Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “RNA interference (RNAi)”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 343 records · Page 19Linked to original sources
PubMed2004

Latrophilin is required for toxicity of black widow spider venom in Caenorhabditis elegans.

Black widow spider venom (BWSV) kills Caenorhabditis elegans after injection owing to the presence of heat- and detergent-sensitive components, which are high-molecular-mass latrotoxins. A C. elegans homologue of latrophilin/CIRL (calcium-independent receptor for latrotoxin), B0457.1, was identified and shown to have five conserved domains. RNAi (RNA interference) of this gene rendered C. elegans resistant to BWSV, whereas RNAi for CYP37A1 or a neurexin I homologue, and a deletion mutant of the related B0286.2 gene, had no effect on BWSV toxicity. The latrophilin RNAi mutants exhibit changes in defaecation cycle and alterations in drug sensitivity. These results demonstrate that latrophilin mediates the toxicity of BWSV and provide evidence for a physiological function of this receptor.

Amino Acid Sequence↗
PubMed2005

[Experimental strategies of the application of RNAi technique in mammalian cells].

As a novel and effective tool for gene function analysis, RNA interference (RNAi) technique has been developed rapidly and made great progress in mammalian cell studies. The mechanisms and experimental operation of RNAi in mammalian cells have become research hotspots, and RNAi mechanisms were supposed to be different from those in plant. In this paper, a review of RNAi mechanisms, the latest research progress and the experimental strategies of RNAi in mammalian cells were presented, including siRNA sequence design, siRNA delivery approaches and RNAi effect detection, so as to offer valuable references for RNAi studies.

Animals↗
PubMedDate not supplied

SiRNA drug delivery by biodegradable polymeric nanoparticles.

RNA interference (RNAi) is an emerging technology in which the introduction of double-stranded RNA (dsRNA) into a diverse range of organisms and cell types causes degradation of the complementary mRNA. It offers a broad spectrum of applications in both biological and medical research. Small interference RNA (siRNA) was recently explored for its therapeutical potential. However, the drug delivery of siRNA oligos is very novel and is in great need of future research. To this end, a biodegradable poly(D,L-lactide-co-glycolide) (PLGA) nanoparticle drug carrier system was prepared to load siRNA oligos with desired physicochemical properties. The nanoparticles were characterized by scanning electron microscopy and laser diffraction particle sizer. The delivery of siRNA into the targeted 293T cells was observed using fluorescent-labeled double-stranded Cy3-oligos. The model siRNA oligos, si-GFP-RNA, were also successfully loaded into PLGA nanoparticles and delivered in 293T cells. The gene silencing effect and the inhibition of GFP expression were investigated using fluorescent microscopy. Both positive and negative controls were used to compare with the new siRNA nanoparticle delivery system. It was found that nanoparticles offered both effective delivery of siRNA and prominent GFP gene silencing effect. Compared to conventional carrier systems, the new biodegradable polymeric nanoparticle system may also offer improved formulation stability, which is practically beneficial and may be used in the future clinical studies of siRNA therapeutics.

Absorbable Implants↗
PubMed2004

Genetic analysis of RNA-mediated transcriptional gene silencing.

The 'nuclear side' of RNA interference (RNAi) is increasingly recognized as an integral part of RNA-mediated gene silencing networks. Current data are consistent with the idea that epigenetic changes, such as DNA (cytosine-5) methylation and histone modifications, can be targeted to identical DNA sequences by short RNAs derived via Dicer cleavage of double-stranded RNA (dsRNA). To determine the relationships among RNA signals, DNA methylation and chromatin structure, we are carrying out a genetic analysis of RNA-mediated transcriptional gene silencing (TGS) in Arabidopsis. Results obtained so far indicate that in response to RNA signals, different site-specific DNA methyltransferases (DMTases) cooperate with each other and eventually with histone-modifying enzymes to establish and maintain a transcriptionally inactive state at a homologous target promoter. Processing of dsRNA in Arabidopsis occurs in the nucleus and in the cytoplasm, where distinct Dicer-like (DCL) activities are thought to generate functionally distinct classes of short RNAs. RNA silencing pathways thus operate throughout the cell to defend against invasive nucleic acids and to regulate genome structure and function.

Animals↗
PubMed2001

A role for the RNase III enzyme DCR-1 in RNA interference and germ line development in Caenorhabditis elegans.

