Результаты поиска

A single amino acid substitution (157 Gly----Val) in a phosphoglycerate kinase variant (PGK Shizuoka) associated with chronic hemolysis and myoglobinuria.

We have determined a single amino acid substitution in a new phosphoglycerate kinase (PGK) variant, PGK Shizuoka, associated with chronic hemolysis and myoglobinuria. PGK Shizuoka had an extremely low enzyme activity with normal kinetic properties and normal electrophoretic mobility. Total blood cell RNA of the patient was reverse-transcribed and amplified by the polymerase chain reaction. A single nucleotide substitution from guanine to thymine at position 473 of PGK messenger RNA was found. This nucleotide change causes a single amino acid substitution from Gly to Val at the 157th position,

DNA Distortion by the Burkholderia thailandensis Transcriptional Regulator FusR2 Is Abolished by Fusaric Acid Binding.

Fusaric acid is a toxic metabolite produced by several Fusarium species, which causes wilt disease in many plants. Among other functions, fusaric acid allows the fungus to outcompete soil bacteria. Understanding mechanisms by which bacteria mitigate the toxic effects of fusaric acid is therefore of interest in terms of controlling Fusarium wilt. The soil bacterium Burkholderia thailandensis encodes a predicted fusaric acid-binding membrane transporter belonging to the FusC2 family. The fusC2 gene is annotated as part of an operon, also encoding an isochorismatase (gene named isoC) and a member

Differential Gene Regulation of the Human Blastocyst Trophectoderm and Inner Cell Mass by Progesterone.

Knowledge of action of progesterone (P4) on the human preimplantation embryo is lacking. The objective of this study was to determine expression of a mitochondrial P4 receptor (PR-M) in the trophectoderm (TE) and the inner cell mass (ICM) of the human blastocyst and to determine P4-induced gene expression during growth from the cleavage to the blastocyst stage. Previously cryopreserved cleavage stage embryos were treated with P4 (10-6 M) or vehicle until blastocyst development. Cells from the TE and the ICM of dissected euploid embryos underwent RNA-seq analysis, while other embryos were used

3-O-trans-p-coumaroyl-alphitolic acid, a triterpenoid from Zizyphus jujuba, leads to apoptotic cell death in human leukemia cells through reactive oxygen species production and activation of the unfolded protein response.

3-O-trans-p-coumaroyl-alphitolic acid (3OTPCA), a triterpenoid isolated from the plant Zizyphus jujuba (ZJ), is known to be cytotoxic to cancer cells; however, the molecular mechanism underlying 3OTPCA-induced cell death remains unknown. Here, we provide novel evidence that 3OTPCA induces apoptotic cell death in human leukemia cells. We found that 3OPTCA induces DNA fragmentation within 24 h after treatment in U937 cells, which was also observed in other leukemia cell lines, including Molt-4 and Jurkat cells. We then investigated other parameters involved in apoptosis, including phosphatidylse

The amino-terminus of the hepatitis C virus (HCV) p7 viroporin and its cleavage from glycoprotein E2-p7 precursor determine specific infectivity and secretion levels of HCV particle types.

Viroporins are small transmembrane proteins with ion channel activities modulating properties of intracellular membranes that have diverse proviral functions. Hepatitis C virus (HCV) encodes a viroporin, p7, acting during assembly, envelopment and secretion of viral particles (VP). HCV p7 is released from the viral polyprotein through cleavage at E2-p7 and p7-NS2 junctions by signal peptidase, but also exists as an E2p7 precursor, of poorly defined properties. Here, we found that ectopic p7 expression in HCVcc-infected cells reduced secretion of particle-associated E2 glycoproteins. Using bioc

Anamorsin, a novel caspase-3 substrate in neurodegeneration.

Activated caspases play a central role in the execution of apoptosis by cleaving endogenous substrates. Here, we developed a high throughput screening method to identify novel substrates for caspase-3 in a neuronal cell line. Critical steps in our strategy consist of two-dimensional electrophoresis-based protein separation and in vitro caspase-3 incubation of immobilized proteins to sort out direct substrates. Among 46 putative substrates identified in MN9D neuronal cells, we further evaluated whether caspase-3-mediated cleavage of anamorsin, a recently recognized cell death-defying factor in

CRISPR-AsCas12a Efficiently Corrects a GPR143 Intronic Mutation in Induced Pluripotent Stem Cells from an Ocular Albinism Patient.

