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 613 records · Page 34Linked to original sources
PubMed2003

Interfering with disease: opportunities and roadblocks to harnessing RNA interference.

RNA interference (RNAi) is an evolutionarily conserved mechanism for silencing gene expression by targeted degradation of mRNA. Short double-stranded RNAs, known as small interfering RNAs (siRNA), are incorporated into an RNA-induced silencing complex that directs degradation of RNA containing a homologous sequence. RNAi has been shown to work in mammalian cells, and can inhibit viral infection and control tumor cell growth in vitro. Recently, it has been shown that intravenous injection of siRNA or of plasmids expressing sequences processed to siRNA can protect mice from autoimmune and viral hepatitis. RNAi could provide an exciting new therapeutic modality for treating infection, cancer, neurodegenerative disease and other illnesses.

Animals↗
PubMed2007

Ribonucleic acid interference for neurological disorders: candidate diseases, potential targets, and current approaches.

OBJECTIVE: Ribonucleic acid (RNA) interference (RNAi) is a conserved evolutionary defense mechanism that is gaining utility for therapeutic application by modulating gene expression or silencing disease-causing genes. METHODS: This strategy has recently achieved success in mammalian cells via synthetic small interfering RNA or short hairpin RNA expressed in vectors for gene delivery. The vector-based RNAi strategy has particular potential because of the possibility of targeted gene delivery, long-term gene expression, and the potential means of penetrating the blood-brain barrier. RESULTS: RNAi-based approaches have been proposed for a variety of neurological disorders, including dominant genetic diseases, neurodegenerative diseases, malignant brain tumors, pain, and viral-induced encephalopathies. CONCLUSION: This review summarizes the current approaches of the RNAi strategy for neurological disorders, focusing on potential targets for therapeutic intervention.

Gene Targeting↗
PubMed2006

Silencing shrimp white spot syndrome virus (WSSV) genes by siRNA.

White spot syndrome virus (WSSV) is a major shrimp pathogen causing large economic losses all over the world. So far, however, there is no efficient approach to control this virus. RNA interference (RNAi), which has been applied to silence virus genes in eukaryotic organisms. In this investigation, a specific 21bp short interfering RNA (vp28-siRNA) targeting a major envelope protein gene (vp28) of WSSV was used to induce gene silencing in vivo in Penaeus japonicus shrimp. It was found that the transcription and expression of vp28 gene were silenced by the sequence-specific vp28-siRNA. However, the RNAi effect disappeared or significantly weakened even if one-nucleotide mutation existed in the vp28-siRNA. As revealed by quantitative PCR, the vp28-siRNA caused a significant reduction in viral DNA production of WSSV-infected shrimp. When treated with the vp28-siRNA, WSSV-infected shrimp had a reduced mortality rate. After three injections of the vp28-siRNA, the virus was completely eradicated from WSSV-infected shrimp. These findings suggest that RNAi is capable of silencing sequence-specific genes of WSSV and might constitute a new therapeutic strategy for WSSV infection in shrimp.

Animals↗
PubMed2005

Subcellular distribution of small interfering RNA: directed delivery through RNA polymerase III expression cassettes and localization by in situ hybridization.

Reduction in the expression of specific genes through small interfering RNAs (siRNAs) is dependent on the colocalization of siRNAs with other components of the RNA interference (RNAi) pathways within the cell. The expression of siRNAs within cells from cassettes that are derived from genes transcribed by RNA polymerase III (pol III) and provide for selective subcellular distribution of their products can be used to direct siRNAs to the cellular pathways. Expression from the human U6 promoter, resulting in siRNA accumulation in the nucleus, is effective in reducing gene expression, whereas cytoplasmic and nucleolar localization of the siRNA when expressed from the 5S or 7 SL promoters is not effective. The distribution of siRNA within the cell is determined by fluorescence in situ hybridization. Although the long uninterrupted duplex of siRNA makes it difficult to detect with DNA oligonucleotide probes, labeled oligonucleotide probes with 2'-O-methyl RNA backbones provide the stability needed for a strong signal. These methods contribute to studies of the interconnected cellular RNAi pathways and are useful in adapting RNAi as a tool to determine gene function and develop RNA-based therapeutics.

Base Sequence↗
PubMed2003

RNAi as a gene therapy approach.

