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Williams, Ryan M., Amanda R. Kulick, Srilakshmi Yedlapalli, Louisa Battistella, Cyrus J. Hajiran, and Letha J. Sooter. "In VitroSelection of a Single-Stranded DNA Molecular Recognition Element Specific for Bromacil." Journal of Nucleic Acids 2014 (2014): 1–8. http://dx.doi.org/10.1155/2014/102968.
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Bromacil is a widely used herbicide that is known to contaminate environmental systems. Due to the hazards it presents and inefficient detection methods, it is necessary to create a rapid and efficient sensing device. Towards this end, we have utilized a stringentin vitroselection method to identify single-stranded DNA molecular recognition elements (MRE) specific for bromacil. We have identified one MRE with high affinity (Kd=9.6 nM) and specificity for bromacil compared to negative targets of selection and other pesticides. The selected ssDNA MRE will be useful as the sensing element in a field-deployable bromacil detection device.
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Bowman, Teresa V., and Eirini Trompouki. "Sensing Stemness." Current Stem Cell Reports 7, no. 4 (October 6, 2021): 219–28. http://dx.doi.org/10.1007/s40778-021-00201-w.
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Abstract Purpose of Review Hematopoietic stem cells (HSCs) are formed embryonically during a dynamic developmental process and later reside in adult hematopoietic organs in a quiescent state. In response to their changing environment, HSCs have evolved diverse mechanisms to cope with intrinsic and extrinsic challenges. This review intends to discuss how HSCs and other stem cells co-opted DNA and RNA innate immune pathways to fine-tune developmental processes. Recent Findings Innate immune receptors for nucleic acids like the RIG-I-like family receptors and members of DNA sensing pathways are expressed in HSCs and other stem cells. Even though the “classic” role of these receptors is recognition of foreign DNA or RNA from pathogens, it was recently shown that cellular transposable element (TE) RNA or R-loops activate such receptors, serving as endogenous triggers of inflammatory signaling that can shape HSC formation during development and regeneration. Summary Endogenous TEs and R-loops activate RNA and DNA sensors, which trigger distinct inflammatory signals to fine-tune stem cell decisions. This phenomenon could have broad implications for diverse somatic stem cells, for a variety of diseases and during aging.
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Booth, Marsilea Adela, Sally Ann Harbison, and Jadranka Travas-Sejdic. "Developing Polypyrrole-Based Oligonucleotide Biosensors." Materials Science Forum 700 (September 2011): 215–18. http://dx.doi.org/10.4028/www.scientific.net/msf.700.215.
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Many medical, forensic science, environmental and general scientific difficulties may be aided by the existence of suitable biosensors such as gene sensors, body fluid detection DNA sensors, disease detection DNA sensors etc. The sensor technology described here uses the conducting polymer polypyrrole (PPy) as both sensing element and transducer of sensing events. Stability and reproducibility are necessary characteristics of practical biosensors. The stability of polymers can be investigated using electrical impedance spectroscopy (EIS). This work discusses research focused towards creating a stable, reproducible sensor surface for oligonucleotide detection. The effect of electropolymerisation conditions (electropolymerisation method, solvent and electrolyte used), post-growth treatment (cycling and EIS experiments), and the sensing-environment conditions on sensor stability and applicability will be discussed.
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Cao, Zhong, Zhong Liang Xiao, Yun Lin Dai, Masao Kamahori, and Maki Shimoda. "Electrical Characteristics of Extended Gate FET Sensing Chip Constructed for Detection of DNA." Advanced Materials Research 97-101 (March 2010): 4189–92. http://dx.doi.org/10.4028/www.scientific.net/amr.97-101.4189.
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An extended gate field effect transistor (EGFET) sensing chip has been constructed by using one gold plate electrode for molecule recognition and FET part for signal transduction. By using a 70.7mV DC voltage onto a Ag/AgCl reference electrode, the electrical characteristics of immobilization of the oligonucleotide probe of P1 and hybridization with the target single strand DNA of P2 on the EGFET sensing chip were examined in detail. The electrical signals on the change of a threshold voltage (VT) shift at a constant ID (3000μA) in VG-ID characteristic were obtained, and the VT shift value due to hybridization was calculated to be 12 mV, which may be attributed to the decreased negative charges after hybridization occurred at the gate surface. The surface density of hybridized dsDNA on gold surface of the FET was evaluated to be about 1 × 1012 molecules/cm2, indicating that the EGFET was a promising sensing element for biochip.
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Balaji, Aditya, Songlin Yang, Jeslyn Wang, and Jin Zhang. "Graphene Oxide-Based Nanostructured DNA Sensor." Biosensors 9, no. 2 (May 30, 2019): 74. http://dx.doi.org/10.3390/bios9020074.
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Quick detection of DNA sequence is vital for many fields, especially, early-stage diagnosis. Here, we develop a graphene oxide-based fluorescence quenching sensor to quickly and accurately detect small amounts of a single strand of DNA. In this paper, fluorescent magnetic nanoparticles (FMNPs) modified with target DNA sequence (DNA-t) were bound onto the modified graphene oxide acting as the fluorescence quenching element. FMNPs are made of iron oxide (Fe3O4) core and fluorescent silica (SiO2) shell. The average particle size of FMNPs was 74 ± 6 nm and the average thickness of the silica shell, estimated from TEM results, was 30 ± 4 nm. The photoluminescence and magnetic properties of FMNPs have been investigated. Target oligonucleotide (DNA-t) was conjugated onto FMNPs through glutaraldehyde crosslinking. Meanwhile, graphene oxide (GO) nanosheets were produced by a modified Hummers method. A complementary oligonucleotide (DNA-c) was designed to interact with GO. In the presence of GO-modified with DNA-c, the fluorescence intensity of FMNPs modified with DNA-t was quenched through a FRET quenching mechanism. Our study indicates that FMNPs can not only act as a FRET donor, but also enhance the sensor accuracy by magnetically separating the sensing system from free DNA and non-hybridized GO. Results indicate that this sensing system is ideal to detect small amounts of DNA-t with limitation detection at 0.12 µM.
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Ida, Jeunice, Soo Chan, Jörn Glökler, Yee Lim, Yee Choong, and Theam Lim. "G-Quadruplexes as An Alternative Recognition Element in Disease-Related Target Sensing." Molecules 24, no. 6 (March 19, 2019): 1079. http://dx.doi.org/10.3390/molecules24061079.
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G-quadruplexes are made up of guanine-rich RNA and DNA sequences capable of forming noncanonical nucleic acid secondary structures. The base-specific sterical configuration of G-quadruplexes allows the stacked G-tetrads to bind certain planar molecules like hemin (iron (III)-protoporphyrin IX) to regulate enzymatic-like functions such as peroxidase-mimicking activity, hence the use of the term DNAzyme/RNAzyme. This ability has been widely touted as a suitable substitute to conventional enzymatic reporter systems in diagnostics. This review will provide a brief overview of the G-quadruplex architecture as well as the many forms of reporter systems ranging from absorbance to luminescence readouts in various platforms. Furthermore, some challenges and improvements that have been introduced to improve the application of G-quadruplex in diagnostics will be highlighted. As the field of diagnostics has evolved to apply different detection systems, the need for alternative reporter systems such as G-quadruplexes is also paramount.
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Hashim, U., Soon Weng Chong, and Wei-Wen Liu. "Fabrication of Silicon Nitride Ion Sensitive Field-Effect Transistor for pH Measurement and DNA Immobilization/Hybridization." Journal of Nanomaterials 2013 (2013): 1–9. http://dx.doi.org/10.1155/2013/542737.
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The fabrication of ion sensitive field-effect transistor (ISFET) using silicon nitride (Si3N4) as the sensing membrane for pH measurement and DNA is reported. For the pH measurement, the Ag/AgCl electrode was used as the reference electrode, and different pH values of buffer solution were used in the ISFET analysis. The ISFET device was tested with pH buffer solutions of pH2, pH3, pH7, pH8, and pH9. The results show that the IV characteristic of ISFET devices is directly proportional and the device’s sensitivity was 43.13 mV/pH. The ISFET is modified chemically to allow the integration with biological element to form a biologically active field-effect transistor (BIOFET). It was found that the DNA immobilization activities which occurred on the sensing membrane caused the drain current to drop due to the negatively charged backbones of the DNA probes repelled electrons from accumulating at the conducting channel. The drain current was further decreased when the DNA hybridization took place.
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Allegra, Alessandro, Claudia Petrarca, Mario Di Gioacchino, Giuseppe Mirabile, and Sebastiano Gangemi. "Electrochemical Biosensors in the Diagnosis of Acute and Chronic Leukemias." Cancers 15, no. 1 (December 26, 2022): 146. http://dx.doi.org/10.3390/cancers15010146.
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Until now, morphological assessment with an optical or electronic microscope, fluorescence in situ hybridization, DNA sequencing, flow cytometry, polymerase chain reactions, and immunohistochemistry have been employed for leukemia identification. Nevertheless, despite their numerous different vantages, it is difficult to recognize leukemic cells correctly. Recently, the electrochemical evaluation with a nano-sensing interface seems an attractive alternative. Electrochemical biosensors measure the modification in the electrical characteristics of the nano-sensing interface, which is modified by the contact between a biological recognition element and the analyte objective. The implementation of nanosensors is founded not on single nanomaterials but rather on compilating these components efficiently. Biosensors able to identify the molecules of deoxyribonucleic acid are defined as DNA biosensors. Our review aimed to evaluate the literature on the possible use of electrochemical biosensors for identifying hematological neoplasms such as acute promyelocytic leukemia, acute lymphoblastic leukemia, and chronic myeloid leukemia. In particular, we focus our attention on using DNA electrochemical biosensors to evaluate leukemias.