An early event in RNA interference (RNAi) is the cleavage of the initiating double-stranded RNA (dsRNA) to short pieces, 21 to 23 nucleotides in length. Here we describe a null mutation in dicer-1 (dcr-1), a gene proposed to encode the enzyme that generates these short RNAs. We find that dcr-1(-/-) animals have defects in RNAi under some, but not all, conditions. Mutant animals have germ line defects that lead to sterility, suggesting that cleavage of dsRNA to short pieces is a requisite event in normal development.

Animals↗
PubMed2006

[Inhibitory effect of silencing hTERT gene by short hairpin RNA on growth of human laryngeal squamous cell carcinoma xenograft in nude mice].

BACKGROUND & OBJECTIVE: RNA interference (RNAi) is triggered by the presence of double-stranded RNA (dsRNA) in the cell and results in rapid destruction and post-transcriptional gene silencing of target mRNA. The roles of RNAi in gene function and antiviral therapy have been reported, but its effect on human telomerase reverse transcriptase (hTERT) gene, which is highly expressed in laryngeal squamous cell carcinoma and other head and neck neoplasms, has seldom been reported. This study was to explore the inhibitory effect of silencing hTERT gene by short hairpin RNA (shRNA) on growth of laryngeal squamous cell carcinoma xenograft in nude mice with RNAi technique. METHODS: Plasmid pshRNA containing fluorescein gene and hTERT cDNA sequences was synthesized. Human laryngeal squamous cell carcinoma Hep-2 cells were transplanted into nude mice to establish xenograft tumors. pshRNA was transfected into the tumors. The tumor volume was observed. Fluorescence expression in the tumors was observed by laser confocal microscopy. Tumor morphology was observed with HE staining. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. The expression of hTERT protein in the tumors was detected by SP immunohistochemistry. The structural change of heart, liver, kidney and spleen was observed, and peripheral blood hematological and biochemical parameters were determined. RESULTS: The tumor volume was significantly smaller in pshRNA group than in normal saline (NS) group and blank plasmid group (P<0.01). The inhibition rate was 76.50% in pshRNA group. After transfection of pshRNA or blank plasmid, many green fluorescent cells were observed under confocal microscope. The necrosis and apoptosis of tumor cells were found under light microscope in shRNA group. The apoptotic index of tumor cells was significantly higher in pshRNA group than in NS group and blank plasmid group [(26.47+/-4.25)% vs. (2.73+/-1.35)% and (3.40+/-1.41)%, P<0.05]. pshRNA directly down-regulated hTERT protein expression in the tumors, but did no damage to the heart, liver, kidney, spleen, and blood system of nude mice. CONCLUSION: shRNA plasmid containing specific sequences of hTERT gene could significantly inhibit the growth of laryngeal carcinoma in nude mice, with no side effect on the heart, liver, kidney, spleen, and blood system.

Animals↗
PubMed2006

["Disease targeting therapy" for neurosurgical disorders: molecular targeting and drug delivery].

Recent progress in molecular biology introduced a concept of molecular targeting therapy against many diseases including neurosurgical disorders. Molecular targeting therapy is promising, because targeting a disease-specific molecule it may provide cure without inducing unfavorable side effects. Focusing mainly on CNS (central nervous system) malignancies, authors discuss the current development of molecular targeting therapy with a special concern to strategies using DNA, RNA, immunotoxin, and molecular selective inhibitors. Strategies using DNA include gene therapy and DNA vaccine. RNA interference (RNAi) is one of the promising molecular targeting strategies using RNA. Immunotoxins such as recombinant chimeric cytotoxin consisting of interleukin-13 and a truncated exotoxin produced by the Pseudomonas aeruginosa bacterium is now under clinical investigation. Many molecular targeted agents such as selective inhibitors for protein kinases are now in preclinical and clinical development for patients with malignant glioma. Additionally, authors introduce their attempts to visualize the local drug delivery using real-time MRI monitoring. Visualization of drug delivery, while achieving effective drug distribution using convection-enhanced delivery (CED), may also contribute to the disease targeting. These strategies together should help us develop the effective 'disease-targeting therapy' for CNS disorders.

Biotechnology↗
PubMed2003

Activation of the interferon system by short-interfering RNAs.

