Detection of double-stranded RNAs (dsRNAs) is a central mechanism of innate immune defense in many organisms. We here discuss several families of dsRNA-binding proteins involved in mammalian ...antiviral innate immunity. These include RIG-I-like receptors, protein kinase R, oligoadenylate synthases, adenosine deaminases acting on RNA, RNA interference systems, and other proteins containing dsRNA-binding domains and helicase domains. Studies suggest that their functions are highly interdependent and that their interdependence could offer keys to understanding the complex regulatory mechanisms for cellular dsRNA homeostasis and antiviral immunity. This review aims to highlight their interconnectivity, as well as their commonalities and differences in their dsRNA recognition mechanisms.
A virus (from the Latin word ‘vīrus’ meaning ‘venom’ or ‘poison’) is a microorganism invisible to the naked eye. Viruses can multiply exclusively by entering a cell and using the cell’s resources to ...create copies of themselves. As the origin of their name suggests, viruses are generally considered dangerous, harmful and often deadly. Some of the most well-studied and widely known viruses, such as HIV and influenza, infect humans. However, viruses can also infect animals, plants and microorganisms, including fungi. Many fungi are medically, ecologically and economically significant, for example, causing diseases to humans, plants and insects or being used in industry to produce bread, cheese, beer and wine. Viruses that infect fungi are called mycoviruses (from the Greek work ‘myco’, meaning ‘fungus’). Mycoviruses do not cause harm to or kill the infected fungus; in contrast, they are ‘friendly’ viruses and we can utilize them to control the growth, pathogenicity and toxin production of fungi. This book describes a range of different mycoviruses and their geographical distribution, transmission and evolution, together with their effects on the fungal hosts and how these are brought about.
Global agriculture loses over $100 billion of produce annually to crop pests such as insects. Many of these crop pests either are not currently controlled by artificial means or have developed ...resistance against chemical pesticides. Long dsRNAs are capable of inducing RNAi in insects and are emerging as novel, highly selective alternatives for sustainable insect management strategies. However, there are significant challenges associated with RNAi efficacy in insects. In this study, we synthesized a range of chemically modified long dsRNAs in an approach to improve nuclease resistance and RNAi efficacy in insects. Our results showed that dsRNAs containing phosphorothioate modifications demonstrated increased resistance to southern green stink bug saliva nucleases. Phosphorothioate-modified and 2′-fluoro-modified dsRNA also demonstrated increased resistance to degradation by soil nucleases and increased RNAi efficacy in Drosophila melanogaster cell cultures. In live insects, we found chemically modified long dsRNAs successfully resulted in mortality in both stink bug and corn rootworm. These results provide further mechanistic insight into the dependence of RNAi efficacy on nucleotide modifications in the sense or antisense strand of the dsRNA in insects and demonstrate for the first time that RNAi can successfully be triggered by chemically modified long dsRNAs in insect cells or live insects.
In recent years, the research on the potential of using RNA interference (RNAi) to suppress crop pests has made an outstanding growth. However, given the variability of RNAi efficiency that is ...observed in many insects, the development of novel approaches toward insect pest management using RNAi requires first to unravel factors behind the efficiency of dsRNA-mediated gene silencing. In this review, we explore essential implications and possibilities to increase RNAi efficiency by delivery of dsRNA through non-transformative methods. We discuss factors influencing the RNAi mechanism in insects and systemic properties of dsRNA. Finally, novel strategies to deliver dsRNA are discussed, including delivery by symbionts, plant viruses, trunk injections, root soaking, and transplastomic plants.
Phytopathogens can continuously cause serious losses worldwide to crop, fruit, and vegetable yields. The excessive use of conventional fungicides in pathogen management has raised severe ...environmental and health side effects and induced fungicide-resistant pathogen strains. RNA-based spray-induced gene silencing (SIGS) has emerged as an ecologically sustainable pathogen control approach. Here, we introduce the SIGS mechanism in plant pathogens, summarize the application of SIGS in controlling plant pathogens, and highlight the major considerations of SIGS. In addition, we propose the future perspectives of SIGS in crop protection and disease management.
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RNA interference (RNAi) is an emerging tool in crop protection by exogenous applications of double-stranded RNAs (dsRNAs) to silence the expression of essential pest genes. Nevertheless, topical ...delivery of dsRNAs to sap-sucking insects is a major challenge due to the insufficient infiltration and translocation to plant vascular. Foliar sprayed layered double hydroxide (LDH) showed insufficient translocation in plant vascular and the supplement of surfactants caused phytotoxicity to plant. Thus, in the present study, LDH was transformed from hydrophilic to amphiphilic by fatty acids modification to obtain a surfactant-like LDH (LDHS) with good leaf adhesion, translocation and excellent biocompatibility. LDHS was synthesized via a mild hydrothermal method using MgCl2, FeCl3 and palm oil as the precursors, resulting in LDH coated with long-chained carboxylates as hydrophobic tails. LDHS showed good binding affinity with dsRNA and high affinity to rice leaves. Once loaded with dsRNA, LDHS were shown to enhance the uptake of dsRNA to rice brown planthoppers (Nilaparvata lugens) through a direct spray. Moreover, in planta, dsRNA-LDHS exhibited translocation from rice leaves to vascular bundles and finally into the planthoppers, delivered 2.5-fold higher dsRNA and caused higher mortalities of planthoppers than hydrophilic LDH with adjuvants. Herein we generate and validate a platform that can deliver dsRNA through either direct sprays or sprayed living plant to brown planthoppers with high efficiency, low toxicity and adjuvant-free. Consequently, it is a green potential selection using amphiphilic LDHS agents instead of the conventional gene carriers to prepare biopesticide nanoparticles.