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Ramsay, Joshua P., Tahlia R. Bastholm, Callum J. Verdonk, Dinah D. Tambalo, John T. Sullivan, Liam K. Harold, Beatrice A. Panganiban, et al. "An epigenetic switch activates bacterial quorum sensing and horizontal transfer of an integrative and conjugative element." Nucleic Acids Research 50, no. 2 (December 14, 2021): 975–88. http://dx.doi.org/10.1093/nar/gkab1217.
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Abstract Horizontal transfer of the integrative and conjugative element ICEMlSymR7A converts non-symbiotic Mesorhizobium spp. into nitrogen-fixing legume symbionts. Here, we discover subpopulations of Mesorhizobium japonicum R7A become epigenetically primed for quorum-sensing (QS) and QS-activated horizontal transfer. Isolated populations in this state termed R7A* maintained these phenotypes in laboratory culture but did not transfer the R7A* state to recipients of ICEMlSymR7A following conjugation. We previously demonstrated ICEMlSymR7A transfer and QS are repressed by the antiactivator QseM in R7A populations and that the adjacently-coded DNA-binding protein QseC represses qseM transcription. Here RNA-sequencing revealed qseM expression was repressed in R7A* cells and that RNA antisense to qseC was abundant in R7A but not R7A*. Deletion of the antisense-qseC promoter converted cells into an R7A*-like state. An adjacently coded QseC2 protein bound two operator sites and repressed antisense-qseC transcription. Plasmid overexpression of QseC2 stimulated the R7A* state, which persisted following curing of this plasmid. The epigenetic maintenance of the R7A* state required ICEMlSymR7A-encoded copies of both qseC and qseC2. Therefore, QseC and QseC2, together with their DNA-binding sites and overlapping promoters, form a stable epigenetic switch that establishes binary control over qseM transcription and primes a subpopulation of R7A cells for QS and horizontal transfer.
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Li, Yong, Tomoki Kimura, John H. Laity, and Glen K. Andrews. "The Zinc-Sensing Mechanism of Mouse MTF-1 Involves Linker Peptides between the Zinc Fingers." Molecular and Cellular Biology 26, no. 15 (August 1, 2006): 5580–87. http://dx.doi.org/10.1128/mcb.00471-06.
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ABSTRACT Mouse metal response element-binding transcription factor-1 (MTF-1) regulates the transcription of genes in response to a variety of stimuli, including exposure to zinc or cadmium, hypoxia, and oxidative stress. Each of these stresses may increase labile cellular zinc, leading to nuclear translocation, DNA binding, and transcriptional activation of metallothionein genes (MT genes) by MTF-1. Several lines of evidence suggest that the highly conserved six-zinc finger DNA-binding domain of MTF-1 also functions as a zinc-sensing domain. In this study, we investigated the potential role of the peptide linkers connecting the four N-terminal zinc fingers of MTF-1 in their zinc-sensing function. Each of these three linkers is unique, completely conserved among all known vertebrate MTF-1 orthologs, and different from the canonical Cys2His2 zinc finger TGEKP linker sequence. Replacing the RGEYT linker between zinc fingers 1 and 2 with TGEKP abolished the zinc-sensing function of MTF-1, resulting in constitutive DNA binding, nuclear translocation, and transcriptional activation of the MT-I gene. In contrast, swapping the TKEKP linker between fingers 2 and 3 with TGEKP had little effect on the metal-sensing functions of MTF-1, whereas swapping the canonical linker for the shorter TGKT linker between fingers 3 and 4 rendered MTF-1 less sensitive to zinc-dependent activation both in vivo and in vitro. These observations suggest a mechanism by which physiological concentrations of accessible cellular zinc affect MTF-1 activity. Zinc may modulate highly specific, linker-mediated zinc finger interactions in MTF-1, thus affecting its zinc- and DNA-binding activities, resulting in translocation to the nucleus and binding to the MT-I gene promoter.
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Pirner, Heike M., and Jürgen Stolz. "Biotin Sensing inSaccharomyces cerevisiaeIs Mediated by a Conserved DNA Element and Requires the Activity of Biotin-Protein Ligase." Journal of Biological Chemistry 281, no. 18 (March 10, 2006): 12381–89. http://dx.doi.org/10.1074/jbc.m511075200.
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Yari, Abdollah, and Marzieh Saidikhah. "Trithiane silver-nanoparticles-decorated polyaniline nanofibers as sensing element for electrochemical determination of Adenine and Guanine in DNA." Journal of Electroanalytical Chemistry 783 (December 2016): 288–94. http://dx.doi.org/10.1016/j.jelechem.2016.10.063.
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Ramsay, Joshua P., Laura G. L. Tester, Anthony S. Major, John T. Sullivan, Christina D. Edgar, Torsten Kleffmann, Jackson R. Patterson-House, et al. "Ribosomal frameshifting and dual-target antiactivation restrict quorum-sensing–activated transfer of a mobile genetic element." Proceedings of the National Academy of Sciences 112, no. 13 (March 18, 2015): 4104–9. http://dx.doi.org/10.1073/pnas.1501574112.
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Symbiosis islands are integrative and conjugative mobile genetic elements that convert nonsymbiotic rhizobia into nitrogen-fixing symbionts of leguminous plants. Excision of the Mesorhizobium loti symbiosis island ICEMlSymR7A is indirectly activated by quorum sensing through TraR-dependent activation of the excisionase gene rdfS. Here we show that a +1 programmed ribosomal frameshift (PRF) fuses the coding sequences of two TraR-activated genes, msi172 and msi171, producing an activator of rdfS expression named Frameshifted excision activator (FseA). Mass-spectrometry and mutational analyses indicated that the PRF occurred through +1 slippage of the tRNAphe from UUU to UUC within a conserved msi172-encoded motif. FseA activated rdfS expression in the absence of ICEMlSymR7A, suggesting that it directly activated rdfS transcription, despite being unrelated to any characterized DNA-binding proteins. Bacterial two-hybrid and gene-reporter assays demonstrated that FseA was also bound and inhibited by the ICEMlSymR7A-encoded quorum-sensing antiactivator QseM. Thus, activation of ICEMlSymR7A excision is counteracted by TraR antiactivation, ribosomal frameshifting, and FseA antiactivation. This robust suppression likely dampens the inherent biological noise present in the quorum-sensing autoinduction circuit and ensures that ICEMlSymR7A transfer only occurs in a subpopulation of cells in which both qseM expression is repressed and FseA is translated. The architecture of the ICEMlSymR7A transfer regulatory system provides an example of how a set of modular components have assembled through evolution to form a robust genetic toggle that regulates gene transcription and translation at both single-cell and cell-population levels.
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Li, Le, Siying Wang, Yin Xiao, and Yong Wang. "Recent Advances in Immobilization Strategies for Biomolecules in Sensors Using Organic Field-Effect Transistors." Transactions of Tianjin University 26, no. 6 (January 27, 2020): 424–40. http://dx.doi.org/10.1007/s12209-020-00234-y.
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Abstract Organic field-effect transistors (OFETs) are fabricated using organic semiconductors (OSCs) as the active layer in the form of thin films. Due to its advantages of high sensitivity, low cost, compact integration, flexibility, and printability, OFETs have been used extensively in the sensing area. For analysis platforms, the construction of sensing layers is a key element for their efficient detection capability. The strategy used to immobilize biomolecules in these devices is especially important for ensuring that the sensing functions of the OFET are effective. Generally, analysis platforms are developed by modifying the gate/electrolyte or OSC/electrolyte interface using biomolecules, such as enzymes, antibodies, or deoxyribonucleic acid (DNA) to ensure high selectivity. To provide better or more convenient biological immobilization methods for researchers in this field and thereby improve detection sensitivity, this review summarizes recent developments in the immobilization strategies used for biological macromolecules in OFETs, including cross-linking, physical adsorption, embedding, and chemical covalent binding. The influences of biomolecules on device performance are also discussed.
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Kim, Youngbae, Chih M. Lew, and Jay D. Gralla. "Escherichia coli pfs Transcription: Regulation and Proposed Roles in Autoinducer-2 Synthesis and Purine Excretion." Journal of Bacteriology 188, no. 21 (September 1, 2006): 7457–63. http://dx.doi.org/10.1128/jb.00868-06.
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ABSTRACT Pfs expression is required for several metabolic pathways and limits the production of autoinducer-2, a molecule proposed to play a central role in interspecies quorum sensing. The present study reveals physiological conditions and promoter DNA elements that regulate Escherichia coli pfs transcription. Pfs transcription is shown to rely on both sigma 70 and sigma 38 (rpoS), and the latter is subject to induction that increases pfs expression. Transcription is maximal as the cells approach stationary phase, and this level can be increased by salt stress through induction of sigma 38-dependent expression. The pfs promoter is shown to contain both positive and negative elements, which can be used by both forms of RNA polymerase. The negative element is contained within the overlapping dgt promoter, which is involved in purine metabolism. Consideration of the physiological roles of sigma 38 and dgt leads to a model for how autoinducer production is controlled under changing physiological conditions.
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Mangach, Hicham, Youssef El Badri, Abdelhamid Hmima, Abdenbi Bouzid, Younes Achaoui, and Shuwen Zeng. "Asymmetrical Dimer Photonic Crystals Enabling Outstanding Optical Sensing Performance." Nanomaterials 13, no. 3 (January 17, 2023): 375. http://dx.doi.org/10.3390/nano13030375.
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The exploration of the propensity of engineered materials to bring forward innovations predicated on their periodic nanostructured tailoring rather than the features of their individual compounds is a continuous pursuit that has propelled optical sensors to the forefront of ultra-sensitive bio-identification. Herein, a numerical analysis based on the Finite Element Method (FEM) was used to investigate and optimize the optical properties of a unidirectional asymmetric dimer photonic crystal (PhC). The proposed device has many advantages from a nanofabrication standpoint compared to conventional PhCs sensors, where integrating defects within the periodic array is imperative. The eigenvalue and transmission analysis performed indicate the presence of a protected, confined mode within the structure, resulting in a Fano-like response in the prohibited states. The optical sensor demonstrated a promising prospect for monitoring the DNA hybridization process, with a quality factor (QF) of roughly 1.53×105 and a detection limit (DL) of 4.4×10−5 RIU. Moreover, this approach is easily scalable in size while keeping the same attributes, which may potentially enable gaze monitoring.
