gebhart/patrepeval huggingface dataset:
Genome-Wide Manipulations of Drosophila melanogaster with Transposons, Flp Recombinase, and ΦC31 Integrase (2012)

Transposable elements, the Flp recombinase, and the ΦC31 integrase are used in Drosophila melanogaster for numerous genome-wide manipulations. Often, their use is combined in a synergistic fashion to alter and engineer the fruit fly genome. Transposons are the foundation for all transgenic technologies in flies and hence almost all innovations in the fruit fly. They have been instrumental in the generation of genome-wide collections of insertions for gene disruption and manipulation. Many important transgenic strains of these collections are available from public repositories. The Flp protein is the most widely used recombinase to induce mitotic clones to study individual gene function. However, Flp has also been used to generate chromosome- and genome-wide collections of precise deletions, inversions, and duplications. Similarly, transposons that contain attP attachment sites for the ΦC31 integrase can be used for numerous applications. This integrase was incorporated into a transgenesis system that allows the integration of small to very large DNA fragments that can be easily manipulated through recombineering. This system allowed the creation of genomic DNA libraries for genome-wide gene manipulations and X chromosome duplications. Moreover, the attP sites are being used to create libraries of tens of thousands of RNAi constructs and tissue-specific GAL4 lines. This chapter focuses on genome-wide applications of transposons, Flp recombinase, and ΦC31 integrase that greatly facilitate experimental manipulation of Drosophila.
Nerve growth factors (NGF, BDNF) enhance axonal regeneration but are not required for survival of adult sensory neurons (1988)

Largely on the basis of studies with nerve growth factor (NGF), it is now widely accepted that development of the peripheral nervous system of vertebrates is dependent in part on the interaction of immature sensory and autonomic neurons with specific survival factors that are derived from peripheral target fields. I have found, in marked contrast to an absolute requirement for NGF during development, that adult rat dorsal root ganglion sensory neurons are not dependent on NGF or other survival factors for long-term (3-4 weeks) maintenance in vitro. When dissociated and enriched, at least 70-80% of adult DRG neurons survived and extended long processes either in the absence of exogenously added NGF or upon the removal of any possible source of endogenous NGF or other neurotrophic activity (i.e., nonneuronal cells, in chemically defined culture medium, in the presence of an excess of anti-NGF antibodies, or when cultured as single neurons in microwells). Although not required for survival or expression of a range of complex morphologies, both NGF and brain-derived neurotrophic factor (BDNF) were found to stimulate the regeneration of axons from adult DRG neurons.
Absence of Direct Antioxidant Effects from Volatile Anesthetics in Primary Mixed Neuronal–Glial Cultures (2001)

Volatile anesthetics reduced cell death induced by oxidative stress only in the context of iron challenge. The likely reason for protection against iron toxicity is inhibition of iron uptake and therefore indirect reduction of subsequent intracellular oxidative stress caused by this challenge. These data argue against a primary antioxidant effect of volatile anesthetics.
PERFORMANCE ENHANCEMENT OF STBC OFDM- CDMA SYSTEM USING CHANNEL CODING TECHNIQUES OVER MULTIPATH FADING CHANNEL (2009)

In this paper, a mitigation method for minimizing the distortion caused by frequency selective fast fading is illustrated. The proposed system combines between two types of diversity schemes; frequency diversity represented by the orthogonal frequency division multiplexing (OFDM) scheme and antenna diversity represented by space time block coding (STBC) scheme these are for mitigating the distortion caused by frequency selective fading. The multiplexing technique proposed here is the code division multiple access (CDMA) scheme which is considered the solution for eliminating the distortion caused by fast fading. The main problem of the OFDM system is the high peak-to-average power ratio (PAR). In the proposed system, three techniques used for reducing the PAR are applied together. And in order to reduce the bit error rate caused by intersymbol interference (ISI) or multiple access interference (MAI) channel coding is applied which can make error detection and correction at the receiver and simulation results have shown that by applying any of channel coding techniques, the bit error rate (BER) has reached a satisfying level at a low signal-to-noise ratio (SNR).
CRISPR-CasΦ from huge phages is a hypercompact genome editor (2020)

