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Two decades of research on RNA interference (RNAi) have transformed a breakthrough discovery in biology into a robust platform for a new class of medicines that modulate mRNA expression. Here we provide an overview of the trajectory of small-interfering RNA (siRNA) drug development, including the first approval in 2018 of a liver-targeted siRNA interference (RNAi) therapeutic in lipid nanoparticles and subsequent approvals of five more RNAi drugs, which used metabolically stable siRNAs combined with N -acetylgalactosamine ligands for conjugate-based liver delivery. We also consider the remaining challenges in the field, such as delivery to muscle, brain and other extrahepatic organs. Today’s RNAi therapeutics exhibit high specificity, potency and durability, and are transitioning from applications in rare diseases to widespread, chronic conditions. With six approved drugs, siRNA is now an established therapeutic modality poised for expansion.

CRISPR–Cas interference is mediated by Cas effector nucleases that are either components of multisubunit complexes—in class 1 CRISPR–Cas systems—or domains of a single protein—in class 2 systems 1 – 3 . Here we show that the subtype III-E effector Cas7-11 is a single-protein effector in the class 1 CRISPR–Cas systems originating from the fusion of a putative Cas11 domain and multiple Cas7 subunits that are derived from subtype III-D. Cas7-11 from Desulfonema ishimotonii ( Di Cas7-11), when expressed in Escherichia coli , has substantial RNA interference effectivity against mRNAs and bacteriophages. Similar to many class 2 effectors—and unique among class 1 systems— Di Cas7-11 processes pre-CRISPR RNA into mature CRISPR RNA (crRNA) and cleaves RNA at positions defined by the target:spacer duplex, without detectable non-specific activity. We engineered Cas7-11 for RNA knockdown and editing in mammalian cells. We show that Cas7-11 has no effects on cell viability, whereas other RNA-targeting tools (such as short hairpin RNAs and Cas13) show substantial cell toxicity 4 , 5 . This study illustrates the evolution of a single-protein effector from multisubunit class 1 effector complexes, expanding our understanding of the diversity of CRISPR systems. Cas7-11 provides the basis for new programmable RNA-targeting tools that are free of collateral activity and cell toxicity. Cas7-11—the fusion of a putative Cas11 domain and four Cas7 subunits—cleaves RNA without detectable non-specific activity and, when optimized for RNA knockdown and editing in mammalian cells, has no effects on cell viability.

The class 2 type VI RNA-guided RNA-targeting CRISPR–Cas effector Cas13 can be engineered for RNA knockdown and binding, expanding the CRISPR toolset with a flexible platform for studying RNA in mammalian cells and therapeutic development. A CRISPR way to knockdown RNA CRISPR–Cas prokaryotic defence systems have provided versatile tools for DNA editing. Here, the authors demonstrate that the class 2 type VI RNA-guided RNA-targeting CRISPR–Cas effector Cas13a (previously known as C2c2) can be engineered for RNA knockdown and binding in mammalian cells. This addition to the CRISPR toolbox expands its potential uses to transcript tracking and knockdown. RNA has important and diverse roles in biology, but molecular tools to manipulate and measure it are limited. For example, RNA interference 1 , 2 , 3 can efficiently knockdown RNAs, but it is prone to off-target effects 4 , and visualizing RNAs typically relies on the introduction of exogenous tags 5 . Here we demonstrate that the class 2 type VI 6 , 7 RNA-guided RNA-targeting CRISPR–Cas effector Cas13a 8 (previously known as C2c2) can be engineered for mammalian cell RNA knockdown and binding. After initial screening of 15 orthologues, we identified Cas13a from Leptotrichia wadei (LwaCas13a) as the most effective in an interference assay in Escherichia coli . LwaCas13a can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts with comparable levels of knockdown as RNA interference and improved specificity. Catalytically inactive LwaCas13a maintains targeted RNA binding activity, which we leveraged for programmable tracking of transcripts in live cells. Our results establish CRISPR–Cas13a as a flexible platform for studying RNA in mammalian cells and therapeutic development.

Protection against microbial infection in eukaryotes is provided by diverse cellular and molecular mechanisms. Here, we present a comparative view of the antiviral activity of virus-derived small interfering RNAs in fungi, plants, invertebrates and mammals, detailing the mechanisms for their production, amplification and activity. We also highlight the recent discovery of viral PIWI-interacting RNAs in animals and a new role for mobile host and pathogen small RNAs in plant defence against eukaryotic pathogens. In turn, viruses that infect plants, insects and mammals, as well as eukaryotic pathogens of plants, have evolved specific virulence proteins that suppress RNA interference (RNAi). Together, these advances suggest that an antimicrobial function of the RNAi pathway is conserved across eukaryotic kingdoms.

We present and discuss perspectives of current developments on advanced quantum optical circuits monolithically integrated in the lithium niobate platform. A set of basic components comprising photon pair sources based on parametric down conversion (PDC), passive routing elements and active electro-optically controllable switches and polarisation converters are building blocks of a toolbox which is the basis for a broad range of diverse quantum circuits. We review the state-of-the-art of these components and provide models that properly describe their performance in quantum circuits. As an example for applications of these models we discuss design issues for a circuit providing on-chip two-photon interference. The circuit comprises a PDC section for photon pair generation followed by an actively controllable modified mach-Zehnder structure for observing Hong-Ou-Mandel interference. The performance of such a chip is simulated theoretically by taking even imperfections of the properties of the individual components into account.