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Todd, Bridget L., Emerson V. Stewart, John S. Burg, Adam L. Hughes, and Peter J. Espenshade. "Sterol Regulatory Element Binding Protein Is a Principal Regulator of Anaerobic Gene Expression in Fission Yeast." Molecular and Cellular Biology 26, no. 7 (April 1, 2006): 2817–31. http://dx.doi.org/10.1128/mcb.26.7.2817-2831.2006.
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ABSTRACT Fission yeast sterol regulatory element binding protein (SREBP), called Sre1p, functions in an oxygen-sensing pathway to allow adaptation to fluctuating oxygen concentrations. The Sre1p-Scp1p complex responds to oxygen-dependent sterol synthesis as an indirect measure of oxygen availability. To examine the role of Sre1p in anaerobic gene expression in Schizosaccharomyces pombe, we performed transcriptional profiling experiments after a shift to anaerobic conditions for 1.5 h. Of the 4,940 genes analyzed, expression levels of 521 (10.5%) and 686 (13.9%) genes were significantly increased and decreased, respectively, under anaerobic conditions. Sre1p controlled 68% of genes induced ≥2-fold. Oxygen-requiring biosynthetic pathways for ergosterol, heme, sphingolipid, and ubiquinone were primary targets of Sre1p. Induction of glycolytic genes and repression of mitochondrial oxidative phosphorylation genes largely did not require Sre1p. Using chromatin immunoprecipitation, we demonstrated that Sre1p acts directly at target gene promoters and stimulates its own transcription under anaerobic conditions. sre1 + promoter analysis identified two DNA elements that are both necessary and sufficient for oxygen-dependent, Sre1p-dependent transcription. Interestingly, these elements are homologous to sterol regulatory elements bound by mammalian SREBP, highlighting the evolutionary conservation between Sre1p and SREBP. We conclude that Sre1p is a principal activator of anaerobic gene expression, upregulating genes required for nonrespiratory oxygen consumption.
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Gabriel, Scott E., Faith Miyagi, Ahmed Gaballa, and John D. Helmann. "Regulation of the Bacillus subtilis yciC Gene and Insights into the DNA-Binding Specificity of the Zinc-Sensing Metalloregulator Zur." Journal of Bacteriology 190, no. 10 (March 14, 2008): 3482–88. http://dx.doi.org/10.1128/jb.01978-07.
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ABSTRACT The Bacillus subtilis Zur protein regulates zinc homeostasis by repressing at least 10 genes in response to zinc sufficiency. One of these genes, yciC, encodes an abundant protein postulated to function as a metallochaperone. Here, we used a genetic approach to identify the cis-acting elements and trans-acting factors contributing to the tight repression of yciC. Initial studies led to the identification of only trans-acting mutations, and, when the selection was repeated using a transposon library, all recovered mutants contained insertionally inactivated zur. Using a zur merodiploid strain, we obtained two cis-acting mutations that contained large deletions in the yciC regulatory region. We demonstrate that the yciC regulatory region contains two functional Zur boxes: a primary site (C2) overlapping a σA promoter ∼200 bp upstream of yciC and a second site near the translational start point (C1). Zur binds to both of these sites to mediate strong, zinc-dependent repression of yciC. Deletion studies indicate that either Zur box is sufficient for repression, although repression by Zur bound to C2 is more efficient. Binding studies demonstrate that both sites bind Zur with high affinity. Sequence alignment of these and previously described Zur boxes suggest that Zur recognizes a more extended operator than other Fur family members. We used synthetic oligonucleotides to identify bases critical for DNA binding by Zur. Unlike Fur and PerR, which bind efficiently to sequences containing a core 7-1-7 repeat element, Zur requires a 9-1-9 inverted repeat for high-affinity binding.
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Hartati, Yeni Wahyuni, Irkham Irkham, Iis Sumiati, Santhy Wyantuti, Shabarni Gaffar, Salma Nur Zakiyyah, Muhammad Ihda H. L. Zein, and Mehmet Ozsoz. "The Optimization of a Label-Free Electrochemical DNA Biosensor for Detection of Sus scrofa mtDNA as Food Adulterations." Biosensors 13, no. 6 (June 15, 2023): 657. http://dx.doi.org/10.3390/bios13060657.
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Fast, sensitive, and easy-to-use methods for detecting DNA related to food adulteration, health, religious, and commercial purposes are evolving. In this research, a label-free electrochemical DNA biosensor method was developed for the detection of pork in processed meat samples. Gold electrodeposited screen-printed carbon electrodes (SPCEs) were used and characterized using SEM and cyclic voltammetry. A biotinylated probe DNA sequence of the Cyt b S. scrofa gene mtDNA used as a sensing element containing guanine substituted by inosine bases. The detection of probe-target DNA hybridization on the streptavidin-modified gold SPCE surface was carried out by the peak guanine oxidation of the target using differential pulse voltammetry (DPV). The optimum experimental conditions of data processing using the Box–Behnken design were obtained after 90 min of streptavidin incubation time, at the DNA probe concentration of 1.0 µg/mL, and after 5 min of probe-target DNA hybridization. The detection limit was 0.135 µg/mL, with a linearity range of 0.5–1.5 µg/mL. The resulting current response indicated that this detection method was selective against 5% pork DNA in a mixture of meat samples. This electrochemical biosensor method can be developed into a portable point-of-care detection method for the presence of pork or food adulterations.
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Leichert, Lars Ingo Ole, Christian Scharf, and Michael Hecker. "Global Characterization of Disulfide Stress in Bacillus subtilis." Journal of Bacteriology 185, no. 6 (March 15, 2003): 1967–75. http://dx.doi.org/10.1128/jb.185.6.1967-1975.2003.
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ABSTRACT We used DNA macroarray and proteome analysis to analyze the regulatory networks in Bacillus subtilis that are affected by disulfide stress. To induce disulfide stress, we used the specific thiol oxidant diamide. After addition of 1 mM diamide to an exponentially growing culture, cell growth stopped until the medium was cleared of diamide. Global analysis of the mRNA expression pattern during growth arrest revealed 350 genes that were induced by disulfide stress by greater than threefold. Strongly induced genes included known oxidative stress genes that are under the control of the global repressor PerR and heat shock genes controlled by the global repressor CtsR. Other genes that were strongly induced encode putative regulators of gene expression and proteins protecting against toxic elements and heavy metals. Many genes were substantially repressed by disulfide stress, among them most of the genes belonging to the negative stringent response. Two-dimensional gels of radioactively labeled protein extracts allowed us to visualize and quantitate the massive changes in the protein expression pattern that occurred in response to disulfide stress. The observed dramatic alteration in the protein pattern reflected the changes found in the transcriptome experiments. The response to disulfide stress seems to be a complex combination of different regulatory networks, indicating that redox-sensing cysteines play a key role in different signaling pathways sensing oxidative stress, heat stress, toxic element stress, and growth inhibition.
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Shinzato, Naoya, Miho Enoki, Hiroaki Sato, Kohei Nakamura, Toru Matsui, and Yoichi Kamagata. "Specific DNA Binding of a Potential Transcriptional Regulator, Inosine 5′-Monophosphate Dehydrogenase-Related Protein VII, to the Promoter Region of a Methyl Coenzyme M Reductase I-Encoding Operon Retrieved from Methanothermobacter thermautotrophicus Strain ΔH." Applied and Environmental Microbiology 74, no. 20 (August 29, 2008): 6239–47. http://dx.doi.org/10.1128/aem.02155-07.
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ABSTRACT Two methyl coenzyme M reductases (MCRs) encoded by the mcr and mrt operons of the hydrogenotrophic methanogen Methanothermobacter thermautotrophicus ΔH are expressed in response to H2 availability. In the present study, cis elements and trans-acting factors responsible for the gene expression of MCRs were investigated by using electrophoretic mobility shift assay (EMSA) and affinity particle purification. A survey of their operator regions by EMSA with protein extracts from mrt-expressing cultures restricted them to 46- and 41-bp-long mcr and mrt upstream regions, respectively. Affinity particle purification of DNA-binding proteins conjugated with putative operator regions resulted in the retrieval of a protein attributed to IMP dehydrogenase-related protein VII (IMPDH VII). IMPDH VII is predicted to have a winged helix-turn-helix DNA-binding motif and two cystathionine β-synthase domains, and it has been suspected to be an energy-sensing module. EMSA with oligonucleotide probes with unusual sequences showed that the binding site of IMPDH VII mostly overlaps the factor B-responsible element-TATA box of the mcr operon. The results presented here suggest that IMPDH VII encoded by MTH126 is a plausible candidate for the transcriptional regulator of the mcr operon in this methanogen.
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Deng, Meng, Haitao Guo, Renate König, Maximilian Riess, Jinyao Mo, Lu Zhang, Alex Petrucelli, et al. "NLRX1 promotes HIV-1 and DNA viruses replication by blocking STING-TBK1 innate immune signaling." Journal of Immunology 198, no. 1_Supplement (May 1, 2017): 158.1. http://dx.doi.org/10.4049/jimmunol.198.supp.158.1.
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Abstract Microbial infections are the most common cause of death in humans. Type I interferon (IFN-I) is a key element bridging the host innate and adaptive immune response against infections. Delineating the molecular regulation network of IFN-I signaling is critical for developing novel antiviral strategy and benefiting rational therapy. Using an unbiased siRNA screen, we find NLRX1, one nucleotide-binding leucine-rich-repeat-containing protein, is a host factor that promotes an early step of HIV-1 infection. NLRX1 suppresses type-I interferon (IFN-I) and cytokines in response to HIV-1 reverse-transcribed DNA and enhances the nuclear import of HIV-1 DNA. In addition to HIV, NLRX1 also reduces STING-dependent host response to cytosolic DNA, c-di-GMP, cGAMP, and DNA virus. Mechanistically, NLRX1 associates with STING in mitochondria-associated ER membranes, and prevents STING recruiting TBK1 and activating downstream interferon signaling. By using purified recombinant proteins, we found NLRX1 interacts directly with STING. Furthermore, DNA virus infected Nlrx1−/− mice exhibited enhanced innate immunity and reduced morbidity and viral load. In summary, these findings reveal that NLRX1 is a checkpoint protein for DNA sensing adaptor STING and may represent a novel precision target for anti-viral therapy.