CRISPR-Cas systems are found widely in prokaryotes, where they provide adaptive immunity against virus infection and plasmid transformation. We describe a minimal functional CRISPR-Cas system, comprising a single ~70-kilodalton protein, CasΦ, and a CRISPR array, encoded exclusively in the genomes of huge bacteriophages. CasΦ uses a single active site for both CRISPR RNA (crRNA) processing and crRNA-guided DNA cutting to target foreign nucleic acids. This hypercompact system is active in vitro and in human and plant cells with expanded target recognition capabilities relative to other CRISPR-Cas proteins. Useful for genome editing and DNA detection but with a molecular weight half that of Cas9 and Cas12a genome-editing enzymes, CasΦ offers advantages for cellular delivery that expand the genome editing toolbox.
In Vivo Assessment of Local Phosphodiesterase Activity Using Tailored Cyclic Nucleotide–Gated Channels as Camp Sensors (2001)

Phosphodiesterases (PDEs) catalyze the hydrolysis of the second messengers cAMP and cGMP. However, little is known about how PDE activity regulates cyclic nucleotide signals in vivo because, outside of specialized cells, there are few methods with the appropriate spatial and temporal resolution to measure cyclic nucleotide concentrations. We have previously demonstrated that adenovirus-expressed, olfactory cyclic nucleotide-gated channels provide real-time sensors for cAMP produced in subcellular compartments of restricted diffusion near the plasma membrane (Rich, T.C., K.A. Fagan, H. Nakata, J. Schaack, D.M.F. Cooper, and J.W. Karpen. 2000. J. Gen. Physiol. 116:147-161). To increase the utility of this method, we have modified the channel, increasing both its cAMP sensitivity and specificity, as well as removing regulation by Ca(2)+-calmodulin. We verified the increased sensitivity of these constructs in excised membrane patches, and in vivo by monitoring cAMP-induced Ca(2)+ influx through the channels in cell populations. The improved cAMP sensors were used to monitor changes in local cAMP concentration induced by adenylyl cyclase activators in the presence and absence of PDE inhibitors. This approach allowed us to identify localized PDE types in both nonexcitable HEK-293 and excitable GH4C1 cells. We have also developed a quantitative framework for estimating the K(I) of PDE inhibitors in vivo. The results indicate that PDE type IV regulates local cAMP levels in HEK-293 cells. In GH4C1 cells, inhibitors specific to PDE types I and IV increased local cAMP levels. The results suggest that in these cells PDE type IV has a high K(m) for cAMP, whereas PDE type I has a low K(m) for cAMP. Furthermore, in GH4C1 cells, basal adenylyl cyclase activity was readily observable after application of PDE type I inhibitors, indicating that there is a constant synthesis and hydrolysis of cAMP in subcellular compartments near the plasma membrane. Modulation of constitutively active adenylyl cyclase and PDE would allow for rapid control of cAMP-regulated processes such as cellular excitability.
Desaturases fused to their electron donor (2001)

Fatty acid desaturations in the carboxy-terminal segment from C1—C10 are catalyzed in many, but not in all cases, by desaturase enzymes which are fused to their electron donor cytochrome b5. Several of these enzymes (“front-end desaturases”) from a wide variety of organisms have been cloned and functionally expressed for proof of regio-, stereo- and chain length-selectivity. In most cases the actual status of the substrate chain, whether coenzyme A thioester or component of a membrane lipid, is not known. The cytochrome b5 domain is located N-terminally, internally or C-terminally. Compared to the free cytochrome b5 , the fused domains show a significant reduction of acidic amino acid residues on the surface of the four helices enclosing the heme group. It is discussed how this may contribute to hydrophobic domain pairing required for interdomain electron transport. This is in contrast to the mode of interaction of free cytochrome b5 with its partners, which is governed by electrostatic charge pairing. A look at crystallized or computer-simulated models involving fused or free cytochrome b5 helps to outline the problems encountered by optimizing the docking of partners and the exchange of electrons between domains of different degrees of mobility.
Genetic relatedness and colinearity of genomes of equine herpesvirus types 1 and 3 (1986)

