2, an inducer of osteogenesis, and increases the differentiation of MSCs to osteoblasts. Additionally, A2BAR null mice have impaired bone formation at baseline and after injury, and increased cartilage. To investigate whether the A2BAR is regulating fate determination of the MSC, we chose to study another lineage, the chondrocyte. We considered the possibility that the A2BAR upregulating effect on RUNX2 leads to differentiation of chondrocytes by a default pathway, involving RUNX2‐mediated downregulation of Sox9. Here, we report that A2BAR activation inhibits differentiation to chondrocytes by causing the downregulation of the key transcription factor Sox9, prior to any observed effect on RUNX2 transcription. This effect is at least partially due to cAMP signaling, as direct activation of adenylyl cyclase mimics the effect. These results support the contention that A2BAR‐induced cAMP elevation influences the lineage determination of the MSC via a direct stimulatory or inhibitory effect on key regulators of the osteoblastic or chrondrocytic lineage, respectively.This work was supported, in whole or in part, by National Institutes of Health Grant HL93149
The effect of IL‐17 on the extracellular matrix expression in dermal fibroblasts
Authors: Jinnin Masatoshi , Nakashima Taiji , Fukushima Satoshi , Ihn Hironobu 
Published: June 2021

Among interleukin (IL)‐17 family, IL‐17A and IL‐17F share amino acid sequence similarity and IL‐17 receptor type A. IL‐17 signaling is implicated in the pathogenesis of various autoimmune diseases, but its role in the regulatory mechanism of extracellular matrix (ECM) expression remains to be elucidated. We found that dermal fibroblasts express type A receptor for IL‐17. We then determined whether IL‐17 affects ECM expression in dermal fibroblasts.The cDNA array of ECM genes and Immunoblot showed IL‐17A, not IL‐17F, reduced the expression of connective tissue growth factor (CTGF) as well as α1(I) collagen. miR‐129–5p increased by IL‐17A and mediated the α1(I) collagen reduction. These results suggest that IL‐17A signaling, not IL‐17F, has an anti‐fibrogenic effect via the up‐regulation of miR‐129–5p and the down‐regulation of CTGF and α1(I) collagen. Expression of IL‐17 receptor in cultured dermal fibroblasts derived from patients with scleroderma, one of the fibrotic skin diseases, was significantly down‐regulated, which may contribute to the tissue fibrosis.IL‐17A may play important roles in tissue remodeling of the skin. Clarifying the novel regulatory mechanisms of tissue remodeling by IL‐17A signal may lead to a new therapeutic approach against various skin diseases with connective tissue abnormalities.
Association of lipocalin‐2 in colorectal cancer metastasis to the liver
Authors: Elamparo Fides , Zhang Yu , Hughes Daniel , Pena Maria 
Published: June 2021

Metastasis of colorectal cancer (CRC) to the liver reduces the five‐year survival rate to less than 12% and is the major cause of death from CRC. Metastasis occurs due to productive collaborations between tumor cells and host‐derived cells in the tumor microenvironment. A pre‐metastatic niche is created to prime for cancer cell invasion into the target organ. In Balb/c mice bearing cecal tumors derived from a highly metastatic CRC cell line, microarray analysis revealed that lipocalin‐2 (LCN2) is the most highly expressed protein in the liver prior to metastasis. Western blot analysis and examination of blood serum by ELISA illustrated increased levels of LCN2 as tumors progressed into metastasis. RT‐PCR analysis showed that greater levels of LCN2 mRNA were found in highly metastatic cells as compared to less metastatic CRC cells. Our results show that LCN2 is highly associated with the development of colorectal cancer metastasis to the liver, with increased levels indicating advanced stages of metastatic progression.Center for Colon Cancer Research, University of South Carolina, Columbia, South Carolina
Effects of high‐vegetable, energy‐restricted Bibimbap on weight loss and fasting blood glucose and blood lipid parameters in overweight and obese adults
Authors: Hwang Ji‐Yun , Lee Mi Jung , Ha Ji Won , Han Jung‐Ah , Hong Wansoo , Park Kwan Hwa , Lee Jin‐Sil 
Published: June 2021

This study was to evaluate the effects of high‐vegetable Bibimbap consumption on weight loss, body composition, and blood parameters. Overweight subjects (BMI>;=23) were assigned to either a daily single meal of high‐vegetable Bibimbap (HV diet; 12 g fiber/meal/day) or a weight‐loss regimen (WL diet), with the dietary counseling of same energy intake (1800kcal for males; 1500kcal for females) for 12 weeks. Fifty‐five subjects (23 males and 32 females; 26.8¡¾8.1 years) completed the study. From the baseline to 6th week, weight and BMI were decreased significantly in both HV (−1.7kg, P=0.000; −0.62kg/m2, P=0.000 ) and WL(− 1.1kg, P=0.000; −0.40kg/m2, P=0.000) groups but such findings existed only in the HV group(−1.0kg, P=0.010; −0.37kg/m2, P=0.008) from 6th to 12th week. After 12 weeks fasting blood glucose levels were decreased in both HV(−7.0mg/Dl, P=0.000) and WL(−3.5mg/Dl, P=0.042) groups and were lower in the HV group (P=0.030). Blood levels of total cholesterol(P=0.039) and HDL(P=0.050) were lower in the HV group than in the WL group at the 6th week but not at the 12th week. Incorporating a daily high‐vegetable Bibimbap into a weight‐loss regimen improved consistent weight loss and would be effective in the weight‐loss programs.This research was supported by grants from the Globalization of Korean Foods R & D program, funded by the Ministry of Food, Agriculture, Forestry and Fisheries, Republic of Korea.