RNA interference (RNAi) has been widely adopted to repress specific gene expression and is easily achieved by designing small interfering RNAs (siRNAs) with perfect sequence complementarity to the intended target mRNAs. Although siRNAs direct Argonaute (Ago), a core component of the RNA-induced silencing complex (RISC), to recognize and silence target mRNAs, they also inevitably function as microRNAs (miRNAs) and suppress hundreds of off-targets. Such miRNA-like off-target repression is potentially detrimental, resulting in unwanted toxicity and phenotypes. Despite early recognition of the severity of miRNA-like off-target repression, this effect has often been overlooked because of difficulties in recognizing and avoiding off-targets. However, recent advances in genome-wide methods and knowledge of Ago–miRNA target interactions have set the stage for properly evaluating and controlling miRNA-like off-target repression. Here, we describe the intrinsic problems of miRNA-like off-target effects caused by canonical and noncanonical interactions. We particularly focus on various genome-wide approaches and chemical modifications for the evaluation and prevention of off-target repression to facilitate the use of RNAi with secured specificity.

Rhizobial infection and root nodule formation in legumes require recognition of signal molecules produced by the bacteria and their hosts. Here, we show that rhizobial transfer RNA (tRNA)-derived small RNA fragments (tRFs) are signal molecules that modulate host nodulation. Three families of rhizobial tRFs were confirmed to regulate host genes associated with nodule initiation and development through hijacking the host RNA-interference machinery that involves ARGONAUTE 1. Silencing individual tRFs with the use of short tandem target mimics or by overexpressing their targets represses root hair curling and nodule formation, whereas repressing these targets with artificial microRNAs identical to the respective tRFs or mutating these targets with CRISPR-Cas9 promotes nodulation. Our findings thus uncover a bacterial small RNA–mediated mechanism for prokaryote-eukaryote interaction and may pave the way for enhancing nodulation efficiency in legumes.

Materials with good flexibility and high conductivity that can provide electromagnetic interference (EMI) shielding with minimal thickness are highly desirable, especially if they can be easily processed into films. Two-dimensional metal carbides and nitrides, known as MXenes, combine metallic conductivity and hydrophilic surfaces. Here, we demonstrate the potential of several MXenes and their polymer composites for EMI shielding. A 45-micrometer-thick Ti₃C₂Tx film exhibited EMI shielding effectiveness of 92 decibels (>50 decibels for a 2.5-micrometer film), which is the highest among synthetic materials of comparable thickness produced to date. This performance originates from the excellent electrical conductivity of Ti₃C₂Tx films (4600 Siemens per centimeter) and multiple internal reflections from Ti₃C₂Tx flakes in free-standing films. The mechanical flexibility and easy coating capability offered by MXenes and their composites enable them to shield surfaces of any shape while providing high EMI shielding efficiency.

Background CRISPR/Cas systems confer immunity against invading nucleic acids and phages in bacteria and archaea. CRISPR/Cas13a (known previously as C2c2) is a class 2 type VI-A ribonuclease capable of targeting and cleaving single-stranded RNA (ssRNA) molecules of the phage genome. Here, we employ CRISPR/Cas13a to engineer interference with an RNA virus, Turnip Mosaic Virus (TuMV), in plants. Results CRISPR/Cas13a produces interference against green fluorescent protein (GFP)-expressing TuMV in transient assays and stable overexpression lines of Nicotiana benthamiana. CRISPR RNA (crRNAs) targeting the HC-Pro and GFP sequences exhibit better interference than those targeting other regions such as coat protein (CP) sequence. Cas13a can also process pre-crRNAs into functional crRNAs. Conclusions Our data indicate that CRISPR/Cas13a can be used for engineering interference against RNA viruses, providing a potential novel mechanism for RNA-guided immunity against RNA viruses and for other RNA manipulations in plants.

The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) adaptive immune systems use small guide RNAs, the CRISPR RNAs (crRNAs), to mark foreign genetic material, e.g. viral nucleic acids, for degradation. Archaea and bacteria encode a large variety of Cas proteins that bind crRNA molecules and build active ribonucleoprotein surveillance complexes. The evolution of CRISPR-Cas systems has resulted in a diversification of cas genes and a classification of the systems into three types and additional subtypes characterized by distinct surveillance and interfering complexes. Recent crystallographic and biochemical advances have revealed detailed insights into the assembly and DNA/RNA targeting mechanisms of the various complexes. Here, we review our knowledge on the molecular mechanism involved in the DNA and RNA interference stages of type I (Cascade: CRISPR-associated complex for antiviral defense), type II (Cas9) and type III (Csm, Cmr) CRISPR-Cas systems. We further highlight recently reported structural and mechanistic themes shared among these systems. This review details and compares the assembly and the DNA/RNA targeting mechanisms of the various surveillance complexes of prokaryotic CRISPR-Cas immune systems.