title: Specific inhibition of diverse pathogens in human cells by synthetic microRNA-like oligonucleotides inferred from RNAi screens.
abstract: Systematic genetic perturbation screening in human cells remains technically challenging. Typically, large libraries of chemically synthesized siRNA oligonucleotides are used, each designed to degrade a specific cellular mRNA via the RNA interference (RNAi) mechanism. Here, we report on data from three genome-wide siRNA screens, conducted to uncover host factors required for infection of human cells by two bacterial and one viral pathogen. We find that the majority of phenotypic effects of siRNAs are unrelated to the intended "on-target" mechanism, defined by full complementarity of the 21-nt siRNA sequence to a target mRNA. Instead, phenotypes are largely dictated by "off-target" effects resulting from partial complementarity of siRNAs to multiple mRNAs via the "seed" region (i.e., nucleotides 2-8), reminiscent of the way specificity is determined for endogenous microRNAs. Quantitative analysis enabled the prediction of seeds that strongly and specifically block infection, independent of the intended on-target effect. This prediction was confirmed experimentally by designing oligos that do not have any on-target sequence match at all, yet can strongly reproduce the predicted phenotypes. Our results suggest that published RNAi screens have primarily, and unintentionally, screened the sequence space of microRNA seeds instead of the intended on-target space of protein-coding genes. This helps to explain why previously published RNAi screens have exhibited relatively little overlap. Our analysis suggests a possible way of identifying "seed reagents" for controlling phenotypes of interest and establishes a general strategy for extracting valuable untapped information from past and future RNAi screens.
authors.affiliation: Institute of Molecular Life Sciences and Swiss Institute of Bioinformatics, University of Zurich, CH-8057 Zurich, Switzerland.
authors.forename: Andrea
authors.initials: A
authors.lastname: Franceschini
authors.forename: Roger
authors.initials: R
authors.lastname: Meier
authors.forename: Alain
authors.initials: A
authors.lastname: Casanova
authors.forename: Saskia
authors.initials: S
authors.lastname: Kreibich
authors.forename: Neha
authors.initials: N
authors.lastname: Daga
authors.forename: Daniel
authors.initials: D
authors.lastname: Andritschke
authors.forename: Sabrina
authors.initials: S
authors.lastname: Dilling
authors.forename: Pauli
authors.initials: P
authors.lastname: Rämö
authors.forename: Mario
authors.initials: M
authors.lastname: Emmenlauer
authors.forename: Andreas
authors.initials: A
authors.lastname: Kaufmann
authors.forename: Raquel
authors.initials: R
authors.lastname: Conde-Álvarez
authors.forename: Shyan Huey
authors.initials: SH
authors.lastname: Low
authors.forename: Lucas
authors.initials: L
authors.lastname: Pelkmans
authors.forename: Ari
authors.initials: A
authors.lastname: Helenius
authors.forename: Wolf-Dietrich
authors.initials: WD
authors.lastname: Hardt
authors.forename: Christoph
authors.initials: C
authors.lastname: Dehio
authors.forename: Christian
authors.initials: C
authors.lastname: von Mering
mesh_terms: D001483:Base Sequence; D002003:Brucella abortus / Q000187:drug effects* / Q000235:genetics; D002043:Bunyaviridae / Q000187:drug effects* / Q000235:genetics; D005798:Genes, Bacterial; D006367:HeLa Cells; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D009841:Oligonucleotides / Q000494:pharmacology*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000235:genetics
chemical_list: D035683:MicroRNAs; D009841:Oligonucleotides; D034741:RNA, Small Interfering
keywords: antimicrobials; high-throughput RNAi screening