gebhart/patrepeval huggingface dataset:
A new direct-viewing chemotaxis chamber (1991)

A new form of chamber for studying chemotaxis, similar in principle to the Zigmond chamber, allows the behaviour of the cells in a linear concentration gradient to be observed directly. The chamber was developed mainly for studying chemotaxis in fibroblasts using interferometric microscopy and the main design criteria were that it should have better optical characteristics, a higher dimensional precision and better long-term stability than the Zigmond chamber. It is made entirely from glass by grinding a blind circular well centrally in the counting platform of a Helber bacteria counting chamber. This procedure leaves an annular 'bridge', approximately 1 mm wide, between the new inner circular well and the original outer annular well. This bridge fulfils the same function as the linear bridge of the Zigmond chamber but the precise construction of the counting chamber ensures that a gap of 20 microns between bridge and coverslip can be accurately and repeatedly achieved when the chamber is assembled. It is envisaged that the improved optical clarity, dimensional accuracy and long-term stability of the new chamber will be advantageous in other applications, particularly in studies requiring critical microscopy or a precise knowledge of the gradient and in studies of cells, such as fibroblasts, that move much more slowly than neutrophils.
Invariant amino acids essential for decoding function of polypeptide release factor eRF1 (2005)

In eukaryotic ribosome, the N domain of polypeptide release factor eRF1 is involved in decoding stop signals in mRNAs. However, structure of the decoding site remains obscure. Here, we specifically altered the stop codon recognition pattern of human eRF1 by point mutagenesis of the invariant Glu55 and Tyr125 residues in the N domain. The 3D structure of generated eRF1 mutants was not destabilized as demonstrated by calorimetric measurements and calculated free energy perturbations. In mutants, the UAG response was most profoundly and selectively affected. Surprisingly, Glu55Arg mutant completely retained its release activity. Substitution of the aromatic ring in position 125 reduced response toward all stop codons. This result demonstrates the critical importance of Tyr125 for maintenance of the intact structure of the eRF1 decoding site. The results also suggest that Tyr125 is implicated in recognition of the 3d stop codon position and probably forms an H-bond with Glu55. The data point to a pivotal role played by the YxCxxxF motif (positions 125-131) in purine discrimination of the stop codons. We speculate that eRF1 decoding site is formed by a 3D network of amino acids side chains.
Relevance of Nicotine-Derived N-Nitrosamines in Tobacco Carcino-Genesis (1991)

Tobacco-specific N-nitrosamines (TSNA) are formed from the Nicotiana alkaloids by N-nitrosation. Among identified TSNA are powerful animal carcinogens, namely N′-nitrosonornicotine (NNN), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL). In animals and in human tissues these nitrosamines are metabolically activated by α-hydroxylation to highly reactive species which form adducts with protein and DNA. These three TSNA contribute significantly to the increased risk of snuff dippers for oral cancer and to the risk of smokers for cancer of the lung and oral cavity and likely also for cancer of the esophagus and pancreas. Chemopreventive approaches towards reduction of risk of TSNA-related cancer are discussed, as is the need for developing methods for the reduction of TSNA in tobacco and tobacco smoke. Future research should concentrate on biomarkers for the exposure of tobacco users to TSNA, the DNA binding of TSNA in humans, and on the question of endogenous formation of TSNA.
Overview of DNA Sequencing Strategies (2011)

Abstract Efficient and cost‐effective DNA sequencing technologies are critical to the progress of molecular biology. This overview of DNA sequencing strategies provides a high‐level review of seven distinct approaches to DNA sequencing: (a) dideoxy sequencing; (b) solid phase sequencing; (c) sequencing‐by‐hybridization; (d) mass spectrometry; (e) cyclic array sequencing; (f) microelectrophoresis; and (g) nanopore sequencing. Other platforms currently in development are also briefly described. The primary focus here is on Sanger dideoxy sequencing, which has been the dominant technology since 1977, and on cyclic array strategies, for which several competitive implementations have been developed since 2005. Because the field of DNA sequencing is changing rapidly, this unit represents a snapshot as of September, 2011. Curr. Protoc. Mol. Biol . 96:7.1.1‐7.1.23. © 2011 by John Wiley &amp; Sons, Inc.
Target gene identification: Target specific transcriptional activation by three murine Homeodomain/VP16 hybrid proteins inSaccharomyces cerevisiae (1996)

