Intron-derived small RNAs for silencing viral RNAs in mosquito cells

Aedes aegypti and Ae. albopictus are the main vectors of mosquito-borne viruses of medical and veterinary significance. Many of these viruses have RNA genomes. Exogenously provided, e.g. transgene encoded, small RNAs could be used to inhibit virus replication, breaking the transmission cycle. We tested, in Ae. aegypti and Ae. albopictus cell lines, reporter-based strategies for assessing the ability of two types of small RNAs to inhibit a chikungunya virus (CHIKV) derived target. Both types of small RNAs use a Drosophila melanogaster pre-miRNA-1 based hairpin for their expression, either with perfect base-pairing in the stem region (shRNA-like) or containing two mismatches (miRNA-like). The pre-miRNA-1 stem loop structure was encoded within an intron; this allows co-expression of one or more proteins, e.g. a fluorescent protein marker tracking the temporal and spatial expression of the small RNAs in vivo. Three reporter-based systems were used to assess the relative silencing efficiency of ten shRNA-like siRNAs and corresponding miRNA-like designs. Two systems used a luciferase reporter RNA with CHIKV RNA inserted either in the coding sequence or within the 3’ UTR. A third reporter used a CHIKV derived split replication system. All three reporters demonstrated that while silencing could be achieved with both miRNA-like and shRNA-like designs, the latter were substantially more effective. Dcr-2 was required for the shRNA-like siRNAs as demonstrated by loss of inhibition of the reporters in Dcr-2 deficient cell lines. These positive results in cell culture are encouraging for the potential use of this pre-miRNA-1-based system in transgenic mosquitoes.     

Small RNAs in viral infection and host defense

Small RNAs are the key mediators of RNA silencing and related pathways in plants and other eukaryotic organisms. Silencing pathways couple the destruction of double-stranded RNA with the use of the resulting small RNAs to target other nucleic acid molecules that contain the complementary sequence. This discovery has revolutionized our ideas about host defense and genetic regulatory mechanisms in eukaryotes. Small RNAs can direct the degradation of mRNAs and single-stranded viral RNAs, the modification of DNA and histones, and the inhibition of translation. Viruses might even use small RNAs to do some targeting of their own to manipulate host gene expression. This review highlights the current understanding and new insights concerning the roles of small RNAs in virus infection and host defense in plants.     

Multiple roles for small RNAs during plant reproduction

Germline development and early embryogenesis in eukaryotes are characterized by large-scale genome reprogramming events. In companion cells of the Arabidopsis male gametophyte, epigenome reorganization leads to loss of heterochromatin and production of a distinct small RNA (sRNA) population. A specific class of sRNA derived from transposons appears to be mobile and can accumulate in germ cells. In the germline of maize, rice, and Arabidopsis, specific ARGONAUTE-sRNA silencing complexes appear to play key roles in reproductive development, including meiosis and regulation of germ cell fate. These results reveal new roles for sRNAs during plant reproduction and suggest that mobility of sRNAs could be critical for some of these functions.     

The roles of pathogen small RNAs

Regulatory small RNAs (sRNAs), also known as non-coding RNA, are not translated into proteins and widespread in prokaryotes and eukaryotes. sRNAs involve in multiple fundamental cellular events. They are emerging regulatory elements that are gaining momentum. Knowledge of sRNA largely originates from eukaryotes. Prokaryotic sRNAs, particularly those of pathogen are only recently explored. The main types, function, and methodology to predict pathogen sRNAs are summarized in this review. Special focus is sRNAs regulating pathogen gene expression, particularly that of Mycobacterium tuberculosis, which is hitherto the most successful pathogen afflicting mankind.     

小分子RNA抵御转座元件对基因组的侵袭

真核生物在漫长的进化繁衍过程中,一直处在抵御转座元件对基因组侵害的"斗争"中。在过去的十多年中,越来越多的证据指明,小分子RNA扮演了这样一个抵御转座子侵袭的角色。尽管这种抵御侵害的策略对于不同物种各具特点,但它们都呈现出惊人相似的共同特征。基本上,所有的机制都包含三个组成部分:首先,转座元件促使产生小分子RNA,在某些物种中主要是Piwi-interacting RNAs,而在其他物种中主要是small interfering RNAs;第二,作用于活跃转座子的小分子RNA通过RNA依赖性RNA聚合酶或切割机制进行扩增;第三,这些小分子RNA与含有Argonaute蛋白或Piwi蛋白的效应复合物相结合,从而作用于目标转座子的转录本,实现转录后沉默,或作用于目标转座子DNA,抑制染色质修饰和DNA甲基化。这些属性特征构成了一个限制由转座元件活动所造成的严重后果的系统,从而防止转座子侵袭所带来的突变积累,基因表达谱的改变,以及生殖腺发育不良和不育。     

The many roles of small RNAs in leaf development

Leaf development involves many complex genetic interactions,signals between adjacent cells or between more distant tissues and consequent changes in cell fate.This review describes three stages in leaf development where regulation by small RNAs have been used to modulate gene expression patterns.     