RNA duplexes of 21 - 23 nucleotides (nts), with approximately 2 nt 3' overhangs (called small interfering RNAs or siRNAs), have recently been shown to mediate sequence-specific inhibition of gene expression in mammalian cells via a post-transcriptional gene silencing (PTGS) mechanism termed RNA interference (RNAi). RNAi has been rapidly adopted as a functional genomics tool in a wide range of species, has been adapted to allow for the transient or stable knockdown of gene expression generation in cell lines and animals, and has been developed for high-throughput analysis of gene function in Caenorhabditis elegans. With an increasing list of genes successfully knocked-down by RNAi in mammalian cells and improvements in the delivery of siRNAs to cells, including in vivo delivery to mice, attention is now turning to assessing the potential RNAi has as a gene therapy approach. RNAi is likely to have the greatest impact as a therapeutic tool in two key clinical areas, cancer and infectious disease, but it also has the potential as a therapy for other disorders including some dominant genetic diseases. This review will describe the status of the science behind this novel mechanism and will illustrate the therapeutic potential of RNAi-based technologies, using examples from these critical clinical research areas.

Animals↗
PubMed2025

Generation of Cdc20 RNAi-Sensitive Cell Lines to Study Mitotic Exit.

Accurate mitotic progression ensures the fidelity of genome passage. Cdc20 is a key mitotic regulator. It promotes mitotic exit by activating the anaphase-promoting complex or cyclosome (APC/C) and monitors kinetochore-microtubule attachment through activating the spindle assembly checkpoint (SAC). Precise characterization of Cdc20 requires efficient depletion of endogenous Cdc20, which is extremely difficult to achieve by RNA interference (RNAi). This chapter describes the methodology to generate Cdc20 RNAi-sensitive cell lines with the help of CRISPR/Cas9 technology. These cell lines are highly sensitive to Cdc20 RNAi and provide a very useful tool for Cdc20 functionality investigation without the interference of endogenous Cdc20 protein. Similar strategy could be applied to other genes.

Cdc20 Proteins↗
PubMed2006

Functional and selective RNA interference in developing axons and growth cones.

Developing axons and growth cones contain "local" mRNAs that are translated in response to various extracellular signaling molecules and have roles in several processes during axonal development, including axonal pathfinding, orientation of axons in chemotactic gradients, and in the regulation of neurotransmitter release. The molecular mechanisms that regulate mRNA translation within axons and growth cones are unknown. Here we show that proteins involved in RNA interference (RNAi), including argonaute-3 and argonaute-4, Dicer, and the fragile X mental retardation protein, are found in developing axons and growth cones. These proteins assemble into functional RNA-induced silencing complexes as transfection of small interfering RNAs selectively into distal axons results in distal axon-specific mRNA knock-down, without reducing transcript levels in proximal axons or associated diffusion of small interfering RNA into proximal axons or cell bodies. RhoA mRNA is localized to axons and growth cones, and intra-axonal translation of RhoA is required for growth cone collapse elicited by Semaphorin 3A (Sema3A), an axonal guidance cue. Selective knock-down of axonal RhoA mRNA abolishes Sema3A-dependent growth cone collapse. Our results demonstrate functional and potent RNAi in axons and identify an approach to spatially regulate mRNA transcripts at a subcellular level in neurons.

Animals↗
PubMed2006

A novel siRNA validation system for functional screening and identification of effective RNAi probes in mammalian cells.

Small interfering RNAs (siRNAs) have become the most powerful and widely used gene silencing reagents for reverse functional genomics and molecular therapeutics. The key challenge for achieving effective gene silencing in particular for the purpose of the therapeutics is primarily dependent on the effectiveness and specificity of the RNAi targeting sequence. However, only a limited number of siRNAs is capable of inducing highly effective and sequence-specific gene silencing by RNA interference (RNAi) mechanism. In addition, the efficacy of siRNA-induced gene silencing can only be experimentally measured based on inhibition of the target gene expression. Therefore, it is important to establish a fully robust and comparative validating system for determining the efficacy of designed siRNAs. In this study, we have developed a reliable and quantitative reporter-based siRNA validation system that consists of a short synthetic DNA fragment containing an RNAi targeting sequence of interest and two expression vectors for targeting reporter and triggering siRNA expression. The efficacy of the siRNAs is measured by their abilities to inhibit expression of the targeting reporter gene with easily quantified readouts including enhanced green fluorescence protein (EGFP) and firefly luciferase. Using fully analyzed siRNAs against human hepatitis B virus (HBV) surface antigen (HBsAg) and tumor suppressor protein p53, we have demonstrated that this system could effectively and faithfully report the efficacy of the corresponding siRNAs. In addition, we have further applied this system for screening and identification of the highly effective siRNAs that could specifically inhibit expression of mouse matrix metalloproteinase-7 (MMP-7), Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1), and human serine/threonine kinase AKT1. Since only a readily available short synthetic DNA fragment is needed for constructing this novel reporter-based siRNA validation system, this system not only provides a powerful strategy for screening highly effective siRNAs but also implicates in the use of RNAi for studying novel gene function in mammals.