The mammalian homeodomain proteins encoded by Hox genes play an important role in embryonic development by providing positional queues which define developmental identities along the anteroposterior axis of developing organisms. These proteins bind DNA specifically through their homeodomain to sequences containing ATTA cores, and thereby are thought to exert their effect regulating downstream genes. Little is known about the specificity of binding of homeodomain proteins to their sequences and the identity of their target genes. We have developed a transcriptional activation assay in yeast which employs a homeobox/VP16 fusion gene as a transcriptional activator and a target construct in which test fragments of DNA are inserted upstream to a reporter gene. Using this assay, we compared transcriptional activation by three chimeric proteins containing the homeodomains of the mouse homeobox genes, Hoxa-5, Hoxb-6, and Hoxc-8. When tested on previously defined target sequences, strong differential specificities of activation were observed. In an effort to identify enhancers that normally respond to homeodomain transcriptional activators, random fragments of mouse genomic DNA were cloned upstream of the reporter gene. Genomic DNA fragments with distinct activation profiles were obtained and were found to share matches beyond the ATTA core with previously described enhancers. These results demonstrate that the transcriptional activation system in yeast can be used as a convenient system to detect DNA motifs which bind homeodomain proteins, and subsequently, to identify authentic target genes responsive to Hox gene proteins.
VCP/p97 is essential for maturation of ubiquitin-containing autophagosomes and this function is impaired by mutations that cause IBMPFD (2010)

VCP (VCP/p97) is a ubiquitously expressed member of the AAA(+)-ATPase family of chaperone-like proteins that regulates numerous cellular processes including chromatin decondensation, homotypic membrane fusion and ubiquitin-dependent protein degradation by the proteasome. Mutations in VCP cause a multisystem degenerative disease consisting of inclusion body myopathy, Paget disease of bone, and frontotemporal dementia (IBMPFD). Here we show that VCP is essential for autophagosome maturation. We generated cells stably expressing dual-tagged LC3 (mCherry-EGFP-LC3) which permit monitoring of autophagosome maturation. We determined that VCP deficiency by RNAi-mediated knockdown or overexpression of dominant-negative VCP results in significant accumulation of immature autophagic vesicles, some of which are abnormally large, acidified and exhibit cathepsin B activity. Furthermore, expression of disease-associated VCP mutants (R155H and A232E) also causes this autophagy defect. VCP was found to be essential to autophagosome maturation under basal conditions and in cells challenged by proteasome inhibition, but not in cells challenged by starvation, suggesting that VCP might be selectively required for autophagic degradation of ubiquitinated substrates. Indeed, a high percentage of the accumulated autophagic vesicles contain ubiquitin-positive contents, a feature that is not observed in autophagic vesicles that accumulate following starvation or treatment with Bafilomycin A. Finally, we show accumulation of numerous, large LAMP-1 and LAMP-2-positive vacuoles and accumulation of LC3-II in myoblasts derived from patients with IBMPFD. We conclude that VCP is essential for maturation of ubiquitin-containing autophagosomes and that defect in this function may contribute to IBMPFD pathogenesis.
Lifetime Overproduction of Circulating Angiotensin-(1-7) Attenuates Deoxycorticosterone Acetate-Salt Hypertension-Induced Cardiac Dysfunction and Remodeling (2010)