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Porter, Joshua R., Chih-Yung S. Lee, Peter J. Espenshade, and Pablo A. Iglesias. "Regulation of SREBP during hypoxia requires Ofd1-mediated control of both DNA bindingand degradation." Molecular Biology of the Cell 23, no. 18 (September 15, 2012): 3764–74. http://dx.doi.org/10.1091/mbc.e12-06-0451.
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Cells adapt to changes in ambient oxygen by changing their gene expression patterns. In fission yeast, the sterol regulatory element–binding protein Sre1 is proteolytically cleaved under low oxygen, and its N-terminal segment (Sre1N) serves as a hypoxic transcription factor. When oxygen is present, the prolyl hydroxylase Ofd1 down-regulates Sre1N activity in two ways: first, by inhibiting its binding to DNA, and second, by accelerating its degradation. Here we use a mathematical model to assess what each of these two regulatory functions contributes to the hypoxic response of the cell. By disabling individual regulatory functions in the model, which would be difficult in vivo, we found that the Ofd1 function of inhibiting Sre1N binding to DNA is essential for oxygen-dependent Sre1N regulation. The other Ofd1 function of accelerating Sre1N degradation is necessary for the yeast to quickly turn off its hypoxic response when oxygen is restored. In addition, the model predicts that increased Ofd1 production at low oxygen plays an important role in the hypoxic response, and the model indicates that the Ofd1 binding partner Nro1 tunes the response to oxygen. This model quantifies our understanding of a novel oxygen-sensing mechanism that is widely conserved.
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Ivanov, Yuri D., Kristina V. Goldaeva, Kristina A. Malsagova, Tatyana O. Pleshakova, Rafael A. Galiullin, Vladimir P. Popov, Nikolay E. Kushlinskii, et al. "Nanoribbon Biosensor in the Detection of miRNAs Associated with Colorectal Cancer." Micromachines 12, no. 12 (December 18, 2021): 1581. http://dx.doi.org/10.3390/mi12121581.
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A nanoribbon biosensor (NRBS) was developed to register synthetic DNAs that simulate and are analogous to miRNA-17-3p associated with colorectal cancer. Using this nanoribbon biosensor, the ability to detect miRNA-17-3p in the blood plasma of a patient diagnosed with colorectal cancer has been demonstrated. The sensing element of the NRBS was a nanochip based on a silicon-on-insulator (SOI) nanostructure. The nanochip included an array of 10 nanoribbons and was designed with the implementation of top-down technology. For biospecific recognition of miRNA-17-3p, the nanochip was modified with DNA probes specific for miRNA-17-3p. The performance of the nanochip was preliminarily tested on model DNA oligonucleotides, which are synthetic analogues of miRNA-17-3p, and a detection limit of ~10−17 M was achieved. The results of this work can be used in the development of serological diagnostic systems for early detection of colorectal cancer.
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Li, Szu-Yuan, Jihwan Park, Yuting Guan, Kiwung Chung, Rojesh Shrestha, Matthew B. Palmer, and Katalin Susztak. "DNMT1 in Six2 Progenitor Cells Is Essential for Transposable Element Silencing and Kidney Development." Journal of the American Society of Nephrology 30, no. 4 (March 8, 2019): 594–609. http://dx.doi.org/10.1681/asn.2018070687.
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BackgroundCytosine methylation of regulatory regions, such as promoters and enhancers, plays a key role in regulating gene expression, however, its role in kidney development has not been analyzed.MethodsTo identify functionally important epigenome-modifying enzymes and genome regions where methylation modifications are functionally important for kidney development, we performed genome-wide methylation analysis, expression profiling, and systematic genetic targeting of DNA methyltransferases (Dnmt1, Dnmt3a, and Dnmt3b) and Ten-eleven translocation methylcytosine hydroxylases (Tet2) in nephron progenitor cells (Six2Cre) in mice.ResultsGenome-wide methylome analysis indicated dynamic changes on promoters and enhancers during development. Six2CreDnmt3af/f, Six2CreDnmt3bf/f, and Six2CreTet2f/f mice showed no significant structural or functional renal abnormalities. In contrast, Six2CreDnmt1f/f mice died within 24 hours of birth, from a severe kidney developmental defect. Genome-wide methylation analysis indicated a marked loss of methylation of transposable elements. RNA sequencing detected endogenous retroviral transcripts. Expression of intracellular viral sensing pathways (RIG-I), early embryonic, nonrenal lineage genes and increased cell death contributed to the phenotype development. In podocytes, loss of Dnmt1, Dnmt3a, Dnmt3b, or Tet2 did not lead to functional or structural differences at baseline or after toxic injury.ConclusionsGenome-wide cytosine methylation and gene expression profiling showed that by silencing embryonic, nonrenal lineage genes and transposable elements, DNMT1-mediated cytosine methylation is essential for kidney development.
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van der Ploeg, Jan R. "Regulation of Bacteriocin Production in Streptococcus mutans by the Quorum-Sensing System Required for Development of Genetic Competence." Journal of Bacteriology 187, no. 12 (June 15, 2005): 3980–89. http://dx.doi.org/10.1128/jb.187.12.3980-3989.2005.
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ABSTRACT In Streptococcus mutans, competence for genetic transformation and biofilm formation are dependent on the two-component signal transduction system ComDE together with the inducer peptide pheromone competence-stimulating peptide (CSP) (encoded by comC). Here, it is shown that the same system is also required for expression of the nlmAB genes, which encode a two-peptide nonlantibiotic bacteriocin. Expression from a transcriptional nlmAB′-lacZ fusion was highest at high cell density and was increased up to 60-fold following addition of CSP, but it was abolished when the comDE genes were interrupted. Two more genes, encoding another putative bacteriocin and a putative bacteriocin immunity protein, were also regulated by this system. The regions upstream of these genes and of two further putative bacteriocin-encoding genes and a gene encoding a putative bacteriocin immunity protein contained a conserved 9-bp repeat element just upstream of the transcription start, which suggests that expression of these genes is also dependent on the ComCDE regulatory system. Mutations in the repeat element of the nlmAB promoter region led to a decrease in CSP-dependent expression of nlmAB′-lacZ. In agreement with these results, a comDE mutant and mutants unable to synthesize or export CSP did not produce bacteriocins. It is speculated that, at high cell density, bacteriocin production is induced to liberate DNA from competing streptococci.
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Hoffmann, Kerstin, Antje Wollherr, Michael Larsen, Michael Rachinger, Heiko Liesegang, Armin Ehrenreich, and Friedhelm Meinhardt. "Facilitation of Direct Conditional Knockout of Essential Genes in Bacillus licheniformis DSM13 by Comparative Genetic Analysis and Manipulation of Genetic Competence." Applied and Environmental Microbiology 76, no. 15 (June 11, 2010): 5046–57. http://dx.doi.org/10.1128/aem.00660-10.
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ABSTRACT The genetic manageability of the biotechnologically important Bacillus licheniformis is hampered due to its poor transformability, whereas Bacillus subtilis efficiently takes up DNA during genetic competence, a quorum-sensing-dependent process. Since the sensor histidine kinase ComP, encoded by a gene of the quorum-sensing module comQXPA of B. licheniformis DSM13, was found to be inactive due to an insertion element within comP, the coding region was exchanged with a functional copy. Quorum sensing was restored, but the already-poor genetic competence dropped further. The inducible expression of the key regulator for the transcription of competence genes, ComK, in trans resulted in highly competent strains and facilitated the direct disruption of genes, as well as the conditional knockout of an essential operon. As ComK is inhibited at low cell densities by a proteolytic complex in which MecA binds ComK and such inhibition is antagonized by the interaction of MecA with ComS (the expression of the latter is controlled by cell density in B. subtilis), we performed an in silico analysis of MecA and the hitherto unidentified ComS, which revealed differences for competent and noncompetent strains, indicating that the reduced competence possibly is due to a nonfunctional coupling of the comQXPA-encoded quorum module and ComK. The obtained increased genetic tractability of this industrial workhorse should improve a wide array of scientific investigations.
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Zhang, Yinxin, Olof Stefan Dallner, Tomoyoshi Nakadai, Gulya Fayzikhodjaeva, Yi-Hsueh Lu, Mitchell A. Lazar, Robert G. Roeder, and Jeffrey M. Friedman. "A noncanonical PPARγ/RXRα-binding sequence regulates leptin expression in response to changes in adipose tissue mass." Proceedings of the National Academy of Sciences 115, no. 26 (June 11, 2018): E6039—E6047. http://dx.doi.org/10.1073/pnas.1806366115.
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Leptin expression decreases after fat loss and is increased when obesity develops, and its proper quantitative regulation is essential for the homeostatic control of fat mass. We previously reported that a distant leptin enhancer 1 (LE1), 16 kb upstream from the transcription start site (TSS), confers fat-specific expression in a bacterial artificial chromosome transgenic (BACTG) reporter mouse. However, this and the other elements that we identified do not account for the quantitative changes in leptin expression that accompany alterations of adipose mass. In this report, we used an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) to identify a 17-bp noncanonical peroxisome proliferator-activated receptor gamma (PPARγ)/retinoid X receptor alpha (RXRα)-binding site, leptin regulatory element 1 (LepRE1), within LE1, and show that it is necessary for the fat-regulated quantitative control of reporter (luciferase) expression. While BACTG reporter mice with mutations in this sequence still show fat-specific expression, luciferase is no longer decreased after food restriction and weight loss. Similarly, the increased expression of leptin reporter associated with obesity inob/obmice is impaired. A functionally analogous LepRE1 site is also found in a second, redundant DNA regulatory element 13 kb downstream of the TSS. These data uncouple the mechanisms conferring qualitative and quantitative expression of the leptin gene and further suggest that factor(s) that bind to LepRE1 quantitatively control leptin expression and might be components of a lipid-sensing system in adipocytes.