Mutations in the GPR143 gene cause X-linked ocular albinism type 1 (OA1), a disease that severely impairs vision. We recently generated induced pluripotent stem cells (iPSCs) from skin fibroblasts of an OA1 patient carrying a point mutation in intron 7 of GPR143. This mutation activates a new splice site causing the incorporation of a pseudoexon. In this study, we present a high-performance CRISPR-Cas ribonucleoprotein strategy to permanently correct the GPR143 mutation in these patient-derived iPSCs. Interestingly, the two single-guide RNAs available for SpCas9 did not allow the cleavage of t

Shared-Output Aptamer-Gated DNA Nanodevices Program NbaSPARDA Trans-Cleavage for Metabolite and Drug Sensing.

Routing chemically distinct small-molecule aptamers into a common nuclease trans-cleavage pathway is constrained by the target-specific output handles of conventional structure-switching aptamers (SSAs). Shared-output aptamer-gated DNA nanodevices are designed so that recognition cores control release of the same short trigger DNA (tDNA). The released tDNA hybridizes with a common guide RNA and activates NbaSPARDA, a short prokaryotic Argonaute-associated nuclease complex, for reporter trans-cleavage. Analyses across two sequence scaffolds reveal sequence-dependent activation and identify a pr

Entropy-driven reactions for controlling CRISPR/Cas12a and constructing an electrochemical biosensor for cardiac biomarkers detection.

An electrochemical biosensor is reported for controlling CRISPR/Cas12a activity through the utilization of entropy-driven reactions, alongside the construction of a highly sensitive biosensor for B-type natriuretic peptide (BNP) detection. In the biosensor, entropy-driven reactions are employed to regulate the activity of CRISPR/Cas12a - a gene editing tool - capable of nonspecific cleavage of single-stranded DNA (ssDNA). The biosensor architecture encompasses an electrode that is modified with ssDNA probes designed to hybridize with target BNP aptamers. These aptamers, furnished with labeled

mRNA-encoded Cas13 can be used to treat dengue infections in mice.

Dengue is a major global health threat, and there are no approved antiviral agents. Prior research using Cas13 only demonstrated dengue mitigation in vitro. Here we demonstrate that systemic delivery of mRNA-encoded Cas13a and guide RNAs formulated in lipid nanoparticles can be used to treat dengue virus (DENV) 2 and 3 in mice. First, we identified guides against DENV 2 and 3 that demonstrated in vitro efficacy. Next, we confirmed that Cas13 enzymatic activity is necessary for DENV 2 or DENV 3 mitigation in vitro. Last, we show that a single dose of lipid-nanoparticle-formulated mRNA-encoded C

Targeting bacterial TPP riboswitches using pVEC-conjugated chimeric antisense oligonucleotides.

Antisense oligonucleotides (ASOs) provide a sequence-specific approach for targeting bacterial RNAs and suppressing the expression of genes … , preparation of an in vitro transcribed TPP aptamer RNA substrate, and assessment of ASO-dependent RNA cleavage using an RNase H assay. The protocol further describes bacterial growth-inhibition assays in Listeria monocytogenes and Bacillus subtilis, the use of Escherichia coli as a specificity control, and evaluation of pVEC-ASO-1 cytotoxicity in A549 human …

Alternative poly(A) site utilization during adenovirus infection coincides with a decrease in the activity of a poly(A) site processing factor.

The recognition and processing of a pre-mRNA to create a poly(A) addition site, a necessary step in mRNA biogenesis, can also be a regulatory event in instances in which the frequency of use of a poly(A) site varies. One such case is found during the course of an adenovirus infection. Five poly(A) sites are utilized within the major late transcription unit to produce more than 20 distinct mRNAs during the late phase of infection. The proximal half of the major late transcription unit is also expressed during the early phase of a viral infection. During this early phase of expression, the L1 po

RNabel─A Standalone Software Tool for Annotating Tandem Mass Spectra of Modified Ribonucleic Acids.

Ribonucleic acid (RNA) modifications, with over 170 identified types, play diverse roles in cellular processes. The past decade has witnessed a surging demand for accurate identification and localization of RNA modifications in both endogenous and synthetic therapeutic RNAs. With accurate spectral annotation for RNA, tandem mass spectrometry (MS/MS) can meet this demand. Here, we present RNabel, a user-friendly software tool for in-depth annotation of MS/MS spectra of RNA oligonucleotides. RNabel considers a full set of backbone-cleavage ions (a, b, c, d, a-B, w, x, y, z) in which the ribonucl

Hsc70 regulates the IRES activity and serves as an antiviral target of enterovirus A71 infection.

Enterovirus A71 (EV-A71) is a small positive-stranded RNA virus that causes human hand, foot and mouth disease (HFMD) and fatal neurological disorders in some cases without effective treatment. Here we show that heat shock cognate protein 70 (Hsc70), a molecular chaperone, displays pivotal role in viral infections. Knockdown of Hsc70 significantly suppresses viral replication evidenced by reducing not only the level of both viral replication intermediates (negative stranded RNA) and viral genomic RNA (positive stranded RNA), but also the level of viral protein expression; whereas ectopic expre

Perfluorooctanoic acid inhibits androgen biosynthesis in rat immature Leydig cells.