We evaluated the development of arterial hypertension, cardiac function, and collagen deposition, as well as the level of components of the renin-angiotensin system in the heart of transgenic rats that overexpress an angiotensin (Ang)-(1-7)-producing fusion protein, TGR(A1-7)3292 (TG), which induces a lifetime increase in circulating levels of this peptide. After 30 days of the induction of the deoxycorticosterone acetate (DOCA)-salt hypertension model, DOCA-TG rats were hypertensive but presented a lower systolic arterial pressure in comparison with DOCA-Sprague-Dawley (SD) rats. In contrast to DOCA-SD rats that presented left ventricle (LV) hypertrophy and diastolic dysfunction, DOCA-TG rats did not develop cardiac hypertrophy or changes in ventricular function. In addition, DOCA-TG rats showed attenuation in mRNA expression for collagen type I and III compared with the increased levels of DOCA-SD rats. Ang II plasma and LV levels were reduced in SD and TG hypertensive rats in comparison with normotensive animals. DOCA-TG rats presented a reduction in plasma Ang-(1-7) levels; however, there was a great increase in Ang-(1-7) ( approximately 3-fold) accompanied by a decrease in mRNA expression of both angiotensin-converting enzyme and angiotensin-converting enzyme 2 in the LV. The mRNA expression of Mas and Ang II type 1 receptors in the LV was not significantly changed in DOCA-SD or DOCA-TG rats. This study showed that TG rats with increased circulating levels of Ang-(1-7) are protected against cardiac dysfunction and fibrosis and also present an attenuated increase in blood pressure after DOCA-salt hypertension. In addition, DOCA-TG rats showed an important local increase in Ang-(1-7) levels in the LV, which might have contributed to the attenuation of cardiac dysfunction and prefibrotic lesions.
Glutamate receptor subunit expression in primary neuronal and secondary glial cultures (2001)

We report on the expression of ionotropic glutamate receptor subunits in primary neuronal cultures from rat cortex, hippocampus and cerebellum and of metabotropic glutamate (mGlu) receptor subtypes in these neuronal cultures as well as in cortical astroglial cultures. We found that the NMDA receptor (NR) subunits NR1, NR2A and NR2B were expressed in all three cultures. Each of the three cultures showed also expression of the four AMPA receptor subunits. Although RT-PCR detected mRNA of all kainate (KA) subunits in the three cultures, western blot showed only expression of Glu6 and KA2 receptor subunits. The expression analysis of mGlu receptors indicated the presence of all mGlu receptor subtype mRNAs in the three neuronal cultures, except for mGlu2 receptor mRNA, which was not detected in the cortical and cerebellar culture. mGlu1a/alpha, -2/3 and -5 receptor proteins were present in all three cultures, whereas mGlu4a and mGlu8a receptor proteins were not detected. Astroglial cultures were grown in either serum-containing or chemically defined medium. Only mGlu5 receptor protein was found in astroglial cultures grown in serum-containing medium. When astrocytes were cultured in chemically defined medium, mGlu3, -5 and -8 receptor mRNAs were detected, but at the protein level, still only mGlu5 receptor was found.
Molecular cloning, nucleotide sequence, and characterization of a 40,000-molecular-weight lipoprotein of Haemophilus somnus (1992)

A gene of Haemophilus somnus encoding the major 40,000-molecular-weight antigen (LppA) was cloned on a 2-kb Sau3AI fragment. The nucleotide sequence of the entire DNA insert was determined. One open reading frame, encoding a 247-residue polypeptide with a calculated molecular weight of 27,072, was identified. This reading frame was confirmed by sequencing the fusion joint of two independent IppA::TnphoA gene fusions. The 21 amino-terminal amino acids of the deduced polypeptide showed strong sequence homology to the signal peptide of secreted proteins, and the sequence Leu-Leu-Ala-Ala-Cys at the putative cleavage site is identical to the consensus cleavage sequence of lipoproteins from gram-negative bacteria. The presence of the lipid moiety on the protein was shown by incorporation of radioactive palmitic acid into the natural H. somnus protein. Palmitic acid could also be incorporated into the recombinant protein in Escherichia coli. Synthesis of the mature LppA lipoprotein was inhibited by globomycin, showing that cleavage of the signal peptide is mediated by signal peptidase II in both organisms. By using site-directed mutagenesis, the cysteine residue at the cleavage site was changed to glycine. Radiolabelled palmitate was not incorporated into the mutated protein, showing that lipid modification occurs at the Cys-22 residue.
Targeted cleavage of mRNA in vitro by RNase P from Escherichia coli. (1992)