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Orisakwe, Orish Ebere, Cecilia Nwadiuto Amadi, and Chiara Frazzoli. "Management of Iron Overload in Resource Poor Nations: A Systematic Review of Phlebotomy and Natural Chelators." Journal of Toxicology 2020 (January 27, 2020): 1–14. http://dx.doi.org/10.1155/2020/4084538.
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Iron is an essential element and the most abundant trace metal in the body involved in oxygen transport and oxygen sensing, electron transfer, energy metabolism, and DNA synthesis. Excess labile and unchelated iron can catalyze the formation of tissue-damaging radicals and induce oxidative stress. English abstracts were identified in PubMed and Google Scholar using multiple and various search terms based on defined inclusion and exclusion criteria. Full-length articles were selected for systematic review, and secondary and tertiary references were developed. Although bloodletting or phlebotomy remains the gold standard in the management of iron overload, this systematic review is an updated account of the pitfalls of phlebotomy and classical synthetic chelators with scientific justification for the use of natural iron chelators of dietary origin in resource-poor nations.
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Yari, Abdollah, and Somayeh Derki. "New MWCNT-Fe3O4@PDA-Ag nanocomposite as a novel sensing element of an electrochemical sensor for determination of guanine and adenine contents of DNA." Sensors and Actuators B: Chemical 227 (May 2016): 456–66. http://dx.doi.org/10.1016/j.snb.2015.12.088.
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Banerjee, Soma, Mahsa Askary Hemmat, Shambhavi Shubham, Agnivo Gosai, Sivaranjani Devarakonda, Nianyu Jiang, Charith Geekiyanage, et al. "Structurally Different Yet Functionally Similar: Aptamers Specific for the Ebola Virus Soluble Glycoprotein and GP1,2 and Their Application in Electrochemical Sensing." International Journal of Molecular Sciences 24, no. 5 (February 27, 2023): 4627. http://dx.doi.org/10.3390/ijms24054627.
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The Ebola virus glycoprotein (GP) gene templates several mRNAs that produce either the virion-associated transmembrane protein or one of two secreted glycoproteins. Soluble glycoprotein (sGP) is the predominant product. GP1 and sGP share an amino terminal sequence of 295 amino acids but differ in quaternary structure, with GP1 being a heterohexamer with GP2 and sGP a homodimer. Two structurally different DNA aptamers were selected against sGP that also bound GP1,2. These DNA aptamers were compared with a 2′FY-RNA aptamer for their interactions with the Ebola GP gene products. The three aptamers have almost identical binding isotherms for sGP and GP1,2 in solution and on the virion. They demonstrated high affinity and selectivity for sGP and GP1,2. Furthermore, one aptamer, used as a sensing element in an electrochemical format, detected GP1,2 on pseudotyped virions and sGP with high sensitivity in the presence of serum, including from an Ebola-virus-infected monkey. Our results suggest that the aptamers interact with sGP across the interface between the monomers, which is different from the sites on the protein bound by most antibodies. The remarkable similarity in functional features of three structurally distinct aptamers suggests that aptamers, like antibodies, have preferred binding sites on proteins.
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PALOMINO, Aaron, Pilar HERRERO, and Fernando MORENO. "Rgt1, a glucose sensing transcription factor, is required for transcriptional repression of the HXK2 gene in Saccharomyces cerevisiae." Biochemical Journal 388, no. 2 (May 24, 2005): 697–703. http://dx.doi.org/10.1042/bj20050160.
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Expression of HXK2, a gene encoding a Saccharomyces cerevisiae bifunctional protein with catalytic and regulatory functions, is controlled by glucose availability, being activated in the presence of glucose and inhibited when the levels of the sugar are low. In the present study, we identified Rgt1 as a transcription factor that, together with the Med8 protein, is essential for repression of the HXK2 gene in the absence of glucose. Rgt1 represses HXK2 expression by binding specifically to the motif (CGGAAAA) located at −395 bp relative to the ATG translation start codon in the HXK2 promoter. Disruption of the RGT1 gene causes an 18-fold increase in the level of HXK2 transcript in the absence of glucose. Rgt1 binds to the RGT1 element of HXK2 promoter in a glucose-dependent manner, and the repression of target gene depends on binding of Rgt1 to DNA. The physiological significance of the connection between two glucose-signalling pathways, the Snf3/Rgt2 that causes glucose induction and the Mig1/Hxk2 that causes glucose repression, was also analysed.
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Quazi, Mohzibudin Z., Taeyoung Kim, Jinhwan Yang, and Nokyoung Park. "Tuning Plasmonic Properties of Gold Nanoparticles by Employing Nanoscale DNA Hydrogel Scaffolds." Biosensors 13, no. 1 (December 24, 2022): 20. http://dx.doi.org/10.3390/bios13010020.
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Noble metals have always fascinated researchers due to their feasible and facile approach to plasmonics. Especially the extensive utilization of gold (Au) has been found in biomedical engineering, microelectronics, and catalysis. Surface plasmonic resonance (SPR) sensors are achievable by employing plasmonic nanoparticles. The past decades have seen colossal advancement in noble metal nanoparticle research. Surface plasmonic biosensors are advanced in terms of sensing accuracy and detection limit. Likewise, gold nanoparticles (AuNPs) have been widely used to develop distinct biosensors for molecular diagnosis. DNA nanotechnology facilitates advanced nanostructure having unique properties that contribute vastly to clinical therapeutics. The critical element for absolute control of materials at the nanoscale is the engineering of optical and plasmonic characteristics of the polymeric and metallic nanostructure. Correspondingly, AuNP’s vivid intense color expressions are dependent on their size, shape, and compositions, which implies their strong influence on tuning the plasmonic properties. These plasmonic properties of AuNPs have vastly exerted the biosensing and molecular diagnosis applications without any hazardous effects. Here, we have designed nanoscale X-DNA-based Dgel scaffolds utilized for tuning the plasmonic properties of AuNPs. The DNA nanohydrogel (Dgel) scaffolds engineered with three different X-DNAs of distinct numbers of base pairs were applied. We have designed X-DNA base pair-controlled size-varied Dgel scaffolds and molar ratio-based nano assemblies to tune the plasmonic properties of AuNPs. The nanoscale DNA hydrogel’s negatively charged scaffold facilitates quaternary ammonium ligand-modified positively charged AuNPs to flocculate around due to electrostatic charge attractions. Overall, our study demonstrates that by altering the DNA hydrogel scaffolds and the physical properties of the nanoscale hydrogel matrix, the SPR properties can be modulated. This approach could potentially benefit in monitoring diverse therapeutic biomolecules.
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Mahfuz, Mohammad Al, Md Anwar Hossain, Emranul Haque, Nguyen Hoang Hai, Yoshinori Namihira, and Feroz Ahmed. "A Bimetallic-Coated, Low Propagation Loss, Photonic Crystal Fiber Based Plasmonic Refractive Index Sensor." Sensors 19, no. 17 (September 1, 2019): 3794. http://dx.doi.org/10.3390/s19173794.
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In this paper, a low-loss, spiral lattice photonic crystal fiber (PCF)-based plasmonic biosensor is proposed for its application in detecting various biomolecules (i.e., sugar, protein, DNA, and mRNA) and biochemicals (i.e., serum and urine). Plasmonic material gold (Au) is employed externally to efficiently generate surface plasmon resonance (SPR) in the outer surface of the PCF. A thin layer of titanium oxide (TiO2) is also introduced, which assists in adhering the Au layer to the silica fiber. The sensing performance is investigated using a mode solver based on the finite element method (FEM). Simulation results show a maximum wavelength sensitivity of 23,000 nm/RIU for a bio-samples refractive index (RI) detection range of 1.32–1.40. This sensor also exhibits a very low confinement loss of 0.22 and 2.87 dB/cm for the analyte at 1.32 and 1.40 RI, respectively. Because of the ultra-low propagation loss, the proposed sensor can be fabricated within several centimeters, which reduces the complexity related to splicing, and so on.
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Nguyen, Duy-Khiem, and Chang-Hyun Jang. "A Simple and Ultrasensitive Colorimetric Biosensor for Anatoxin-a Based on Aptamer and Gold Nanoparticles." Micromachines 12, no. 12 (December 8, 2021): 1526. http://dx.doi.org/10.3390/mi12121526.
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Here, we designed a simple, rapid, and ultrasensitive colorimetric aptasensor for detecting anatoxin-a (ATX-a). The sensor employs a DNA aptamer as the sensing element and gold nanoparticles (AuNPs) as probes. Adsorption of the aptamer onto the AuNP surface can protect AuNPs from aggregation in NaCl solution, thus maintaining their dispersion state. In the presence of ATX-a, the specific binding of the aptamer with ATX-a results in a conformational change in the aptamer, which facilitates AuNP aggregation and, consequently, a color change of AuNPs from red to blue in NaCl solution. This color variation is directly associated with ATX-a concentration and can be easily measured using a UV/Vis spectrophotometer. The absorbance variation is linearly proportional to ATX-a concentration across the concentration range of 10 pM to 200 nM, with a detection limit of 4.45 pM and high selectivity against other interferents. This strategy was successfully applied to the detection of ATX-a in lake water samples. Thus, the present aptasensor is a promising alternative method for the rapid detection of ATX-a in the environment.
