Genetic architecture of mitochondrial editing in Arabidopsis thaliana

We have analyzed the mitochondrial editing behavior of two Arabidopsis thaliana accessions, Landsberg erecta (Ler) and Columbia (Col). A survey of 362 C-to-U editing sites in 33 mitochondrial genes was conducted on RNA extracted from rosette leaves. We detected 67 new editing events in A. thaliana rosette leaves that had not been observed in a prior study of mitochondrial editing in suspension cultures. Furthermore, 37 of the 441 C-to-U editing events reported in A. thaliana suspension cultures were not observed in rosette leaves. Forty editing sites that are polymorphic in extent of editing were detected between Col and Ler. Silent editing sites, which do not change the encoded amino acid, were found in a large excess compared to nonsilent sites among the editing events that differed between accessions and between tissue types. Dominance relationships were assessed for 15 of the most polymorphic sites by evaluating the editing values of the reciprocal hybrids. Dominance is more common in nonsilent sites than in silent sites, while additivity was observed only in silent sites. A maternal effect was detected for 8 sites. QTL mapping with recombinant inbred lines detected 12 major QTL for 11 of the 13 editing traits analyzed, demonstrating that efficiency of editing of individual mitochondrial C targets is generally governed by a major factor.     

ADAR2 A-->I editing: site selectivity and editing efficiency are separate events

ADAR enzymes, adenosine deaminases that act on RNA, form a family of RNA editing enzymes that convert adenosine to inosine within RNA that is completely or largely double-stranded. Site-selective A→I editing has been detected at specific sites within a few structured pre-mRNAs of metazoans. We have analyzed the editing selectivity of ADAR enzymes and have chosen to study the naturally edited R/G site in the pre-mRNA of the glutamate receptor subunit B (GluR-B). A comparison of editing by ADAR1 and ADAR2 revealed differences in the specificity of editing. Our results show that ADAR2 selectively edits the R/G site, while ADAR1 edits more promiscuously at several other adenosines in the double-stranded stem. To further understand the mechanism of selective ADAR2 editing we have investigated the importance of internal loops in the RNA substrate. We have found that the immediate structure surrounding the editing site is important. A purine opposite to the editing site has a negative effect on both selectivity and efficiency of editing. More distant internal loops in the substrate were found to have minor effects on site selectivity, while efficiency of editing was found to be influenced. Finally, changes in the RNA structure that affected editing did not alter the binding abilities of ADAR2. Overall these findings suggest that binding and catalysis are independent events.     

ADAR2 induces reproducible changes in sequence and abundance of mature microRNAs in the mouse brain

Adenosine deaminases that act on RNA (ADARs) deaminate adenosines to inosines in double-stranded RNAs including miRNA precursors. A to I editing is widespread and required for normal life. By comparing deep sequencing data of brain miRNAs from wild-type and ADAR2 deficient mouse strains, we detect editing sites and altered miRNA processing at high sensitivity. We detect 48 novel editing events in miRNAs. Some editing events reach frequencies of up to 80%. About half of all editing events depend on ADAR2 while some miRNAs are preferentially edited by ADAR1. Sixty-four percent of all editing events are located within the seed region of mature miRNAs. For the highly edited miR-3099, we experimentally prove retargeting of the edited miRNA to novel 3′ UTRs. We show further that an abundant editing event in miR-497 promotes processing by Drosha of the corresponding pri-miRNA. We also detect reproducible changes in the abundance of specific miRNAs in ADAR2-deficient mice that occur independent of adjacent A to I editing events. This indicates that ADAR2 binding but not editing of miRNA precursors may influence their processing. Correlating with changes in miRNA abundance we find misregulation of putative targets of these miRNAs in the presence or absence of ADAR2.     

基于PacBio测序数据的蜜蜂球囊菌转录因子、融合基因及RNA编辑事件的鉴定

【目的】本研究旨在利用已获得的PacBio单分子实时(single molecule real-time, SMRT)测序数据对蜜蜂球囊菌Ascosphaera apis菌丝(AaM)和孢子(AaS)中的转录因子(TF)、融合基因和RNA编辑事件进行鉴定和分析,以期丰富蜜蜂球囊菌的相关信息,并为进一步探究它们的功能提供理论依据。【方法】利用BLASTx工具将AaM和AaS的全长转录本序列比对到Nr, Swiss-Prot和KEGG数据库以获得一致性最高的蛋白序列,再利用hmmscan软件将上述蛋白序列比对到Plant TFdb数据库以获得TF的分类及注释信息。采用TOFU软件中的fusion_finder.py程序进行融合基因的预测,进而分析融合基因的序列和位置信息。使用SAMtools预测AaM和AaS中的RNA编辑事件,再利用ANNOVAR软件对RNA编辑事件进行注释,进而采用相关生物信息学软件对RNA编辑位点基因进行GO功能和KEGG通路注释。【结果】在AaS中共鉴定到17个TF家族的213个TF,其中C2H2家族包含的TF成员最多。在AaM和AaS中分别鉴定到921和510个融合基因,二者共有的融合基因为510个,特有的融合基因分别为411和0个。在AaM和AaS中分别鉴定到547和191次RNA编辑事件,其中AaM中同义单核苷酸突变的数量最多,AaS中非同义单核苷酸突变的数量最多。此外,在AaM中鉴定到12种碱基替换类型,其中发生C->T的RNA编辑事件数量最多,达到158次;在AaS中鉴定到9种碱基替换类型,其中发生C->T和G->T的RNA编辑事件数量最多,均有42次。AaM和AaS中RNA编辑位点基因分别涉及19和24个GO功能条目;此外还能注释到11和20条KEGG通路。【结论】蜜蜂球囊菌的菌丝和孢子中含有丰富的TF、融合基因和RNA编辑位点;转录因子C2H2家族与蜜蜂球囊菌菌丝和孢子的生长发育和细胞活动具有潜在关联;RNA编辑事件的碱基替换类型在蜜蜂球囊菌和其他物种中具有物种特异性;RNA编辑可能在蜜蜂球囊菌菌丝和孢子的生长和代谢中发挥作用。     

Virus-specific editing identification approach reveals the landscape of A-to-I editing and its impacts on SARS-CoV-2 characteristics and evolution

Upon SARS-CoV-2 infection, viral intermediates specifically activate the IFN response through MDA5-mediated sensing and accordingly induce ADAR1 p150 expression, which might lead to viral A-to-I RNA editing. Here, we developed an RNA virus-specific editing identification pipeline, surveyed 7622 RNA-seq data from diverse types of samples infected with SARS-CoV-2, and constructed an atlas of A-to-I RNA editing sites in SARS-CoV-2. We found that A-to-I editing was dynamically regulated, varied between tissue and cell types, and was correlated with the intensity of innate immune response. On average, 91 editing events were deposited at viral dsRNA intermediates per sample. Moreover, editing hotspots were observed, including recoding sites in the spike gene that affect viral infectivity and antigenicity. Finally, we provided evidence that RNA editing accelerated SARS-CoV-2 evolution in humans during the epidemic. Our study highlights the ability of SARS-CoV-2 to hijack components of the host antiviral machinery to edit its genome and fuel its evolution, and also provides a framework and resource for studying viral RNA editing.