RNA interference (RNAi) is a powerful tool used to manipulate gene expression or determine gene function. One technique of expressing the short double-stranded (ds) RNA intermediates required for interference in mammalian systems is the introduction of short-interfering (si) RNAs. Although RNAi strategies are reliant on a high degree of specificity, little attention has been given to the potential non-specific effects that might be induced. Here, we found that transfection of siRNAs results in interferon (IFN)-mediated activation of the Jak-Stat pathway and global upregulation of IFN-stimulated genes. This effect is mediated by the dsRNA-dependent protein kinase, PKR, which is activated by 21-base-pair (bp) siRNAs and required for upregulation of IFN-beta in response to siRNAs. In addition, we show by using cell lines deficient in specific components mediating IFN action that the RNAi mechanism itself is independent of the interferon system. Thus, siRNAs have broad and complicating effects beyond the selective silencing of target genes when introduced into cells. This is of critical importance, as siRNAs are currently being explored for their potential therapeutic use.

Animals↗
PubMed2004

Inhibition of herpesvirus-6B RNA replication by short interference RNAs.

RNA interference (RNAi) is a process of sequence-specific gene silencing, which is initiated by double-stranded RNA (dsRNA). RNAi may also serve as an antiviral system in vertebrates. This study describes the inhibition of herpesvirus-6B (HHV-6B) replication by short interference RNAs (siRNAs) that are targeted to the U38 sequence that encodes DNA polymerase. When virus-infected SupT1 cells were treated by siRNA, these cells blocked the cytopathic effect (CPE) and detected the HHV-6B antibody-negative in indirect immunofluorescence assays (IFA). Our result suggests that RNAi can efficiently block Herpesvirus-6B replication.

Animals↗
PubMed2006

Application of RNAi inducible TNFRI knockdown cells to the analysis of TNFalpha-induced cytotoxicity.

RNA interference (RNAi) has become a popular tool for downregulating in many species including mammalian cells. Therefore, suppression of target genes in mammalian cultured cells using RNAi may represent an ideal alternative to knockout studies for understanding the molecular mechanisms of chemical toxicity. Here, we assessed the potential of RNAi mediated gene knockdown in HeLa and HepG2 cells to cytotoxicity studies. Tumor necrosis factor receptor I (TNFRI) was chosen as a target gene because its signaling has been implicated in xenobiotic-induced toxicity. We optimized the design and performance of a vector-based RNAi experiment and then investigated viability of both HeLa and HepG2 cells exposed to TNFalpha. In addition, we examined gene expression profile of TNFRI knockdown HeLa cells after TNFalpha treatment, and then protein expression levels for several apoptosis-related genes of the cells. In both HeLa and HepG2 cells, TNFalpha exposure resulted in significantly reduced susceptibility of the knockdown cells to the cytotoxicity as compared with those of mock-transfected cells. Furthermore, the gene expression profiling and western blotting revealed that several genes including apoptosis and/or NF-kappaB pathway were downregulated in the knockdown HeLa cells. These results suggest that downregulation of the TNFRI gene in both HeLa and HepG2 cells by RNAi participates in resistance to TNFalpha-induced cytotoxicity. Therefore, this study raises the possibility that RNAi-based gene silencing in mammalian cells may be a valuable tool for elucidating the relationships between phenotypic changes and target gene functions in response to xenobiotic-induced cytotoxicity. Further exposure study using xenobiotics needs to be done to validate the potential utility of RNAi technology.

Bleomycin↗
PubMedDate not supplied

[RNA binding proteins in the RNA interference phenomenon].

The ability of short RNAs (21-27 nucleotides) to silence genes containing homologous nucleotide sequences is related to RNA silencing. The pathways of short RNAs (siRNA and microRNA) biogenesis from their precursors, double stranded and hairpin RNAs respectively, are briefly reviewed. The functioning of specific RNA binding domains found for the first time in the proteins operating in RNA interference (RNAi) is considered. The interactions of these domains with the earlier well known RNA binding modules in RNAi proteins are described.

Animals↗
PubMed2005

Individualised cancer therapeutics: dream or reality? Therapeutics construction.