Perfluorooctanoic acid (PFOA) is a commonly used short-chain synthetic perfluoroalkyl agent. Immature Leydig cells (ILCs) are localized in the testis and responsible for androgen biosynthesis and metabolism. Although PFOA shows toxicity in the reproductive system, it is not clear if it disrupts the function of ILCs. In the present study, primary ILCs were isolated from 35-day-old rats and exposed to a range of PFOA concentrations (0, 0.01, 0.1, or 1 μM). It was determined that 0.1 or 1 μM PFOA reduced total androgen biosynthesis in ILCs. Specifically, 22R-hydroxycholesterol (22R), and pregneno

CRISPR-Cas and catalytic hairpin assembly technology for target-initiated amplification detection of pancreatic cancer specific tsRNAs.

Transfer RNA-derived small RNAs (tsRNAs) tRF-LeuCAG-002 (ts3011a RNA) is a novel class of non-coding RNAs biomarker for pancreatic cancer (PC). Reverse transcription polymerase chain reaction (RT-qPCR) has been unfit for community hospitals that are short of specialized equipment or laboratory setups. It has not been reported whether isothermal technology can be used for detection, because the tsRNAs have rich modifications and secondary structures compared with other non-coding RNAs. Herein, we have employed a catalytic hairpin assembly (CHA) circuit and clustered regularly interspaced short

Антивирусная эффективность японского вируса энцефалита (RNase H-Depent Gapmer Antiense Oligonucleotides).

Зависимые от RNase H антисенсы олигонуклеотиды (ASO) представляют собой перспективный терапевтический подход на основе конкретной последовательности, связывающей и унижающей целевые РНК. Однако эффективность и механизм противовирусных распылителей АСО остаются неясными. Здесь мы изучили ингибиционное воздействие японского энцефалитного вируса (JEV), содержащего запертые нуклеиновые кислоты (LNA displamers) на распространение флавивируса, переносимого комарами. Мы разработали несколько лазеек LNA, направленных на 3' непереведенные области JEV геномных РНК. Проверка in vitro с помощью табличного

Совпадение в аннотации оригинала
RNase H-dependent gapmer antisense oligonucleotides (ASOs) are a promising therapeutic approach via sequence-specific binding to and degradi … that LNA gapmers targeting a stem-loop region effectively inhibit JEV proliferation. Cell-based and RNA cleavage assays using mismatched LNA gapmers exhibited an underlying mechanism where the inhibition of viral production results from JEV RNA degradation by LNA gapmers in a sequence- and modification-dependent manner. Encouragingly, LNA gapmers potently inhi …

Обнаружение геномных последовательности вируса кори в мозговой ткани SSPE по цепной реакции полимеразы.

В качестве предварительного шага была изменена цепная реакция полимеразы (PCR) для обнаружения геномных последовательностей RNA путем создания копий этих последовательностей CDNA. Были синтезированы пары олигонуклеотидов, дополняющие последовательности в каждом из пяти основных структурных генов белка вируса кори (белок нуклеокапсид, фосфопротеин, матрицовый протеин, белок синтеза и протеин гемагглютинина). Продукты ПЦР были в предварительном порядке идентифицированы путем визуализации полос соответствующего размера путем пятнаэтидиумного метила после геля электрофореза, и их идентичность была

CRISPR technologies for detecting DNA and RNA methylation: Mechanisms, platforms, and translational opportunities.

DNA and RNA methylation are key epigenetic and epitranscriptomic modifications involved in gene regulation, genome stability, RNA metabolism, and disease progression. Aberrant methylation patterns in cell-free DNA and RNA have emerged as valuable biomarkers for cancer detection, disease monitoring, and therapeutic stratification. However, conventional methods such as bisulfite sequencing, methylation-specific PCR, MeRIP-seq, SCARLET, and LC-MS/MS often require harsh processing, high sample input, complex instrumentation, or lack site-specific resolution, limiting their clinical and point-of-ca

Potential Applications of RNase P Ribozyme Against Hepatitis B Virus.

Nucleic acid-based gene-interfering molecules, such as antisense oligonucleotides, ribozymes, and small interfering RNA (siRNA), represent exciting gene-targeting agents for therapeutic applications. RNase P ribozymes derived from M1 RNA, the catalytic RNA subunit of RNase P in Escherichia coli, have shown great promise as a novel nucleic acid-based gene interference approach to modulate gene expression. When M1 RNA is covalently linked to a guide sequence (GS), it can be engineered into a sequence-specific endonuclease M1GS ribozyme, which can hydrolyze any mRNA that base-pairs with the guide