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Paonessa, Francesco, Stefania Criscuolo, Silvio Sacchetti, Davide Amoroso, Helena Scarongella, Federico Pecoraro Bisogni, Emanuele Carminati, et al. "Regulation of neural gene transcription by optogenetic inhibition of the RE1-silencing transcription factor." Proceedings of the National Academy of Sciences 113, no. 1 (December 23, 2015): E91—E100. http://dx.doi.org/10.1073/pnas.1507355112.
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Optogenetics provides new ways to activate gene transcription; however, no attempts have been made as yet to modulate mammalian transcription factors. We report the light-mediated regulation of the repressor element 1 (RE1)-silencing transcription factor (REST), a master regulator of neural genes. To tune REST activity, we selected two protein domains that impair REST-DNA binding or recruitment of the cofactor mSin3a. Computational modeling guided the fusion of the inhibitory domains to the light-sensitive Avena sativa light–oxygen–voltage-sensing (LOV) 2-phototrophin 1 (AsLOV2). By expressing AsLOV2 chimeras in Neuro2a cells, we achieved light-dependent modulation of REST target genes that was associated with an improved neural differentiation. In primary neurons, light-mediated REST inhibition increased Na+-channel 1.2 and brain-derived neurotrophic factor transcription and boosted Na+ currents and neuronal firing. This optogenetic approach allows the coordinated expression of a cluster of genes impinging on neuronal activity, providing a tool for studying neuronal physiology and correcting gene expression changes taking place in brain diseases.
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Winckler, Thomas, Negin Iranfar, Peter Beck, Ingo Jennes, Oliver Siol, Unha Baik, William F. Loomis, and Theodor Dingermann. "CbfA, the C-Module DNA-Binding Factor, Plays an Essential Role in the Initiation of Dictyostelium discoideum Development." Eukaryotic Cell 3, no. 5 (October 2004): 1349–58. http://dx.doi.org/10.1128/ec.3.5.1349-1358.2004.
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ABSTRACT We recently isolated from Dictyostelium discoideum cells a DNA-binding protein, CbfA, that interacts in vitro with a regulatory element in retrotransposon TRE5-A. We have generated a mutant strain that expresses CbfA at <5% of the wild-type level to characterize the consequences for D. discoideum cell physiology. We found that the multicellular development program leading to fruiting body formation is highly compromised in the mutant. The cells cannot aggregate and stay as a monolayer almost indefinitely. The cells respond properly to prestarvation conditions by expressing discoidin in a cell density-dependent manner. A genomewide microarray-assisted expression analysis combined with Northern blot analyses revealed a failure of CbfA-depleted cells to induce the gene encoding aggregation-specific adenylyl cyclase ACA and other genes required for cyclic AMP (cAMP) signal relay, which is necessary for aggregation and subsequent multicellular development. However, the cbfA mutant aggregated efficiently when mixed with as few as 5% wild-type cells. Moreover, pulsing cbfA mutant cells developing in suspension with nanomolar levels of cAMP resulted in induction of acaA and other early developmental genes. Although the response was less efficient and slower than in wild-type cells, it showed that cells depleted of CbfA are able to initiate development if given exogenous cAMP signals. Ectopic expression of the gene encoding the catalytic subunit of protein kinase A restored multicellular development of the mutant. We conclude that sensing of cell density and starvation are independent of CbfA, whereas CbfA is essential for the pattern of gene expression which establishes the genetic network leading to aggregation and multicellular development of D. discoideum.
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K, DARSHAN, JAGMOHAN SINGH, SURESH YADAV, VENUGOPALA K. M, and RASHMI AGGARWAL. "Root border cells: A pioneer’s of plant defence in rhizosphere." Indian Journal of Agricultural Sciences 90, no. 10 (December 4, 2020): 1850–55. http://dx.doi.org/10.56093/ijas.v90i10.107884.
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The environment around the root in rhizosphere is a complex region where multiple interactions take place among soil, plant and microorganisms. Plant growth and architecture of entire plant, in fact depends upon the roots which provide sufficient amount of nutrients and water. Root tips while sensing the rhizosphere for availability nutrients counteract with array of harmful microorganisms in the soil. In order to protect from these stresses plant roots have evolved specialized cells known as root border cells which act as front line defence mechanism in rhizosphere. Border cells are individual or bunch of programmed viable cells released from root tip which forms protective sheath between root and external environment. Border cells secret hydrated mucilage that contains antimicrobial compounds and extracellular DNA which governs the behaviour of microbiome in the soil. Production of border cell is regulated by number of factors such as phytohormones, PME enzyme and transcription factor NLP7. Plant breeding and genetic engineering could be used to exploit border cells defence mechanism as a new avenue for disease control. This review summarizes about the importance, properties of border cells, mechanism, and regulation of border cells production and role of RETs as element of plant defence.
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Ogurcovs, Andrejs, Kevon Kadiwala, Eriks Sledevskis, Marina Krasovska, Ilona Plaksenkova, and Edgars Butanovs. "Effect of DNA Aptamer Concentration on the Conductivity of a Water-Gated Al:ZnO Thin-Film Transistor-Based Biosensor." Sensors 22, no. 9 (April 29, 2022): 3408. http://dx.doi.org/10.3390/s22093408.
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Field-effect transistor-based biosensors (bio-FETs) are promising candidates for the rapid high-sensitivity and high-selectivity sensing of various analytes in healthcare, clinical diagnostics, and the food industry. However, bio-FETs still have several unresolved problems that hinder their technological transfer, such as electrical stability. Therefore, it is important to develop reliable, efficient devices and establish facile electrochemical characterization methods. In this work, we have fabricated a flexible biosensor based on an Al:ZnO thin-film transistor (TFT) gated through an aqueous electrolyte on a polyimide substrate. In addition, we demonstrated techniques for establishing the operating range of such devices. The Al:ZnO-based devices with a channel length/width ratio of 12.35 and a channel thickness of 50 nm were produced at room temperature via magnetron sputtering. These Al:ZnO-based devices exhibited high field-effect mobility (μ = 6.85 cm2/Vs) and threshold voltage (Vth = 654 mV), thus showing promise for application on temperature-sensitive substrates. X-ray photoelectron spectroscopy was used to verify the chemical composition of the deposited films, while the morphological aspects of the films were assessed using scanning electron and atomic force microscopies. The gate–channel electric capacitance of 40 nF/cm2 was determined using electrochemical impedance spectroscopy, while the electrochemical window of the gate–channel system was determined as 1.8 V (from −0.6 V to +1.2 V) using cyclic voltammetry. A deionized water solution of 10 mer (CCC AAG GTC C) DNA aptamer (molar weight −2972.9 g/mol) in a concentration ranging from 1–1000 pM/μL was used as an analyte. An increase in aptamer concentration caused a proportional decrease in the TFT channel conductivity. The techniques demonstrated in this work can be applied to optimize the operating parameters of various semiconductor materials in order to create a universal detection platform for biosensing applications, such as multi-element FET sensor arrays based on various composition nanostructured films, which use advanced neural network signal processing.
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Pachauri, Vivek, and Sven Ingebrandt. "Biologically sensitive field-effect transistors: from ISFETs to NanoFETs." Essays in Biochemistry 60, no. 1 (June 30, 2016): 81–90. http://dx.doi.org/10.1042/ebc20150009.
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Biologically sensitive field-effect transistors (BioFETs) are one of the most abundant classes of electronic sensors for biomolecular detection. Most of the time these sensors are realized as classical ion-sensitive field-effect transistors (ISFETs) having non-metallized gate dielectrics facing an electrolyte solution. In ISFETs, a semiconductor material is used as the active transducer element covered by a gate dielectric layer which is electronically sensitive to the (bio-)chemical changes that occur on its surface. This review will provide a brief overview of the history of ISFET biosensors with general operation concepts and sensing mechanisms. We also discuss silicon nanowire-based ISFETs (SiNW FETs) as the modern nanoscale version of classical ISFETs, as well as strategies to functionalize them with biologically sensitive layers. We include in our discussion other ISFET types based on nanomaterials such as carbon nanotubes, metal oxides and so on. The latest examples of highly sensitive label-free detection of deoxyribonucleic acid (DNA) molecules using SiNW FETs and single-cell recordings for drug screening and other applications of ISFETs will be highlighted. Finally, we suggest new device platforms and newly developed, miniaturized read-out tools with multichannel potentiometric and impedimetric measurement capabilities for future biomedical applications.
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Dutta, Ritika, Bruce Tiu, Arya Kaul, Bryan Mitton, and Kathleen M. Sakamoto. "CREB Increases Chemotherapy Resistance through Regulation of the DNA Damage Repair Pathway in AML Cells." Blood 126, no. 23 (December 3, 2015): 1390. http://dx.doi.org/10.1182/blood.v126.23.1390.1390.
