DISE: A Seed-Dependent RNAi Off-Target Effect That Kills Cancer Cells

Putzbach, William; Gao, Quan; Patel, Monal; Haluck-Kangas, Ashley; Murmann, Andrea E.; Peter, Marcus E.

doi:10.1016/j.trecan.2017.11.007

Cited by 23 publications

(30 citation statements)

References 66 publications

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“…Consistently, miR-34a exhibits low toxicity to normal cells in vitro and in vivo (Di Martino et al, 2012). If one assumes most of its toxicity comes from its toxic 6mer seed sequence, these reports are consistent with our observation that mice treated with toxic siRNAs while slowing down tumor growth showed no signs of toxicity to normal tissues (Murmann et al, 2017). In contrast, in cancer cells miR-34a,b,c become highly expressed after genotoxic stress as they are p53-regulated (He et al, 2007).…”

Section: Mir-34a and Its Clinical Utilitysupporting

confidence: 90%

“…We had previously postulated that 6mer seed toxicity could be an anticancer mechanism and predicted that small RNAi active guide RNAs would be involved in killing cancer cells (Putzbach, Gao, Patel, Haluck-Kangas, et al, 2017). Consistent with this hypothesis, an analysis of published data on point mutations in the exomes of 27 different human cancer showed that losing a C is the predominant point mutation across all cancers (Lawrence et al, 2013).…”

Section: Sirnas With Toxic Seeds Target Housekeeping Genes Enriched Imentioning

confidence: 84%

“…We recently postulated the existence of small RNAi-active sequences that are highly toxic to cancer cells (Putzbach, Gao, Patel, Haluck-Kangas, et al, 2017). However, identifying all of them in a screen is not feasible, as a 19mer duplex siRNA would have 270 billion different sequence combinations.…”

Section: Discussion the Toxic Seed Screenmentioning

confidence: 99%

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6mer Seed Toxicity Determines Strand Selection in miRNAs

Gao¹,

Putzbach²,

Murmann³

et al. 2018

Preprint

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SUMMARYMany siRNAs and shRNAs are toxic to cancer cells through a 6mer seed sequence (position 2-7 of the guide strand). A siRNA screen with all 4096 possible 6mer seed sequences in a neutral RNA backbone revealed a preference for guanine in positions 1-3 and a GC content of >80% of the 6mer seed in the most toxic siRNAs. These 6mer seed containing siRNAs exert their toxicity by targeting survival genes which contain GC-rich 3'UTRs. The master tumor suppressor miRNA miR-34a was found to be toxic through such a G-rich 6mer seed suggesting that certain tumor suppressive miRNAs use a toxic 6mer seed to kill cancer cells. An analysis of all mature miRNAs suggests that most miRNAs evolved to avoid guanine at the 5' end of the 6mer seed sequence of the predominantly expressed arm. In contrast, for many tumor suppressive miRNAs the predominant arm contains a G-rich toxic 6mer seed, presumably to eliminate cancer cells. INTRODUCTIONRNA interference (RNAi) is a form of post-transcriptional regulation exerted by 19-21nt long double stranded RNAs that represses expression of target mRNAs that harbor reverse complementarity to the antisense/guide strand of the small RNA. This results in degradation of the targeted mRNA or in translational repression. RNAiactive guide RNAs can be endogenous siRNAs and micro(mi)RNAs. For a miRNA, the RNAi pathway begins in the nucleus with transcription of a primary miRNA precursor (pri-miRNA) . Pri-miRNAs are first processed by the Drosha/DGCR8 Microprocessor complex into pre-microRNAs which are exported from the nucleus to the cytoplasm by Exportin 5 (Yi, Qin, Macara, & Cullen, 2003). Once in the cytoplasm, Dicer processes them further (Bernstein, Caudy, Hammond, & Hannon, 2001;Hutvagner et al., 2001) and these mature dsRNA duplexes are then loaded into Argonaute (Ago) proteins to form the RNAinduced silencing complex (RISC) (Y. Wang, Sheng, Juranek, Tuschl, & Patel, 2008). The sense/passenger strand is ejected/degraded, while the guide strand remains associated with the RISC (Leuschner, Ameres, Kueng, & Martinez, 2006). While si/shRNAs are usually designed with 100% reverse complementarity to their intended targets and induce AGO2 dependent target degradation (Schirle & MacRae, 2012), cleavageindependent RNAi as exerted by miRNAs causes deadenylation/degradation or translational repression via interaction between AGO2 and TNRC6 family proteins that in turn recruit the CCR4-NOT deadenylase complex (Eulalio, Huntzinger, & Izaurralde, 2008). RNAi can be initiated with as little as six nucleotide basepairing between a guide RNA's so-called seed sequence (positions 2 to 7) and the target RNA (Lai, 2002;Lewis, Shih, Jones-Rhoades, Bartel, & Burge, 2003). This seed-based targeting is restricted to binding sites located in the 3'UTR of a target mRNA (Baek et al., 2008;Selbach et al., 2008). miRNAs play important roles in cancer (Esquela-Kerscher & Slack, 2006). While the function of a miRNA depends on the nature of its targets, some miRNA families have predominantly tumor suppressive or oncogenic...