The many roles of small RNAs in leaf development

Leaf development involves many complex genetic interactions,signals between adjacent cells or between more distant tissues and consequent changes in cell fate.This review describes three stages in leaf development where regulation by small RNAs have been used to modulate gene expression patterns.     

Sequence-independent characterization of viruses based on the pattern of viral small RNAs produced by the host

Virus surveillance in vector insects is potentially of great benefit to public health. Large-scale sequencing of small and long RNAs has previously been used to detect viruses, but without any formal comparison of different strategies. Furthermore, the identification of viral sequences largely depends on similarity searches against reference databases. Here, we developed a sequence-independent strategy based on virus-derived small RNAs produced by the host response, such as the RNA interference pathway. In insects, we compared sequences of small and long RNAs, demonstrating that viral sequences are enriched in the small RNA fraction. We also noted that the small RNA size profile is a unique signature for each virus and can be used to identify novel viral sequences without known relatives in reference databases. Using this strategy, we characterized six novel viruses in the viromes of laboratory fruit flies and wild populations of two insect vectors: mosquitoes and sandflies. We also show that the small RNA profile could be used to infer viral tropism for ovaries among other aspects of virus biology. Additionally, our results suggest that virus detection utilizing small RNAs can also be applied to vertebrates, although not as efficiently as to plants and insects.     

小分子RNA在植物激素信号通路中的调控功能

植物激素是调控植物生长发育的信号分子。近年来的研究发现,小分子RNA作为基因表达调控网络的组分,参与植物激素信号途径,在植物生长发育和胁迫反应方面发挥重要作用。本文综述了miRNA和次级siRNA(Short interfering RNAs)介导的基因调控与植物激素信号通路相互作用的研究进展,主要包括生长素、赤霉素、油菜素内酯和脱落酸途径涉及的miRNA及其功能,并对不同发育过程中miRNA参与的不同激素信号通路的交叉和互作进行了讨论。     

Big roles for small RNAs in polyploidy, hybrid vigor, and hybrid incompatibility

Small RNAs, including microRNAs (miRNAs), small interfering RNAs (siRNAs), and trans-acting siRNAs (ta-siRNAs), mediate gene expression and epigenetic regulation. While siRNAs are highly diverged, miRNAs and ta-siRNAs are generally conserved but many are differentially expressed between related species and in interspecific hybrids and allopolyploids. On one hand, combination of diverged maternal and paternal siRNAs in the same nucleus may exert cis-acting and trans-acting effects on transposable elements (TEs) and TE-associated genes, leading to genomic instability and endosperm and embryo failures, constituting a bottleneck for the evolution of hybrids and polyploids. On the other hand, cis and trans-acting small RNAs induce quantitative and qualitative changes in epigenetic regulation, leading to morphological variation and hybrid vigor in F1 hybrids and stable allopolyploids as well as transgressive phenotypes in the progeny, increasing a potential for adaptive evolution.     

Expression profiles and potential roles of transfer RNA-derived small RNAs in atherosclerosis

Knowledge regarding the relationship between the molecular mechanisms underlying atherosclerosis (AS) and transfer RNA-derived small RNAs (tsRNAs) is limited. This study illustrated the expression profile of tsRNAs, thus exploring its roles in AS pathogenesis. Small RNA sequencing was performed with four atherosclerotic arterial and four healthy subject samples. Using bioinformatics, the protein-protein interaction network and cellular experiments were constructed to predict the enriched signalling pathways and regulatory roles of tsRNAs in AS. Of the total 315 tsRNAs identified to be dysregulated in the AS group, 131 and 184 were up-regulated and down-regulated, respectively. Interestingly, the pathway of the differentiated expression of tsRNAs in cell adhesion molecules (CAMs) was implicated to be closely associated with AS. Particularly, tRF-Gly-GCC might participate in AS pathogenesis via regulating cell adhesion, proliferation, migration and phenotypic transformation in HUVECs and VSMCs. In conclusion, tsRNAs might help understand the molecular mechanisms of AS better. tRF-Gly-GCC may be a promising target for suppressing abnormal vessels functions, suggesting a novel strategy for preventing the progression of atherosclerosis.     

'Complementing' viral infection: mechanisms for evading innate immunity

Through co-evolution with their hosts, viruses have developed a variety of immune escape and control mechanisms. In addition to strategies used to avoid the cellular and humoral immune responses, many viral families encode proteins capable of neutralizing the host's first line of defense, complement. The diversity of these complement avoidance mechanisms and proposed mechanisms by which viruses not only avoid, but also use the immune system to their advantage are discussed.     

New functions of small nucleolar RNAs

Small nucleolar RNAs (snoRNAs) are one of the most abundant and well-studied groups of non-coding RNAs. snoRNAs are mostly engaged in processing of rRNA. However, recent data indicate that snoRNAs are also involved in other processes including regulation of alternative splicing, translation and oxidative stress. snoRNAs are also involved in pathogenesis of some hereditary diseases and cancer. Therefore, the range of snoRNAs’ functions is significantly wider than it has been assumed earlier.