Animals↗
PubMed2005

Molecular mechanisms of RNAi: implications for development and disease.

Research over the past few years has led to dramatic new discoveries on the role of double-stranded RNA (dsRNA) in the cell. RNA duplexes have been shown to orchestrate epigenetic changes, repress translation, and direct mRNA degradation in a sequence-specific manner. These diverse effects of dsRNA on gene expression have been termed RNA interference (RNAi). In addition to playing a role in viral defense and silencing transposons, RNAi also has a critical function in a number of developmental processes in the embryo. In this review, we explore these roles and discuss the molecular mechanisms behind dsRNA-mediated gene silencing. Further, we address the use of RNAi as a tool to study gene function in biology, and as a strategy for treating human disease.

Animals↗
PubMed2006

[Effect of silencing of hyaluronidase gene HYAL1 by RNA interference on proliferation of human breast cancer cells].

BACKGROUND & OBJECTIVE: It is reported that hyaluronidase is related to malignant potentiality of human breast cancer. This study was to investigate whether HYAL1 RNA interference (RNAi) could effectively inhibit gene mRNA expression as well as cell proliferation in human breast cancer cells. METHODS: Chemically synthesized double stranded RNA (dsRNA) targeting HYAL1 was transfected into human breast cancer cell lines MDA-MB-231, MDA-MB-453S, ZR-75 and ZR-75-30 using SiPORT lipid. The transfection efficiency was observed under a fluorescence confocal microscopy. Expression of HYAL1 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR). Cell proliferation and cell cycle were determined by MTT and flow cytometry assay, respectively. RESULTS: HYAL1 siRNA effectively inhibited HYAL1 mRNA expression (P<0.05), cell proliferation (P<0.05), and induced cell cycle arrest in G0/G1 phase with a significant decrease of cells in S-phase(P<0.05). CONCLUSION: HYAL1-siRNA may be used as a new approach in human breast cancer gene therapy.

Breast Neoplasms↗
PubMed2006

Short hairpin RNA-expressing bacteria elicit RNA interference in mammals.

RNA-interference (RNAi) is a potent mechanism, conserved from plants to humans for specific silencing of genes, which holds promise for functional genomics and gene-targeted therapies. Here we show that bacteria engineered to produce a short hairpin RNA (shRNA) targeting a mammalian gene induce trans-kingdom RNAi in vitro and in vivo. Nonpathogenic Escherichia coli were engineered to transcribe shRNAs from a plasmid containing the invasin gene Inv and the listeriolysin O gene HlyA, which encode two bacterial factors needed for successful transfer of the shRNAs into mammalian cells. Upon oral or intravenous administration, E. coli encoding shRNA against CTNNB1 (catenin beta-1) induce significant gene silencing in the intestinal epithelium and in human colon cancer xenografts in mice. These results provide an example of trans-kingdom RNAi in higher organisms and suggest the potential of bacteria-mediated RNAi for functional genomics, therapeutic target validation and development of clinically compatible RNAi-based therapies.

Animals↗
PubMedDate not supplied

RNA interference may be more potent than antisense RNA in human cancer cell lines.