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Section: Mir-34a and Its Clinical Utilitysupporting

confidence: 90%

Section: Sirnas With Toxic Seeds Target Housekeeping Genes Enriched Imentioning

confidence: 84%

See 1 more Smart Citation

6mer Seed Toxicity Determines Strand Selection in miRNAs

Gao¹,

Putzbach²,

Murmann³

et al. 2018

Preprint

Self Cite

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“…Peter and colleagues have identified suicide/killer RNA molecules (siRNA, shRNA, miRNA, siRNA+miRNA complex) on numerous cancer types. In addition, they have shown that specific toxic RNAi-active sequences present in the genome can kill cancer cells [40][41][42][43][44]. Rozowsky and colleagues have generated a comprehensive analytic platform for extracellular RNA profiling called "exceRpt" [45].…”

Section: Application Field Of Omics Technology In Molecular Medicinementioning

confidence: 99%

Introductory Chapter: Insight into the OMICS Technologies and Molecular Medicine

Nalbantoglu¹,

Karadağ²

2019

Molecular Medicine

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“…We called this form of cell death DISE, for death induced by survival gene elimination (Putzbach et al, 2018a).…”

Section: Introductionmentioning

confidence: 99%

6mer seed toxicity in viral microRNAs

Murmann

Bartom

Schipma

et al. 2019

Preprint

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Micro(mi)RNAs are short double stranded noncoding RNAs (19-23nts) that regulate gene expression by suppressing mRNAs through RNA interference. Targeting is determined by the seed sequence (position 2-7/8) of the mature miRNA. A minimal G-rich seed of just 6 nucleotides is highly toxic to cells by targeting genes essential for cell survival. A screen of 215 miRNAs encoded by 17 human pathogenic viruses (v-miRNAs) now suggests that a number of v-miRNAs can kill cells through a G-rich 6mer sequence embedded in their seed.Specifically, we demonstrate that miR-K12-6-5p, an oncoviral mimic of the tumor suppressive miR-15/16 family encoded by human Kaposi's sarcoma-associated herpes virus, harbors a noncanonical toxic 6mer seed (position 3-8) and that v-miRNAs are more likely than cellular miRNAs to utilize a noncanonical 6mer seed. Our data suggest that during evolution viruses evolved to use 6mer seed toxicity to kill cells.

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DISE: A Seed-Dependent RNAi Off-Target Effect That Kills Cancer Cells

Cited by 23 publications

References 66 publications

6mer Seed Toxicity Determines Strand Selection in miRNAs

6mer Seed Toxicity Determines Strand Selection in miRNAs

Introductory Chapter: Insight into the OMICS Technologies and Molecular Medicine

6mer seed toxicity in viral microRNAs

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