The analysis of DNA microarray and proteomic data, and the subsequent integration into functional expression sets, provides a circuit map of the hierarchical cellular networks responsible for sustaining the viability and environmental competitiveness of cancer cells, that is, their robust systematics. These technologies can be used to 'snapshot' the unique patterns of molecular derangements and modified interactions in cancer, and allow for strategic selection of therapeutics that best match the individual profile of the tumour. This review highlights technology that can be used to selectively disrupt critical molecular targets and describes possible vehicles to deliver the synthesised molecular therapeutics to the relevant cellular compartments of the malignant cells. RNA interference (RNAi) involves a group of evolutionarily conserved gene silencing mechanisms in which small sequences of double-stranded RNA or intrinsic antisense RNA trigger mRNA cleavage or translational repression, respectively. Although RNAi molecules can be synthesised to 'silence' virtually any gene, even if upregulated, a mechanism for selective delivery of RNAi effectors to sites of malignant disease remains challenging. The authors will discuss gene-modified conditionally replicating viruses as candidate vehicles for the delivery of RNAi.

Animals↗
PubMed2004

Short interfering RNA-mediated inhibition of herpes simplex virus type 1 gene expression and function during infection of human keratinocytes.

RNA interference (RNAi) is an antiviral mechanism that is activated when double-stranded RNA is cleaved into fragments, called short interfering RNA (siRNA), that prime an inducible gene silencing enzyme complex. We applied RNAi against a herpes simplex virus type 1 (HSV-1) gene, glycoprotein E, which mediates cell-to-cell spread and immune evasion. In an in vitro model of infection, human keratinocytes were transfected with siRNA specific for glycoprotein E and then infected with wild-type HSV-1. RNAi-mediated gene silencing reproduced the small plaque phenotype of a gE-deletion mutant virus. The specificity of gene targeting was demonstrated by flow cytometry and Northern blot analyses. Exogenous siRNA can suppress HSV-1 glycoprotein E expression and function during active infection in vitro through RNAi. This work establishes RNAi as a genetic tool for the study of HSV and provides a foundation for development of RNAi as a novel antiviral therapy.

Blotting, Northern↗
PubMed2003

Small RNA: can RNA interference be exploited for therapy?

CONTEXT: RNA interference (RNAi) is the sequence-specific gene-silencing induced by double-stranded RNA (dsRNA), and gives information about gene function quickly, easily, and inexpensively. The use of RNAi for genetic-based therapies is widely studied, especially in viral infections, cancers, and inherited genetic disorders. RNAi has been used to make tissue-specific knockdown mice for studying gene function in a whole animal. Combined with genomics data, RNAi-directed gene-silencing could allow functional determination of any gene expressed in a cell or pathway. The term RNAi came from the discovery that the injection of dsRNAs into Caenorhabditis elegans interferes with the expression of specific genes containing a complementary region to the delivered dsRNA. Although stalled for a time by the non-gene-specific interferon response elicited by dsRNA molecules longer than about 30 nucleotides in mammalian cells, Tom Tuschl's group found that transfection of synthetic 21-nucleotide small-interfering RNA (siRNA) duplexes were highly selective and sequence-specific inhibitors of endogenous genes. STARTING POINT: siRNA expression has been studied with siRNA from plasmid and viral vectors that efficiently deliver siRNAs into both dividing and non-dividing cells, stem cells, zygotes, and their differentiated progeny. A collection of RNA interference vectors that suppress 50 human de-ubiquitinating enzymes allowed Thijn Brummelkamp and colleagues to study this gene family and to identify de-ubiquitinating enzymes in cancer-relevant pathways (Nature 2003; 424: 797-801). These researchers found that the familial cylindromatosis tumour suppressor gene (CYLD), previously of unknown function, could enhance the activation of the transcription factor NF-kappaB, leading to increased resistance to apoptosis. They have now started to investigate the use of CYLD inhibitors in clinical trials. WHERE NEXT: The ability to efficiently and stably produce and deliver sufficient amounts of siRNA to the proper target tissues require refinement before this new technology can be tried clinically. Initial in-vivo studies reported effective transgene suppression in adult mice by chemically synthesised siRNAs. More recently many researchers have used plasmid and viral vectors for transcription of short-hairpin RNAs, both in vitro and in vivo. With these expression systems, gene expression was more stably inhibited than with the transient knockdown recorded with chemically synthesised siRNA. Human trials exploiting these latest findings are likely to soon follow.