1. RNA interference (RNAi) is a newly discovered cellular pathway for the silencing of sequence-specific genes at the mRNA level by the introduction of the cognate double-stranded (ds) RNA. Because antisense (AS) mechanisms have similar effects, we compared these two effects in human cancer cell lines, considering a possible application of RNAi for cancer therapy. 2. We tested RNAi effects by transfecting human hepatoma and pancreatic cancer cell lines with AS and sense (S) RNA expression plasmids corresponding to the exogenous luciferase gene or the endogenous c-raf gene in the form of complexes with a cationic lipopolyamine or a tumour-targeting peptide vector we developed. In addition, we compared the effects of small interfering RNA and AS oligoDNA complexed with the peptide vector. 3. From the viewpoint of AS actions, the effect of the AS RNA may be cancelled by the S RNA, although, interestingly, we found that the combination of the AS and S RNA expression plasmids was more effective than the AS RNA expression plasmids alone in reducing target gene expression, whereas the S RNA expression plasmids had no effects. The combination of the luciferase AS and S RNA had no effects on the expression of either the beta-galactosidase gene or the c-raf gene. In the presence of 2-aminopurine (an inhibitor of dsRNA-activated protein kinase), the inhibitory effect of the combination of AS and S RNA on gene expression did not change in the case of the endogenous c-raf gene, but was reduced in the case of the exogenous luciferase gene. The effect of 22 nucleotide RNA duplexes corresponding to the luciferase gene was by one order stronger than that of the phosphorothioate AS DNA. 4. Thus, it is suggested that RNAi may be more potent than AS RNA in reducing target gene expression in human cancer cell lines, regardless of the length of dsRNA. With further studies on the RNAi phenomenon in cancer cells, RNAi could provide a novel approach for cancer gene therapy.

2-Aminopurine↗
PubMed2004

Inhibition of cathepsin B and MMP-9 gene expression in glioblastoma cell line via RNA interference reduces tumor cell invasion, tumor growth and angiogenesis.

Extracellular proteases have been shown to cooperatively influence matrix degradation and tumor cell invasion through proteolytic cascades, with individual proteases having distinct roles in tumor growth, invasion, migration and angiogenesis. Matrix metalloproteases (MMP)-9 and cathepsin B have been shown to participate in the processes of tumor growth, vascularization and invasion of gliomas. In the present study, we used a cytomegalovirus promoter-driven DNA template approach to induce hairpin RNA (hpRNA)-triggered RNA interference (RNAi) to block MMP-9 and cathepsin B gene expression with a single construct. Transfection of a plasmid vector-expressing double-stranded RNA (dsRNA) for MMP-9 and cathepsin B significantly inhibited MMP-9 and cathepsin B expression and reduced the invasive behavior of SNB19, glioblastoma cell line in Matrigel and spheroid invasion models. Downregulation of MMP-9 and cathepsin B using RNAi in SNB19 cells reduced cell-cell interaction of human microvascular endothelial cells, resulting in the disruption of capillary network formation in both in vitro and in vivo models. Direct intratumoral injections of plasmid DNA expressing hpRNA for MMP-9 and cathepsin B significantly inhibited established glioma tumor growth and invasion in intracranial tumors in vivo. Further intraperitoneal (i.p.) injections of plasmid DNA expressing hpRNA for MMP-9 and cathepsin B completely regressed pre-established tumors for a long time (4 months) without any indication of these tumor cells. For the first time, these observations demonstrate that the simultaneous RNAi-mediated targeting of MMP-9 and cathepsin B has potential application for the treatment of human gliomas.

Animals↗
PubMed2005

Potential roles for short RNAs in lymphocytes.

RNA interference (RNAi) is an ancient and evolutionarily conserved process. In some eukaryotes, RNAi silences parasitic genetic elements. In plants, RNAi serves as an immune system against RNA viruses and transgenes and in worms, RNAi silences transposons. In mammals, RNAi has yet unknown functions. However, emerging roles for short RNAs and the factors that interact with them in other eukaryotes include chromatin modification, DNA deletion and DNA methylation, which may provide clues to the roles for short RNA function in mammals. For example, antigen receptor expression in lymphocytes is a highly regulated process and although much is known about chromatin modification and DNA deletion in the immune system, several molecular details of chromatin regulation remain elusive. This review compares emerging roles for short RNA function to processes required for antigen receptor expression in mammalian lymphocytes and predicts that short RNAs direct events required for successful lymphocyte-restricted gene expression.

Animals↗
PubMed2002

Exon-specific RNAi: a tool for dissecting the functional relevance of alternative splicing.