The arrangement and location of homologous DNA sequences within the genomes of equine herpesvirus type 1 (EHV-1) and EHV-3 were investigated by using Southern blot hybridization analyses conducted under stringent conditions. Recombinant plasmid libraries comprising 95 and 84% of the EHV-1 and EHV-3 genomes, respectively, were labeled with 32P-deoxynucleotides by nick translation and were used as probes in filter hybridization studies. The DNA homology between the EHV-1 and EHV-3 genomes was dispersed throughout the genomes in a colinear arrangement. Significant hybridization was detected between the EHV-1 short region inverted repeat sequences, which are known to encode immediate early transcripts, and the corresponding EHV-3 inverted repeat sequences. Interestingly, probes derived from the EHV-1 heterogeneous region, which is adjacent to the EHV-1 short region, hybridized strongly to EHV-3 DNA sequences within a similar genomic location, but did not reveal any corresponding heterogeneity within the EHV-3 genome. Our results demonstrated that there is a highly conserved evolutionary relationship between EHV-1 and EHV-3 and provided the foundation for further investigations to determine whether similarities in protein function underpin the genetic relatedness between these two herpesviruses.
Elastography: Elasticity Imaging Using Ultrasound with Application to Muscle and Breast in Vivo (1993)

Changes in tissue elasticity are generally correlated with its pathological state. In many cases, despite the difference in elasticity, the small size of a lesion or its location deep in the body preclude its detection by palpation. In general, such a lesion may or may not possess echogenic properties that would make it ultrasonically detectable. Elastography is an ultrasonic method for imaging the elasticity of compliant tissues. The method estimates the local longitudinal strain of tissue elements by ultrasonically assessing the one dimensional local displacements. This information can be combined with first order theoretical estimates of the local stress to yield a quantitative measure of the local elastic properties of tissue. The elasticity information is displayed in the form of a gray scale image called an elastogram. An experimental system for elastography in phantoms based on a single element transducer has been described previously [1]. Here we introduce a new elastography system based on a linear array transducer that is suitable for in vivo scanning. We describe tissue mimicking phantom experiments and preliminary in vivo breast and muscle elastograms confirming the feasibility of performing elastography in vivo. An elastogram of a breast containing an 8 mm palpable cancer nodule clearly shows the lesion. Elastograms and their corresponding sonograms show some similarities and differences in the depiction of tissue structures.
Ternary Complex of Transforming Growth Factor-β1 Reveals Isoform-specific Ligand Recognition and Receptor Recruitment in the Superfamily (2010)

Transforming growth factor (TGF)-beta1, -beta2, and -beta3 are 25-kDa homodimeric polypeptides that play crucial nonoverlapping roles in embryogenesis, tissue development, carcinogenesis, and immune regulation. Here we report the 3.0-A resolution crystal structure of the ternary complex between human TGF-beta1 and the extracellular domains of its type I and type II receptors, TbetaRI and TbetaRII. The TGF-beta1 ternary complex structure is similar to previously reported TGF-beta3 complex except with a 10 degrees rotation in TbetaRI docking orientation. Quantitative binding studies showed distinct kinetics between the receptors and the isoforms of TGF-beta. TbetaRI showed significant binding to TGF-beta2 and TGF-beta3 but not TGF-beta1, and the binding to all three isoforms of TGF-beta was enhanced considerably in the presence of TbetaRII. The preference of TGF-beta2 to TbetaRI suggests a variation in its receptor recruitment in vivo. Although TGF-beta1 and TGF-beta3 bind and assemble their ternary complexes in a similar manner, their structural differences together with differences in the affinities and kinetics of their receptor binding may underlie their unique biological activities. Structural comparisons revealed that the receptor-ligand pairing in the TGF-beta superfamily is dictated by unique insertions, deletions, and disulfide bonds rather than amino acid conservation at the interface. The binding mode of TbetaRII on TGF-beta is unique to TGF-betas, whereas that of type II receptor for bone morphogenetic protein on bone morphogenetic protein appears common to all other cytokines in the superfamily. Further, extensive hydrogen bonds and salt bridges are present at the high affinity cytokine-receptor interfaces, whereas hydrophobic interactions dominate the low affinity receptor-ligand interfaces.
In Situ Fabrication and Actuation of Polymer Magnetic Microstructures (2011)

