MicroRNAs: Genomics, Biogenesis, Mechanism, and Function

抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Double-stranded RNA (dsRNA) induces sequence-specific posttranscriptional gene silencing in many organisms by a process known as RNA interference (RNAi). Using a Drosophila in vitro system, we demonstrate that 21- and 22-nt RNA fragments are the sequence-specific mediators of RNAi. The short interfering RNAs (siRNAs) are generated by an RNase III–like processing reaction from long dsRNA. Chemically synthesized siRNA duplexes with overhanging 3 ends mediate efficient target RNA cleavage in the lysate, and the cleavage site is located near the center of the region spanned by the guiding siRNA. Furthermore, we provide evidence that the direction of dsRNA processing determines whether sense or antisense target RNA can be cleaved by the siRNA–protein complex.

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RNA interference is mediated by 21- and 22-nucleotide RNAs

Posttranscriptional gene silencing (PTGS) is a nucleotide sequence-specific defense mechanism that can target both cellular and viral mRNAs. Here, three types of transgene-induced PTGS and one example of virus-induced PTGS were analyzed in plants. In each case, antisense RNA complementary to the targeted mRNA was detected. These RNA molecules were of a uniform length, estimated at 25 nucleotides, and their accumulation required either transgene sense transcription or RNA virus replication. Thus, the 25-nucleotide antisense RNA is likely synthesized from an RNA template and may represent the specificity determinant of PTGS.

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A species of small antisense RNA in posttranscriptional gene silencing in plants.

Small RNAs of 20-30 nucleotides can target both chromatin and transcripts, and thereby keep both the genome and the transcriptome under extensive surveillance. Recent progress in high-throughput sequencing has uncovered an astounding landscape of small RNAs in eukaryotic cells. Various small RNAs of distinctive characteristics have been found and can be classified into three classes based on their biogenesis mechanism and the type of Argonaute protein that they are associated with: microRNAs (miRNAs), endogenous small interfering RNAs (endo-siRNAs or esiRNAs) and Piwi-interacting RNAs (piRNAs). This Review summarizes our current knowledge of how these intriguing molecules are generated in animal cells.

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Biogenesis of small RNAs in animals.

Induction and suppression of antiviral RNA interference (RNAi) has been observed in mammals during infection with at least seven distinct RNA viruses, including some that are pathogenic in humans. However, while the cell‐autonomous immune response mediated by antiviral RNAi is gradually being recognized, little is known about systemic antiviral RNAi in mammals. Furthermore, extracellular vesicles (EVs) also function in viral signal spreading and host immunity. Here, we show that upon antiviral RNAi activation, virus‐derived small‐interfering RNAs (vsiRNAs) from Nodamura virus (NoV), Sindbis virus (SINV), and Zika virus (ZIKV) enter the murine bloodstream via EVs for systemic circulation. vsiRNAs in the EVs are biologically active, since they confer RNA–RNA homology‐dependent antiviral activity in both cultured cells and infant mice. Moreover, we demonstrate that vaccination with a live‐attenuated virus, rendered deficient in RNAi suppression, induces production of stably maintained vsiRNAs and confers protective immunity against virus infection in mice. This suggests that vaccination with live‐attenuated VSR (viral suppressor of RNAi)‐deficient mutant viruses could be a new strategy to induce immunity.

Mouse circulating extracellular vesicles contain virus‐derived siRNAs active in antiviral immunity

The present invention provides recombinant DNA constructs for inactivation of viral or endogenous genes in a cell, wherein the construct comprises viral sequence sufficient to activate RNA silencing. In another aspect, the invention provides methods for identifying RNA silencing suppressors by sequence analysis and functional tests. In yet another aspect, the invention provides a method for identifying inhibitors of RNA silencing suppressors. In still other aspects, the invention comprises methods for identifying genes in the antiviral RNA silencing pathway, enhancers of the antiviral pathway, and methods of treating or preventing viral infections using enhancers of the pathway.

RNA silencing in animals as an antiviral defense

Virus-host coevolution often drives virus immune escape. However, it remains unknown whether natural variations of plant virus resistance are enriched in genes of RNA interference (RNAi) pathway known to confer essential antiviral defense in plants. Here, we report two genome-wide association study screens to interrogate natural variation among wild-collected Arabidopsis thaliana accessions in quantitative resistance to the endemic cucumber mosaic virus (CMV). We demonstrate that the highest-ranked gene significantly associated with resistance from both screens acts to regulate antiviral RNAi in ecotype Columbia-0. One gene, corresponding to Reduced Dormancy 5 (RDO5), enhances resistance by promoting amplification of the virus-derived small interfering RNAs (vsiRNAs). Interestingly, the second gene, designated Antiviral RNAi Regulator 1 (VIR1), dampens antiviral RNAi so its genetic inactivation by CRISPR/Cas9 editing enhances both vsiRNA production and CMV resistance. Our findings identify positive and negative regulators of the antiviral RNAi defense that may play important roles in virus-host coevolution. 

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Identification of positive and negative regulators of antiviral RNA interference in Arabidopsis thaliana

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/j.febslet.2005.08.036

Shou-Wei Ding

Papers

Mouse circulating extracellular vesicles contain virus‐derived siRNAs active in antiviral immunity

RNA silencing in animals as an antiviral defense

Identification of positive and negative regulators of antiviral RNA interference in Arabidopsis thaliana

RNAi: Mechanisms, biology and applications

Mammalian viral suppressors of RNA interference.