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Abstract CREB (cAMP Response Element Binding Protein) is a nuclear transcription factor that plays a critical role in regulating myeloid cell proliferation and differentiation. CREB is overexpressed in Acute Myeloid Leukemia (AML) cells from the majority of AML patients at diagnosis, and CREB overexpression is associated with a poor prognosis.Transgenic mice overexpressing CREB in myeloid cells develop myelodysplasia/myeloproliferative neoplasms. CREB also cooperates with other oncogenes, such as Sox4, to induce transformation to AML. Knockdown of CREB inhibits AML proliferation but does not affect normal hematopoietic stem cell activity, establishing the crucial role of CREB in AML cell growth and survival. In vitro, CREB overexpression leads to increased resistance to apoptosis in AML cells. Thus, we hypothesized that increased CREB expression confers chemoresistance, as this may represent one reason that patients with high CREB levels have worse prognoses and relapse following therapy. Previous studies have demonstrated that chemotherapy resistance can result from increased DNA damage repair activity, but CREB has never been implicated in these DNA damage repair processes, nor has CREB even been described as an important transcriptional regulator of DNA damage repair genes. The goal of this study was to characterize whether CREB expression confers chemoresistance through regulation of DNA repair genes in AML cells. Firstly, we established that CREB expression levels correlate with chemoresistance by treating KG-1 cells engineered to express lower and higher levels of CREB with etoposide and doxorubicin, both chemotherapy drugs used to treat AML. Cells with CREB overexpression had increased viability compared to CREB knockdown cells after treatment with both chemotherapies at a range of concentrations. To investigate the underlying mechanism, we performed CREB chromatin immunoprecipitation and RNA-seq following small molecule CREB inhibition to identify the sets of genes that are regulated by CREB in AML cells and whose expression levels are sensitive to CREB inhibition. Out of 88 DNA damage repair genes found to be CREB-bound, 41 exhibited at least a 2-fold change in expression after CREB inhibition. qPCR was performed to determine whether the expression of DNA damage repair genes were proportional to CREB levels. Transcription of ATM, ATR, RAD54L, and RAD51, genes important in sensing and repairing DNA damage, were coordinately regulated with CREB expression. ATM, ATR, RAD54L, and RAD51 were reduced by approximately 42.0%±0.1%, 44.8%±0.1%, 40.2%±0.1%, and 27.9%±0.1% respectively in CREB knockdown cells (p≤0.05). Reduced expression of these genes also had a functional consequence. CREB knockdown cells initiated a lesser DNA damage repair response in response to etoposide treatment, as determined by measured phospho-H2AX levels, compared to wild-type CREB-expressing cells. Conversely, cells with CREB overexpression exhibited the strongest DNA damage repair response following etoposide treatment. Taken together, these data demonstrate that CREB overexpression has a protective effect against DNA damage and confers chemoresistance, likely through upregulation of DNA damage repair genes. Future studies will seek to determine if small molecule inhibition of CREB can reduce the transcription of DNA damage repair genes and thus sensitize AML cells to chemotherapeutic agents. Disclosures No relevant conflicts of interest to declare.
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Balabanova, Larissa, Yuri Shkryl, Lubov Slepchenko, Daria Cheraneva, Anna Podvolotskaya, Irina Bakunina, Olga Nedashkovskaya, Oksana Son, and Liudmila Tekutyeva. "Genomic Features of a Food-Derived Pseudomonas aeruginosa Strain PAEM and Biofilm-Associated Gene Expression under a Marine Bacterial α-Galactosidase." International Journal of Molecular Sciences 21, no. 20 (October 16, 2020): 7666. http://dx.doi.org/10.3390/ijms21207666.
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The biofilm-producing strains of P. aeruginosa colonize various surfaces, including food products and industry equipment that can cause serious human and animal health problems. The biofilms enable microorganisms to evolve the resistance to antibiotics and disinfectants. Analysis of the P. aeruginosa strain (serotype O6, sequence type 2502), isolated from an environment of meat processing (PAEM) during a ready-to-cook product storage (−20 °C), showed both the mosaic similarity and differences between free-living and clinical strains by their coding DNA sequences. Therefore, a cold shock protein (CspA) has been suggested for consideration of the evolution probability of the cold-adapted P. aeruginosa strains. In addition, the study of the action of cold-active enzymes from marine bacteria against the food-derived pathogen could contribute to the methods for controlling P. aeruginosa biofilms. The genes responsible for bacterial biofilm regulation are predominantly controlled by quorum sensing, and they directly or indirectly participate in the synthesis of extracellular polysaccharides, which are the main element of the intercellular matrix. The levels of expression for 14 biofilm-associated genes of the food-derived P. aeruginosa strain PAEM in the presence of different concentrations of the glycoside hydrolase of family 36, α-galactosidase α-PsGal, from the marine bacterium Pseudoalteromonas sp. KMM 701 were determined. The real-time PCR data clustered these genes into five groups according to the pattern of positive or negative regulation of their expression in response to the action of α-galactosidase. The results revealed a dose-dependent mechanism of the enzymatic effect on the PAEM biofilm synthesis and dispersal genes.
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Goueli, Said A., Kevin Hsiao, Hui Wang, and Matt Larsen. "Abstract 2765: Detection and quantification of cellular GAMP using homogenous HTS formatted bioluminescent assay." Cancer Research 83, no. 7_Supplement (April 4, 2023): 2765. http://dx.doi.org/10.1158/1538-7445.am2023-2765.
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Abstract The STimulator of INterferon Genes (STING) is a transmembrane protein located at the endoplasmic reticulum, which serves as a molecular hub in response to cytoplasmic cyclic dinucleotide (CDN) second messengers such as cyclic GMP-AMP (cGAMP). Upon stimulation of cyclic GMP-AMP synthase (cGAS) by a wide array of pathogens that provide extracellular microbial DNA and by intracellular (nuclear and mitochondrial) DNA, the second messenger cGAMP is generated, which then binds directly to the STING protein resulting in the activation of the STING pathway. This activation by cytosolic DNA initiates a cascade of events, cGAS-STING-TBK1-IRF3 signaling, that induces type 1 IFN response, and eventually the signaling is terminated by degradation of pathway components and clearance of stimulatory DNA. Given the significant roles of STING as an adaptor for DNA sensors to pathogens and its involvement in immune sensing, tumor growth control, and control of autoinflammatory and autoimmune disorders, the STING pathway has become a very important drug target. cGAS is well-positioned to act as a DNA sensor that triggers innate immune responses and initiates host immune responses against pathogens and cancer, but inappropriate activation of STING signaling causes severe and often fatal autoimmune or autoinflammatory diseases. Credible evidence suggests that the cGAS-cGAMP-STING pathway makes fundamental contributions to at least three major cancer therapies: radiation therapy, chemotherapy, and immunotherapy, and it thus represents a promising drug target. Therefore, there is a need to identify lead compounds that effectively modulate human STING for further drug development. Towards this goal, we have developed a bioluminescent assay to monitor the concentration of cGAMP in intracellular as well as extracellular compartments. The assay relies on the new concept of nanoluciferase (nanoluc) complementation where a small peptide (SmBiT) linked to cGAMP through a linker (cGAMP tracer) can be recognized by an anti cGAMP antibody that is linked to a nanoluc complementary fragment (LgBiT) resulting in bioluminescence upon the addition of nanoluc substrate. The presence of free cGAMP that is generated in the reaction competes with cGAMP tracer and decreases bioluminescence. The assay is sensitive and can detect less than 10 nM cGAMP and less than one ng of cGAS and thus, it is suitable for testing compounds that modulate cGAS activity. We will show data that correlate the activation of cGAS using less than 10 ng of dsDNA stimulation, and the activation of IFN response element reporter assays to detect cytokine release upon cGAS-STING activation. The assay is homogenous, HTS formatted and can be completed in less than one hour. Thus, this assay provides valuable contribution for investigating modulators of the STING pathway and facilitates the search for novel therapeutics. Citation Format: Said A. Goueli, Kevin Hsiao, Hui Wang, Matt Larsen. Detection and quantification of cellular GAMP using homogenous HTS formatted bioluminescent assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2765.
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Li, Zhanhong, Mona A. Mohamed, A. M. Vinu Mohan, Zhigang Zhu, Vinay Sharma, Geetesh K. Mishra, and Rupesh K. Mishra. "Application of Electrochemical Aptasensors toward Clinical Diagnostics, Food, and Environmental Monitoring: Review." Sensors 19, no. 24 (December 10, 2019): 5435. http://dx.doi.org/10.3390/s19245435.
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Aptamers are synthetic bio-receptors of deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) origin selected by the systematic evolution of ligands (SELEX) process that bind a broad range of target analytes with high affinity and specificity. So far, electrochemical biosensors have come up as a simple and sensitive method to utilize aptamers as a bio-recognition element. Numerous aptamer based sensors have been developed for clinical diagnostics, food, and environmental monitoring and several other applications are under development. Aptasensors are capable of extending the limits of current analytical techniques in clinical diagnostics, food, and environmental sample analysis. However, the potential applications of aptamer based electrochemical biosensors are unlimited; current applications are observed in the areas of food toxins, clinical biomarkers, and pesticide detection. This review attempts to enumerate the most representative examples of research progress in aptamer based electrochemical biosensing principles that have been developed in recent years. Additionally, this account will discuss various current developments on aptamer-based sensors toward heavy metal detection, for various cardiac biomarkers, antibiotics detection, and also on how the aptamers can be deployed to couple with antibody-based assays as a hybrid sensing platform. Aptamers can be used in various applications, however, this account will focus on the recent advancements made toward food, environmental, and clinical diagnostic application. This review paper compares various electrochemical aptamer based sensor detection strategies that have been applied so far and used as a state of the art. As illustrated in the literature, aptamers have been utilized extensively for environmental, cancer biomarker, biomedical application, and antibiotic detection and thus have been extensively discussed in this article.
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Tiaden, André, Thomas Spirig, Paula Carranza, Holger Brüggemann, Kathrin Riedel, Leo Eberl, Carmen Buchrieser, and Hubert Hilbi. "Synergistic Contribution of the Legionella pneumophila lqs Genes to Pathogen-Host Interactions." Journal of Bacteriology 190, no. 22 (September 19, 2008): 7532–47. http://dx.doi.org/10.1128/jb.01002-08.
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ABSTRACT The causative agent of Legionnaires' disease, Legionella pneumophila, is a natural parasite of environmental protozoa and employs a biphasic life style to switch between a replicative and a transmissive (virulent) phase. L. pneumophila harbors the lqs (Legionella quorum sensing) cluster, which includes genes encoding the autoinducer synthase LqsA, the sensor kinase LqsS, the response regulator LqsR, and a homologue of HdeD, which is involved in acid resistance in Escherichia coli. LqsR promotes host-cell interactions as an element of the stationary-phase virulence regulatory network. Here, we characterize L. pneumophila mutant strains lacking all four genes of the lqs cluster or only the hdeD gene. While an hdeD mutant strain did not have overt physiological or virulence phenotypes, an lqs mutant showed an aberrant morphology in stationary growth phase and was defective for intracellular growth, efficient phagocytosis, and cytotoxicity against host cells. Cytotoxicity was restored upon reintroduction of the lqs genes into the chromosome of an lqs mutant strain. The deletion of the lqs cluster caused more-severe phenotypes than deletion of only lqsR, suggesting a synergistic effect of the other lqs genes. A transcriptome analysis indicated that in the stationary phase more than 380 genes were differentially regulated in the lqs mutant and wild-type L. pneumophila. Genes involved in protein production, metabolism, and bioenergetics were upregulated in the lqs mutant, whereas genes encoding virulence factors, such as effectors secreted by the Icm/Dot type IV secretion system, were downregulated. A proteome analysis revealed that a set of Icm/Dot substrates is not produced in the absence of the lqs gene cluster, which confirms the findings from DNA microarray assays and mirrors the virulence phenotype of the lqs mutant strain.