RNA interference (RNAi) is a natural mechanism of gene regulation, highly conserved in eukaryotes. Since the elucidation of the gene silencing mechanism, RNAi became an important tool used in insect ...reverse genetics. The demonstration of effective target-gene silencing by ingestion of double-stranded RNA (dsRNA) produced by transgenic plants indicated the RNAi potential to be used in insect pest management, particularly in agriculture. However, the efficiency of gene silencing by RNAi in insects may vary according to the target taxa, and lepidopteran species have been shown to be quite recalcitrant to RNAi. Developing transgenic plants is a time-consuming and labor-intensive process, so alternative oral delivery systems are required to develop and optimize RNAi settings, such as selecting an efficient target gene, and dsRNA design, length, and stability, among other features. We have developed delivery systems to evaluate dsRNAs to silence genes from two important lepidopteran crop pests of tomato (Solanum lycopersicum) and sugarcane (Saccharum × officinarum): Tuta absoluta (Meyrick), the South American Tomato Pinworm, and Diatraea saccharalis (Fabricius), the Sugarcane Borer, respectively. The protocol described here can be used in similar species and includes (a) direct oral delivery by droplets containing dsRNA; (b) oral delivery by tomato leaflets that absorbed dsRNA solution; (c) delivery by Escherichia coli expressing dsRNA; and (d) delivery by transgenic plants expressing dsRNA.
The RNA interference (RNAi) triggered by short small interfering RNA (siRNA) was discovered in nematodes and found to function in most living organisms. RNAi has been widely used as a research tool ...to study gene functions and has shown great potential for the development of novel pest management strategies. RNAi is highly efficient and systemic in coleopterans but highly variable or inefficient in many other insects. Differences in double-stranded RNA (dsRNA) degradation, cellular uptake, inter- and intracellular transports, processing of dsRNA to siRNA, and RNA-induced silencing complex formation influence RNAi efficiency. The basic dsRNA delivery methods include microinjection, feeding, and soaking. To improve dsRNA delivery, various new technologies, including cationic liposome-assisted, nanoparticle-enabled, symbiont-mediated, and plant-mediated deliveries, have been developed. Major challenges to widespread use of RNAi in insect pest management include variable RNAi efficiency among insects, lack of reliable dsRNA delivery methods, off-target and nontarget effects, and potential development of resistance in insect populations.
Mycoviruses exist in all major groups of fungi. With the continuous development of science and technology, the methods of studying viruses are constantly updated, and progressively mycoviruses have ...been discovered where most of these viruses are RNA viruses. Therefore, double-stranded RNA has traditionally been used as the hallmark of RNA mycovirus detection. This report describes in detail the method of mycovirus identification using extraction of dsRNA. Besides, extraction of viral dsRNA, and the assembly methods of viral genome and identification of virus type are presented.
Bi-directional transcription of Human Endogenous Retroviruses (hERVs) is a common feature of autoimmunity, neurodegeneration and cancer. Higher rates of cancer incidence, neurodegeneration and ...autoimmunity but a lower prevalence of autoimmune diseases characterize elderly people. Although the re-expression of hERVs is commonly observed in different cellular models of senescence as a result of the loss of their epigenetic transcriptional silencing, the hERVs modulation during aging is more complex, with a peak of activation in the sixties and a decline in the nineties. What is clearly accepted, instead, is the impact of the re-activation of dormant hERV on the maintenance of stemness and tissue self-renewing properties. An innate cellular immunity system, based on the RLR-MAVS circuit, controls the degradation of dsRNAs arising from the transcription of hERV elements, similarly to what happens for the accumulation of cytoplasmic DNA leading to the activation of cGAS/STING pathway. While agonists and inhibitors of the cGAS–STING pathway are considered promising immunomodulatory molecules, the effect of the RLR-MAVS pathway on innate immunity is still largely based on correlations and not on causality. Here we review the most recent evidence regarding the activation of MDA5-RIG1-MAVS pathway as a result of hERV de-repression during aging, immunosenescence, cancer and autoimmunity. We will also deal with the epigenetic mechanisms controlling hERV repression and with the strategies that can be adopted to modulate hERV expression in a therapeutic perspective. Finally, we will discuss if the RLR-MAVS signalling pathway actively modulates physiological and pathological conditions or if it is passively activated by them.
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