External guide sequences (EGSs) complementary to mRNAs that encode beta-galactosidase from Escherichia coli and nuclease A from Staphylococcus aureus can target these RNAs for cleavage in vitro by RNase P from E. coli. Specific cleavage occurs at locations predicted by the nucleotide sequences of the EGSs. EGSs with regions complementary to the mRNAs that are as short as 13 nucleotides function efficiently and turn over slowly during incubation with the target substrate and the enzyme. EGSs composed of deoxyribonucleotides as well as those composed of ribonucleotides are effective, but cleavage of the targeted substrate with DNA as an EGS is about 10-fold less efficient than that with RNA as an EGS. An RNA EGS inhibited the formation of beta-galactosidase activity in a crude extract (S30) of E. coli that was capable of catalyzing coupled transcription-translation reactions.
An Asparaginyl Endopeptidase Mediates in Vivo Protein Backbone Cyclization (2007)

Proteases can catalyze both peptide bond cleavage and formation, yet the hydrolysis reaction dominates in nature. This presents an interesting challenge for the biosynthesis of backbone cyclized (circular) proteins, which are encoded as part of precursor proteins and require post-translational peptide bond formation to reach their mature form. The largest family of circular proteins are the plant-produced cyclotides; extremely stable proteins with applications as bioengineering scaffolds. Little is known about the mechanism by which they are cyclized in vivo but a highly conserved Asn (occasionally Asp) residue at the C terminus of the cyclotide domain suggests that an enzyme with specificity for Asn (asparaginyl endopeptidase; AEP) is involved in the process. Nicotiana benthamiana does not endogenously produce circular proteins but when cDNA encoding the precursor of the cyclotide kalata B1 was transiently expressed in the plants they produced the cyclotide, together with linear forms not commonly observed in cyclotide-containing plants. Observation of these species over time showed that in vivo asparaginyl bond hydrolysis is necessary for cyclization. When AEP activity was suppressed, either by decreasing AEP gene expression or using a specific inhibitor, the amount of cyclic cyclotide in the plants was reduced compared with controls and was accompanied by the accumulation of extended linear species. These results suggest that an AEP is responsible for catalyzing both peptide bond cleavage and ligation of cyclotides in a single processing event.
Rose Chafer (Coleoptera: Scarabaeidae): Improved Attractants for Adults (1990)

In the years 1982-1985, more than 60 compounds were evaluated in comparison to the standards valeric acid and hexanoic acid for attractancy to adult rose chafers, Macrodactylus subspinosus (F.), under field conditions. Many new, effective lures were discovered, including octyl butyrate; nonyl butyrate; 1-nonanol, a binary mixture of valerie and hexanoic acids (1:1); and the trinary valerie acid + hexanoic acid + octyl butyrate (1: 1:1). Trap color was proven to influence beetle attractancy; white traps were superior to yellow traps.
Blockade of Thrombospondin-1-CD47 Interactions Prevents Necrosis of Full Thickness Skin Grafts (2008)

TSP1 through CD47 limits skin graft survival. Blocking TSP1 binding or suppressing CD47 expression drastically increases graft survival. The therapeutic applications of this approach could include burn patients and the broader group of people requiring grafts or tissue flaps for closure and reconstruction of complex wounds of diverse etiologies.
Rate-determining complexation in catalytic hydrolysis of unactivated esters in neutral water (1989)

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTRate-determining complexation in catalytic hydrolysis of unactivated esters in neutral waterJik Chin and Mariusz BanaszczykCite this: J. Am. Chem. Soc. 1989, 111, 7, 2724–2726Publication Date (Print):March 1, 1989Publication History Published online1 May 2002Published inissue 1 March 1989https://pubs.acs.org/doi/10.1021/ja00189a062https://doi.org/10.1021/ja00189a062research-articleACS PublicationsRequest reuse permissionsArticle Views210Altmetric-Citations25LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-AlertscloseSupporting Info (1)»Supporting Information Supporting Information Get e-Alerts
Automatic and self-contained calibration of a multi-sensorial humanoid's upper body (2012)

