Many commonly used siRNAs risk off-target activity

Snøve, Ola; Holen, Torgeir

doi:10.1016/j.bbrc.2004.04.175

Cited by 131 publications

(88 citation statements)

References 46 publications

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“…Off-targeting effects surfaced as a major issue in RNAi experiments when the effects of RNAi treatment were systematically analyzed Scacheri et al, 2004;Snove and Holen, 2004;Lin et al, 2005;Fedorov et al, 2006). One of the most revealing data came from mammalian cells transfected with different siRNAs targeting the same gene, which were systematically analyzed using microarrays The present document has been produced and adopted by the bodies identified above as authors.…”

Section: Off-targeting -Causes and Remediesmentioning

confidence: 99%

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

Pačes

Nič²,

Novotný³

et al. 2017

EFSA Supporting Publications

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This report is the outcome of an EFSA procurement aiming at investigating and summarising the state of knowledge on (I) the mode-of-action of dsRNA and miRNA pathways, (II) the potential for nontarget gene regulation by dsRNA-derived siRNAs or miRNAs, (III) the determination of siRNA pools in plant tissues and the importance of individual siRNAs for silencing. The report is based on a comprehensive and systematic literature search, starting with the identification and retrieval of~190,000 publications related to the research area and further filtered down with keywords to produce focused collections used for subsequent screening of titles and abstracts. The report is comprised of an (I) Introduction to the field of small RNAs, (II) a Data and Methodologies section containing strategies used for literature search and study selection, and (III) the Results of the literature review organized according to the three main procurement tasks. The outcome of the first task reviews dsRNA and miRNA pathways in mammals (including humans), birds, fish, arthropods, annelids, molluscs, nematodes, and plants. Eight taxon-dedicated chapters are based on~1,400 cumulative references chosen from~10,000 inspected titles and abstracts. We review conserved and divergent aspects of small RNA pathways and dsRNA responses in animals and plants including structure and function of key proteins as well as four basic mechanisms: genome-encoded posttranscriptional regulations (miRNA), degradation of RNAs by short interfering RNA pools generated from long dsRNA (RNAi), transcriptional silencing, and sequence-independent responses to dsRNA. The outcome of the second task focuses on base pairing between small RNAs and their target RNAs and predictability of biological effects of small RNAs in animals and plants. The outcome of the last task reviews methodology, siRNA pools, and movement of small RNAs in plants. Potential transfer of small RNAs between species and circulating miRNAs in mammals is described in the final chapter.

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Section: Off-targeting -Causes and Remediesmentioning

confidence: 99%

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

Pačes

Nič²,

Novotný³

et al. 2017

EFSA Supporting Publications

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“…The specificity of RNAi-mediated gene silencing has been challenged by several studies (Jackson et al, 2003;Snove and Holen, 2004). To ascertain the specificity of esiRNA effects in our study, we next performed rescue experiments.…”

Section: Ectopic Expression Of Mklp1 Rescues Cytokinesis Defect In Mkmentioning

confidence: 99%

Functional Analysis of Human Microtubule-based Motor Proteins, the Kinesins and Dyneins, in Mitosis/Cytokinesis Using RNA Interference