Animals↗
PubMed2006

RNAi-mediated gene silencing in non-human primates.

The opportunity to harness the RNA interference (RNAi) pathway to silence disease-causing genes holds great promise for the development of therapeutics directed against targets that are otherwise not addressable with current medicines. Although there are numerous examples of in vivo silencing of target genes after local delivery of small interfering RNAs (siRNAs), there remain only a few reports of RNAi-mediated silencing in response to systemic delivery of siRNA, and there are no reports of systemic efficacy in non-rodent species. Here we show that siRNAs, when delivered systemically in a liposomal formulation, can silence the disease target apolipoprotein B (ApoB) in non-human primates. APOB-specific siRNAs were encapsulated in stable nucleic acid lipid particles (SNALP) and administered by intravenous injection to cynomolgus monkeys at doses of 1 or 2.5 mg kg(-1). A single siRNA injection resulted in dose-dependent silencing of APOB messenger RNA expression in the liver 48 h after administration, with maximal silencing of >90%. This silencing effect occurred as a result of APOB mRNA cleavage at precisely the site predicted for the RNAi mechanism. Significant reductions in ApoB protein, serum cholesterol and low-density lipoprotein levels were observed as early as 24 h after treatment and lasted for 11 days at the highest siRNA dose, thus demonstrating an immediate, potent and lasting biological effect of siRNA treatment. Our findings show clinically relevant RNAi-mediated gene silencing in non-human primates, supporting RNAi therapeutics as a potential new class of drugs.

Animals↗
PubMed2006

3' UTR seed matches, but not overall identity, are associated with RNAi off-targets.

Off-target gene silencing can present a notable challenge in the interpretation of data from large-scale RNA interference (RNAi) screens. We performed a detailed analysis of off-targeted genes identified by expression profiling of human cells transfected with small interfering RNA (siRNA). Contrary to common assumption, analysis of the subsequent off-target gene database showed that overall identity makes little or no contribution to determining whether the expression of a particular gene will be affected by a given siRNA, except for near-perfect matches. Instead, off-targeting is associated with the presence of one or more perfect 3' untranslated region (UTR) matches with the hexamer or heptamer seed region (positions 2-7 or 2-8) of the antisense strand of the siRNA. These findings have strong implications for future siRNA design and the application of RNAi in high-throughput screening and therapeutic development.

3' Untranslated Regions↗
PubMed2004

RNAi knock-down mice: an emerging technology for post-genomic functional genetics.

RNA interference (RNAi) has been extensively used for sequence-specific silencing of gene function in mammalian cells. The latest major breakthrough in the application of RNAi technology came from experiments demonstrating RNAi-mediated gene repression in mice and rats. After more than two decades of functional mouse research aimed at developing and continuously improving transgenic and knock-out technology, the advent of RNAi knock-down mice represents a valuable new alternative for studying gene function in vivo. In this review we provide some basic insight as to how RNAi can induce gene silencing to then focus on recent findings concerning the applicability of RNAi for regulating gene function in the mouse. Reviewed topics will include delivery methods for RNAi-mediating molecules, a comparison between traditional knock-out and innovative transgenic RNAi technology and the generation of graded RNAi knock-down phenotypes. Apart from the exciting possibilities RNAi provides for studying gene function in mice, we discuss several caveats and limitations to be considered. Finally, we present prospective strategies as to how RNAi technology might be applied for generating conditional and tissue-restricted knock-down mice.

Alleles↗
PubMed2003

Protection from HIV-1 infection of primary CD4 T cells by CCR5 silencing is effective for the full spectrum of CCR5 expression.

Stable gene silencing by RNA interference (RNAi) can be achieved by expression of small hairpin RNAs (shRNAs) from RNA polymerase III promoters. We have tested lentiviral vectors expressing shRNAs targetting CCR5 in primary CD4 T cells from donors representing various CCR5 and CCR2 genetic backgrounds covering the full spectrum of CCR5 expression levels and permissiveness for HIV-1 infection. A linear decrease in CCR5 expression resulted in a logarithmic decrease in cellular infection, giving up to three logs protection from HIV-1 infection in vitro. Protection was maintained at very high multiplicity of infection. This and other recent reports on RNAi should open a debate about the use of RNAi gene therapy for HIV infection.

CD4-Positive T-Lymphocytes↗