We demonstrate a single-exposure in situ magnetic actuator fabrication technique using magnetic nanoparticles (MNs) containing UV curable polymer in a Polydimethylsiloxane (PDMS) channel. Microstructures with a 3-D anchored cantilever as well as free-floating components are fabricated in a single step at a single site without the use of a sacrificial layer. By controlling the location of high oxygen concentration area through PDMS substrate patterning, we can create partially bound and free-floating movement-restricted structures. This allows us to create complex magnetic actuators, such as a 3-D anchored cantilever, motor type, and rail-guided magnetic actuators. The actuating performance of UV photopatterned magnetic microstructures depends on the MN concentration in photopolymer resin and magnetic field intensity. The measured translational velocity of magnetic microactuators with a 1 : 10 MN concentration is 140 μm/s under 1400 G of magnetic field in poly(ethylene glycol) diacrylate resin. Also, we demonstrate selective magnetic actuation of heterogeneous structures composed of magnetic and nonmagnetic parts self-assembled in railed microfluidic channels. Only magnetic parts from the assembly selectively actuated due to the magnetic field without response to the flow. Therefore, we have developed a versatile magnetic microstructure fabrication method that is very simple and fast, enabling rapid in situ fabrication and actuation.
Effect of reductive alkylation of lysine in positions 6 and/or 8 on the histamine-releasing activity of luteinizing hormone-releasing hormone antagonists (1987)

The solid-phase reductive alkylation of the Lys side chain in positions 6 (D) and 8 (L) and position 8 alone of the LH-RH antagonist [N-Ac-D-Nal,D-Ph2,3,D-Arg6,Phe7,D-Ala10]LH-RH was investigated in an attempt to reduce the histamine-releasing activity inherent to most potent antagonists while retaining high antiovulatory activity. The protected parent analogues were prepared by conventional solid-phase peptide synthesis. After selective removal of the Lys Fmoc side chain protection, the resin-bound peptides were readily and conveniently alkylated at the epsilon amino groups with various aldehydes and ketones in the presence of NaBH3CN. The analogues were then cleaved from the resin with simultaneous deprotection by anhydrous hydrogen fluoride and purified to homogeneity in two stages: gel permeation followed by preparative reversed-phase liquid chromatography. The analogues were assayed in standard rat antiovulatory and in vitro histamine-release assays.
Selective disruption of PPARγ2 impairs the development of adipose tissue and insulin sensitivity (2004)

Peroxisome proliferator-activated receptor gamma (PPARgamma) is a nuclear receptor that plays a pivotal role in obesity and diabetes. PPARgamma has two isoforms, PPARgamma1 and PPARgamma2. We investigated the functional differences between PPARgamma1 and PPARgamma2 by selectively disrupting PPARgamma2 in mice. In contrast to the embryonic lethality of PPARgamma-deficient mice, PPARgamma2(-/-) mice survived. Although normal development was identified in other tissues we examined, PPARgamma2(-/-) mice exhibited an overall reduction in white adipose tissue, less lipid accumulation, and decreased expression of adipogenic genes in adipose tissue. In addition, insulin sensitivity was impaired in male PPARgamma2(-/-) mice, with dramatically decreased expression of insulin receptor substrate 1 and glucose transporter 4 in the skeletal muscle, but thiazolidinediones were able to normalize this insulin resistance. Consistent with in vivo data, PPARgamma2(-/-) mouse embryonic fibroblasts showed a dramatically reduced capacity for adipogenesis in vitro compared with wild-type mouse embryonic fibroblasts. Taken together, our data demonstrate that PPARgamma2 deficiency impairs the development of adipose tissue and insulin sensitivity. PPARgamma2(-/-) mice may provide a tool to study the role of PPARgamma2 in obesity and diabetes.
Filtration of bacteria and yeast by ultrasound-enhanced sedimentation (1997)