Zhu

Zhao

Bibikova

et al. 2005

MBoC

369

337

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Microtubule (MT)-based motor proteins, kinesins and dyneins, play important roles in multiple cellular processes including cell division. In this study, we describe the generation and use of an Escherichia coli RNase III-prepared human kinesin/dynein esiRNA library to systematically analyze the functions of all human kinesin/dynein MT motor proteins. Our results indicate that at least 12 kinesins are involved in mitosis and cytokinesis. Eg5 (a member of the kinesin-5 family), Kif2A (a member of the kinesin-13 family), and KifC1 (a member of the kinesin-14 family) are crucial for spindle formation; KifC1, MCAK (a member of the kinesin-13 family), CENP-E (a member of the kinesin-7 family), Kif14 (a member of the kinesin-3 family), Kif18 (a member of the kinesin-8 family), and Kid (a member of the kinesin-10 family) are required for chromosome congression and alignment; Kif4A and Kif4B (members of the kinesin-4 family) have roles in anaphase spindle dynamics; and Kif4A, Kif4B, MKLP1, and MKLP2 (members of the kinesin-6 family) are essential for cytokinesis. Using immunofluorescence analysis, time-lapse microscopy, and rescue experiments, we investigate the roles of these 12 kinesins in detail. INTRODUCTIONThe mitotic spindle, a unique cellular apparatus assembled at the beginning of mitosis, plays a pivotal role in regulating mitosis and cytokinesis. Although tubulins are the most abundant protein in the mitotic spindle, many additional proteins are involved in regulating its formation and function. Most prominent among these are members of the kinesin and dynein families of MT-based motor proteins (Mandelkow and Mandelkow, 2002;Vale, 2003), which generate directional movement along microtubules. The kinesin proteins share a conserved, ϳ340 amino acid, motor domain that utilizes ATP to fuel their movement along MTs. Outside the motor domain, kinesins also contain different "stalk" and "tail" domains that mediate oligomerization, regulation of motor activity, and interactions with their specific cargos (Vale, 2003).The roles of kinesins in mitosis and cytokinesis have been extensively studied in Saccharomyces cerevisiae, in which there are six kinesin genes belonging to five subfamilies. Gene disruption experiments have demonstrated that five of these kinesins play essential roles in regulating the formation, orientation, and elongation of the mitotic spindle and the segregation of chromosomes in mitosis (Hildebrandt and Hoyt, 2000). In addition, one cytoplasmic dynein motorcontaining polypeptide, the dynein heavy chain (DHC), has also been implicated in mitotic spindle function (Hildebrandt and Hoyt, 2000). Using RNAi techniques, Goshima and Vale recently examined the mitotic functions of cytoplasmic dynein and all 25 fly kinesins in cultured Drosophila cells (Goshima and Vale, 2003). They found that four kinesins are needed for bipolar spindle assembly and four are crucial for metaphase chromosome alignment. Dynein plays a role in the metaphase-to-anaphase transition, and one kinesin is required for cytokinesis.Th...

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“…The vector control clonal cell line demonstrated no reduction in RTN3 levels compared with the parental HeLa cell (lane 1 versus lane 2). The siRNA technique sometimes introduces nonspecific off-target gene knockdowns (33,34). This knowledge prompted us to reintroduce the RTN3 gene into the knockdown cell line.…”

Section: Rtn3 Is Required and Sufficient For Immediate Early Virus Rementioning

confidence: 99%

Reticulon 3 Binds the 2C Protein of Enterovirus 71 and Is Required for Viral Replication

Tang¹,

Yang²,

Wu³

et al. 2007

Journal of Biological Chemistry

131

119

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Enterovirus 71 is an enterovirus of the family Picornaviridae. The 2C protein of poliovirus, a relative of enterovirus 71, is essential for viral replication. The poliovirus 2C protein is associated with host membrane vesicles, which form viral replication complexes where viral RNA synthesis takes place. We have now identified a host-encoded 2C binding protein called reticulon 3, which we found to be associated with the replication complex through direct interaction with the enterovirus 71-encoded 2C protein. We observed that the N terminus of the 2C protein, which has both RNA-and membrane-binding activity, interacted with reticulon 3. This region of interaction was mapped to its reticulon homology domain, whereas that of 2C was encoded by the 25th amino acid, isoleucine. Reticulon 3 could also interact with the 2C proteins encoded by other enteroviruses, such as poliovirus and coxsackievirus A16, implying that it is a common factor for such viral replication. Reduced production of reticulon 3 by RNA interference markedly reduced the synthesis of enterovirus 71-encoded viral proteins and replicative doublestranded RNA, reducing plaque formation and apoptosis. Furthermore, reintroduction of nondegradable reticulon 3 into these knockdown cells rescued enterovirus 71 infectivity, and viral protein and double-stranded RNA synthesis. Thus, reticulon 3 is an important component of enterovirus 71 replication, through its potential role in modulation of the sequential interactions between enterovirus 71 viral RNA and the replication complex.

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Many commonly used siRNAs risk off-target activity

Cited by 131 publications

References 46 publications

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

Functional Analysis of Human Microtubule-based Motor Proteins, the Kinesins and Dyneins, in Mitosis/Cytokinesis Using RNA Interference

Reticulon 3 Binds the 2C Protein of Enterovirus 71 and Is Required for Viral Replication

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