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Wu, Liang, Erhu Xiong, Xia Zhang, Xiaohua Zhang, and Jinhua Chen. "Nanomaterials as signal amplification elements in DNA-based electrochemical sensing." Nano Today 9, no. 2 (April 2014): 197–211. http://dx.doi.org/10.1016/j.nantod.2014.04.002.
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Arvand, Majid, and Masoomeh Sayyar Ardaki. "Poly- l -cysteine/electrospun copper oxide nanofibers-zinc oxide nanoparticles nanocomposite as sensing element of an electrochemical sensor for simultaneous determination of adenine and guanine in biological samples and evaluation of damage to dsDNA and DNA purine bases by UV radiation." Analytica Chimica Acta 986 (September 2017): 25–41. http://dx.doi.org/10.1016/j.aca.2017.07.057.
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Prakoso, Agung, Riski Kurniawan, and Farchan Mauluddin. "Penanganan dan Analisa Kegagalan Pada Bleed Air Leak Detection System Pesawat Airbus A330-300 di Hanggar 3 PT.GMF AEROASIA." Jurnal Teknologi dan Riset Terapan (JATRA) 3, no. 2 (December 29, 2021): 73–78. http://dx.doi.org/10.30871/jatra.v3i2.3714.
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Sistem bleed air leak detection pada pesawat digunakan untuk mendeteksi adanya kebocoran pada bleed air duct. Sistem ini dapat mendeteksi over heat temperature di luar duct apabila terdapat kebocoran pada sekitar duct. Bleed air leak detection dilengkapi dengan sensing element yang berfungsi untuk mendeteksi kenaikan suhu apabila terjadi kebocoran disekitar bleed air duct. Sensing element ini dipasang pada engine pylon, wing, dan fuselage. Kenaikan suhu yang terdeteksi oleh sensing element dapat dimonitor di bleed monitoring computer (BMC) dan kemudian akan diteruskan ke engine bleed switch di cockpit. Troubleshooting Manual (TSM) Task 36-22-00-810-911-A digunakan untuk menyelesaikan permasalahan sistem bleed air leak detection. Metode fault tree analysis juga digunakan untuk mendapatkan kemungkinan-kemungkinan penyebab kegagalan pada sistem bleed air leak detection. Pada penelitian ini ditemukan kerusakan pada sensing element pada zone 71 HF. Sensing element tersebut telah mencapai usia 24 tahun 8 bulan dari yang ditetapkan oleh manufaktur yaitu 25 tahun, sehingga perlu dilakukan pergantian dengan sensing element yang baru. Berdasarkan hasil fault tree analysis di dapatkan beberapa kemungkinan penyebab kegagalan sistem bleed air leak detection yaitu electorstatic, short circuit, lost continuity, connector fault, control logic fault, thermistor disconnected, degradasi prestasi, dislokasi komponen, dan corrosion.
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Yu, Valen Zhuoyou, Shan Shan So, Bryan Chee-Chad Lung, Carissa Wing-Yan Wong, Ian Yu-Hong Wong, Claudia Lai-Yin Wong, Desmond Kwan-Kit Chan, et al. "Abstract 2611: p63 constrains cancer cell transposable element expressions and viral mimicry response to sustain esophageal cancer development and indicates therapeutic vulnerability." Cancer Research 83, no. 7_Supplement (April 4, 2023): 2611. http://dx.doi.org/10.1158/1538-7445.am2023-2611.
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Abstract Background: Esophageal squamous cell carcinoma (ESCC) is the predominant form of esophageal cancer in developing areas and is among the top 10 deadliest cancers with a dismal 5-year survival rate of 10~20%. ESCC molecular pathogenesis has not been well characterized, while chemoradiotherapy remains the routine treatment option besides surgery, highlighting the need for a thorough understanding of ESCC and new treatment options. Tumor protein p63 (TP63), encoding p63, plays fundamental roles in stratified epithelial homeostasis. In the esophagus, ΔNp63 is the predominant isoform and is required for normal epithelial development. ESCC retains high ΔNp63 expression level, but its contribution to ESCC development is not fully understood. Results: We found ΔNp63 maintains ESCC development, depletion of which results in great attenuation and regression of cell line-derived xenograft growth in mice. Transcriptomic profiling showed type-I interferon (IFN-I) signaling-related pathways are the top pathways enriched in ΔNp63-depleted cells. This was further verified by quantitative PCR confirming up-regulations of interferon-stimulated genes upon ΔNp63 depletion in cell lines, and by transcriptomic profiling on our latest panel of naïve ESCC patient-derived organoids (PDOs) showing TP63 expression level negatively associated with IFN-I signaling-related pathways. Elevated endogenous retrotransposon-encoded RNA expression induces cancer cell IFN-I signaling through mediating tumor-suppressive viral mimicry response, an anti-viral state triggered by endogenous stimuli (1). We found cancer cell ΔNp63 depletion results in increased retrotransposon expression triggering dsRNA sensing and downstream signaling activation; Interferon regulatory factor 1 plays a critical role in mediating viral mimicry response downstream of ΔNp63. We further showed cells with lower ΔNp63 level and higher IFN-I signaling activity (ΔNp63-depleted cell lines and ΔNp63lo PDOs) display stronger responses to Decitabine, an anti-cancer drug and viral mimicry booster (1), as compared to control cell lines or ΔNp63hi PDOs, respectively. Conclusion: We identified a novel function of ΔNp63 in repressing cancer retrotransposon expression and explored the therapeutic potential of enhancing viral mimicry response, which may guide future ΔNp63/viral mimicry response-targeted therapy. Acknowledgement: We acknowledge the Research Grants Council (TRS T12-701/17-R to MLL) and the Food and Health Bureau (HMRF 06171566 to VZY) of Hong Kong SAR for funding supports. We acknowledge DSMZ for the KYSE cell lines. We acknowledge the HKUMed-CPOS for providing imaging facilities. Reference: 1.Chiappinelli KB, et al. Inhibiting DNA Methylation Causes an Interferon Response in Cancer via dsRNA Including Endogenous Retroviruses. Cell 2015 Citation Format: Valen Zhuoyou Yu, Shan Shan So, Bryan Chee-Chad Lung, Carissa Wing-Yan Wong, Ian Yu-Hong Wong, Claudia Lai-Yin Wong, Desmond Kwan-Kit Chan, Fion Siu-Yin Chan, Betty Tsz-Ting Law, Ka-On Lam, Anthony Wing-Ip Lo, Josephine Mum-Yee Ko, Wei Dai, Alfred King-Yin Lam, Dora Lai-Wan Kwong, Simon Law, Maria Li Lung. p63 constrains cancer cell transposable element expressions and viral mimicry response to sustain esophageal cancer development and indicates therapeutic vulnerability [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2611.
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Heo, Yun-Jeong, In-Young Chung, Wan-Je Cho, Bo-Young Lee, Jung-Hoon Kim, Kyoung-Hee Choi, Jin-Won Lee, Daniel J. Hassett, and You-Hee Cho. "The Major Catalase Gene (katA) of Pseudomonas aeruginosa PA14 Is under both Positive and Negative Control of the Global Transactivator OxyR in Response to Hydrogen Peroxide." Journal of Bacteriology 192, no. 2 (November 20, 2009): 381–90. http://dx.doi.org/10.1128/jb.00980-09.
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ABSTRACT The adaptive response to hydrogen peroxide (H2O2) in Pseudomonas aeruginosa involves the major catalase, KatA, and OxyR. However, neither the molecular basis nor the relationship between the aforementioned proteins has been established. Here, we demonstrate that the transcriptional activation of the katA promoter (katAp) in response to H2O2 was abrogated in the P. aeruginosa PA14 oxyR null mutant. Promoter deletion analyses revealed that H2O2-mediated induction was dependent on a region of DNA −76 to −36 upstream of the H2O2-responsive transcriptional start site. This region harbored the potential operator sites (OxyR-responsive element [ORE]) of the Escherichia coli OxyR binding consensus. Deletion of the entire ORE not only abolished H2O2-mediated induction but also elevated the basal transcription, suggesting the involvement of OxyR and the ORE in both transcriptional activation and repression. OxyR bound to the ORE both in vivo and in vitro, demonstrating that OxyR directly regulates the katAp. Three distinct mobility species of oxidized OxyR were observed in response to 1 mM H2O2, as assessed by free thiol trapping using 4-acetamido-4′-maleimidylstilbene-2,2′-disulfonic acid. These oxidized species were not observed for the double mutants with mutations in the conserved cysteine (Cys) residues (C199 and C208). The uninduced transcription of katAp was elevated in an oxyR mutant with a mutation of Cys to serine at 199 (C199S) and even higher in the oxyR mutant with a mutation of Cys to alanine at 199 (C199A) but not in oxyR mutants with mutations in C208 (C208S and C208A). In both the C199S and the C208S mutant, however, katAp transcription was still induced by H2O2 treatment, unlike in the oxyR null mutant and the C199A mutant. The double mutants with mutations in both Cys residues (C199S C208S and C199A C208S) did not differ from the C199A mutant. Taken together, our results suggest that P. aeruginosa OxyR is a bona fide transcriptional regulator of the katA gene, sensing H2O2 based on the conserved Cys residues, involving more than one oxidation as well as activation state in vivo.