Continuous flow filtration of suspensions of eukaryotic cells by ultrasonic standing wave enhanced sedimentation has recently been reported. The filtration efficiency for Escherichia coli in such a filter has been characterized at frequencies of 1 and 3 MHz in the present work and compared with results for Saccharomyces cerevisiae. The yeast can be filtered at greater than 99% efficiency at a flow rate of 5 ml min-1 at either frequency. The filtration efficiency of the smaller E. coli at 3 MHz is in excess of 80% at concentrations in the region of 10(10) ml-1 but decreased at lower concentrations. However, E. coli in a mixed suspension with yeast were, because of inter-particle interactions, removed with the filtrate at an efficiency ranging from 80 to 50% over the eight orders of bacterial concentrations tested (down to 10(3) ml-1) at 3 MHz. Quantitative considerations show that poor filtration of pure suspensions of the smaller cells at the lower frequency arises because, at reasonable flow rates, the residence time is not sufficient for the cells to reach the pressure nodal cell concentration regions. The filtration efficiencies of both cell types are comparable at 3 MHz. It is suggested that the more comparable efficiencies arise because concentration regions are narrower at the high frequency and Stokes drag by the filter bulk flow inhibits sedimentation of the concentrated cells.
The serine proteinase inhibitor from summer squash (Cucurbita pepo): some structural features, stability and proteolytic degradation. (1985)

The serine proteinase inhibitor from summer squash seeds (CPTI-II) with Mr of about 3250 contains three disulphide bridges and is unusually resistant to denaturing agents (e.g. 10% trichloroacetic acid at about 100 degrees C), thermolysin and proteinase V8 from Staphylococcus aureus. The inhibitor is digested by pepsin; the digestion of the virgin form proceeds more rapidly than when the peptide bond of the reactive site is broken. The inhibitor is not specifically reduced by sodium borohydride at pH 8.8, and almost full reactivation of the inhibitor reduced by dithiothreitol takes place at pH 8.15 in the presence of EDTA and the reduced + oxidized glutathione system. The inhibitor was crystallized from methanol. CD spectra point to the occurrence of beta-turns in the secondary structure of the inhibitor.
Polytope vaccines for the codelivery of multiple CD8T-cell epitopes (2002)

Vaccines against a number of diseases, including HIV, Epstein Barr virus, malaria and several cancers, are believed to require the coinduction of multiple alphabeta CD8+ cytotoxic T-lymphocyte responses that are directed towards a number of different target antigens. The difficulties associated with making large recombinant vaccines that contain numerous antigens has led to the development of alphabeta CD8+ cytotoxic T-lymphocyte polyepitope or polytope vaccine approach, where multiple (usually 8-10 amino acids long) alphabeta CD8+ cytotoxic T-lymphocyte epitopes, derived from several antigens are conjoined into single artificial constructs. Such polytope constructs can be delivered using a number of different vaccine vector modalities with each epitope in the construct emerging as individually immunogenic.
Fc-fusion technology and recombinant FVIII and FIX in the management of the hemophilias (2014)

Abstract: Prophylaxis with regular infusions of factor VIII (FVIII)- or factor IX (FIX)- containing products is the mainstay of modern hemophilia care. However, this therapeutic regimen is inconvenient, requiring repeated intravenous injections from childhood. Approaches meant to prolong the half-life of FVIII and FIX in plasma have been developed in order to improve the feasibility and acceptability of replacement therapy, extending protection from bleeding, reducing infusion frequency and hence the need for venous access devices in young children. Several strategies have been implemented to enhance the pharmacokinetics of clotting factors, including conjugation with polyethylene glycol and the production by genetic engineering of fusion proteins containing the coagulation factors linked to a long-lived plasma protein such as albumin or the Fc fragment of immunoglobulin (Ig)G. The latter technology is one of the most promising, since the prolongation of FVIII and FIX half-life is obtained by exploiting the physiological binding of the Fc domain to the neonatal Fc receptor. Fc fusion monomers have been obtained with both recombinant FVIII (rFVIIIFc) and FIX (rFIXFc), and data from preclinical and clinical studies showed improved pharmacokinetics for both factors, which are produced in human embryonic kidney (HEK) 293 cells, thus ensuring full human post-translational modifications. In Phase I/IIa studies, rFVIIIFc and rFIXFc showed 1.5–1.7 fold and 3.0–4.0 fold longer elimination half-life, respectively. Similar data have been obtained in the Phase III clinical studies with rFVIIIFc and rFIX-Fc published recently. Both drugs were satisfactorily safe, particularly with respect to immunogenicity, and no serious adverse event was observed. Keywords: factor VIII, factor IX, long-acting molecules