Complex manipulation tasks require an accurate interplay of actuation and sensing. This accuracy can only be achieved by calibrating the relevant components beforehand. Typically calibration procedures are time-consuming and often include subsequent calibration steps, involve multiple people and require external tools. In this paper we alleviate these issues by auto-calibrating the different sensors of DLR's humanoid Rollin' Justin in a single, completely automatic and self-contained procedure, i.e. without calibration plate. By observing a single point feature on each wrist while moving the robot's head, the stereo cameras' intrinsic and extrinsic parameters are calibrated together with the arm joint elasticities and joint angle offsets. Additionally, we use the head motion to calibrate an Inertial Measurement Unit (IMU) extrinsically. Parameters are obtained by formulating the calibration problem as a batch-optimization problem that estimates all parameters jointly. A rough initial guess, as is, e.g., available when re-calibrating, is needed for the estimation and to facilitate marker detection. The procedure is validated on real hardware and reduces the effort considerably allowing rapid (5 min movement time), automatic, and accurate calibration by simply “pushing a button”.
Royal Jelly Inhibits the Production of Proinflammatory Cytokines by Activated Macrophages (2004)

In this study, we have examined the anti-inflammatory actions of royal jelly (RJ) at a cytokine level. When supernatants of RJ suspensions were added to a culture of mouse peritoneal macrophages stimulated with lipopolysaccharide and IFN-gamma, the production of proinflammatory cytokines, such as TNF-alpha, IL-6, and IL-1, was efficiently inhibited in a dose-dependent manner without having cytotoxic effects on macrophages. This suggests that RJ contains factor(s) responsible for the suppression of proinflammatory cytokine secretion. We named the factor for honeybees RJ-derived anti-inflammatory factor (HBRJ-AIF), and further investigated the molecular aspects of it. Size fractionation study showed that HBRJ-AIF is composed of substances of low (< 5 kDa) and high (> 30 kDa) molecular weights, with the former being a major component. Chromatographic analysis showed that MRJP3 is one candidate for the HBRJ-AIF with high molecular weights. Thus, our results suggest that RJ has anti-inflammatory actions through inhibiting proinflammatory cytokine production by activated macrophages.
IL-2 immunotoxin denileukin diftitox reduces regulatory T cells and enhances vaccine-mediated T-cell immunity (2007)

CD4+CD25+Foxp3+ regulatory T (Treg) cells have been implicated in the lack of effective antitumor immunity. Denileukin diftitox (DAB(389)IL-2), a fusion protein of interleukin 2 (IL-2) and diphtheria toxin, provides a means of targeting Treg cells. In this study, we examined (1) the effect of denileukin diftitox on the deletion of Treg cells in various lymphoid compartments and (2) the dose scheduling of denileukin diftitox in combination with a recombinant poxviral vaccine to enhance antigen-specific immune responses. Treg cells in spleen, peripheral blood, and bone marrow of normal C57BL/6 mice were variously reduced after a single intraperitoneal injection of denileukin diftitox; the reduction was evident within 24 hours and lasted approximately 10 days. Injection of denileukin diftitox 1 day before vaccination enhanced antigen-specific T-cell responses above levels induced by vaccination alone. These studies show for the first time in a murine model (1) the differential effects of denileukin diftitox on Treg cells in different cellular compartments, (2) the advantage of combining denileukin diftitox with a vaccine to enhance antigen-specific T-cell immune responses, (3) the lack of inhibition by denileukin diftitox of host immune responses directed against a live viral vector, and (4) the importance of dose scheduling of denileukin diftitox when used in combination with a vaccine.
Neuropilin-2 is a receptor for the vascular endothelial growth factor (VEGF) forms VEGF-145 and VEGF-165. (2000)

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