The goal of functional genomics is to determine the function of each protein encoded by an organism. Typically, this is done by inactivating individual genes and, subsequently, analyzing the phenotype of the modified organisms. In higher eukaryotes, where a tremendous amount of alternative splicing occurs, such approaches are not feasible because they have the potential to simultaneously affect multiple proteins that could have quite distinct and important functions. Thus, it is necessary to develop techniques that inactivate only a subset of proteins synthesized from genes encoding alternatively spliced mRNAs. Here we demonstrate that RNA interference (RNAi) can be used to selectively degrade specific alternatively spliced mRNA isoforms in cultured Drosophila cells. This is achieved by treating the cells with double-stranded RNA corresponding to an alternatively spliced exon. This technique may prove to be a powerful tool to assess the function of proteins synthesized from alternatively spliced mRNAs. In addition, these results have implications regarding the mechanism of RNAi in Drosophila.

Alternative Splicing↗
PubMed2006

siRNA in human cells selectively localizes to target RNA sites.

Recent observations of RNA interference (RNAi) in the nuclei of human cells raise key questions about the extent to which nuclear and cytoplasmic RNAi pathways are shared. By directly visualizing the localization of small interfering RNA (siRNA) in live human cells, we show here that siRNA either selectively localizes in the cytoplasm or translocates into the nucleus, depending on where the silencing target RNA resides. Two siRNAs that target the small nuclear 7SK and U6 RNAs localize into the nucleus as duplexes. In contrast, an siRNA targeting the cytoplasmic hepatitis C virus replicon RNA dissociates, and only antisense strand distributes in the cytoplasm of the cells harboring the target RNA, whereas sense strand gets degraded. At the same time, both strands of the latter siRNA are distributed throughout the cytoplasm and nucleus in cells lacking the silencing target RNA. These results suggest the existence of a mechanism by which the RNAi machinery orchestrates a target-determined localization of the siRNA and the corresponding RNAi activity, and also provide evidence for formation of nuclear-programmed active RNA induced silencing complexes directly in the nucleus.

Animals↗
PubMed2006

RNA interference of cytosolic leucine aminopeptidase reduces fecundity in the hard tick, Haemaphysalis longicornis.

Ticks are effective vectors of pathogens because of their blood feeding and high fecundity. This high fecundity is related to the size of the blood meal. Therefore, knowledge of how blood proteins are degraded and converted to proteins, including yolk protein, is important for the development of ways to inhibit the utilization of blood proteins by ticks. RNA interference (RNAi) is becoming a powerful post-transcriptional gene silencing technique that provides insight into gene function. We constructed a double-stranded RNA (dsRNA) based on a previously cloned Haemaphysalis longicornis leucine aminopeptidase (HlLAP) gene to reevaluate the biological role in tick blood digestion. Gene specific transcriptional, translational, and functional disruptions were achieved by the introduction of dsRNA into the ticks. Significantly delayed onset of egg-laying and reduced egg oviposition resulted from the RNAi for the HlLAP gene. These results suggest that HlLAP actually works as a blood digestive enzyme and affects tick fecundity via unknown mechanisms. The reduction of egg oviposition may be caused by a decrease in nutrients, especially free amino acids generated by HlLAP, from the blood meal. This is the first report of an impact on tick reproduction caused by gene silencing of a blood digestion-related molecule.

Animals↗
PubMed2006

Telomere elongation is under the control of the RNAi-based mechanism in the Drosophila germline.

Telomeres in Drosophila are maintained by transposition of specialized telomeric retroelements HeT-A, TAHRE, and TART instead of the short DNA repeats generated by telomerase in other eukaryotes. Here we implicate the RNA interference machinery in the control of Drosophila telomere length in ovaries. The abundance of telomeric retroelement transcripts is up-regulated owing to mutations in the spn-E and aub genes, encoding a putative RNA helicase and protein of the Argonaute family, respectively, which are related to the RNA interference (RNAi) machinery. These mutations cause an increase in the frequency of telomeric element retrotransposition to a broken chromosome end. spn-E mutations eliminate HeT-A and TART short RNAs in ovaries, suggesting an RNAi-based mechanism in the control of telomere maintenance in the Drosophila germline. Enhanced frequency of TART, but not HeT-A, attachments in individuals carrying one dose of mutant spn-E or aub alleles suggests that TART is a primary target of the RNAi machinery. At the same time, we detected enhanced HeT-A attachments to broken chromosome ends in oocytes from homozygous spn-E mutants. Double-stranded RNA (dsRNA)-mediated control of telomeric retroelement transposition may occur at premeiotic stages, resulting in the maintenance of appropriate telomere length in gamete precursors.

Adenosine Triphosphatases↗