RIP-seq of BmAgo2-associated small RNAs reveal various types of small non-coding RNAs in the silkworm, Bombyx moriReport as inadecuate




RIP-seq of BmAgo2-associated small RNAs reveal various types of small non-coding RNAs in the silkworm, Bombyx mori - Download this document for free, or read online. Document in PDF available to download.

BMC Genomics

, 14:661

Multicellular invertebrate genomics

Abstract

BackgroundSmall non-coding RNAs ncRNAs are important regulators of gene expression in eukaryotes. Previously, only microRNAs miRNAs and piRNAs have been identified in the silkworm, Bombyx mori. Furthermore, only ncRNAs 50-500nt of intermediate size have been systematically identified in the silkworm.

ResultsHere, we performed a systematic identification and analysis of small RNAs 18-50nt associated with the Bombyx mori argonaute2 BmAgo2 protein. Using RIP-seq, we identified various types of small ncRNAs associated with BmAGO2. These ncRNAs showed a multimodal length distribution, with three peaks at ~20nt, ~27nt and ~33nt, which included tRNA-, transposable element TE-, rRNA-, snoRNA- and snRNA-derived small RNAs as well as miRNAs and piRNAs. The tRNA-derived fragments tRFs were found at an extremely high abundance and accounted for 69.90% of the BmAgo2-associated small RNAs. Northern blotting confirmed that many tRFs were expressed or up-regulated only in the BmNPV-infected cells, implying that the tRFs play a prominent role by binding to BmAgo2 during BmNPV infection. Additional evidence suggested that there are potential cleavage sites on the D, anti-codon and TψC loops of the tRNAs. TE-derived small RNAs and piRNAs also accounted for a significant proportion of the BmAgo2-associated small RNAs, suggesting that BmAgo2 could be involved in the maintenance of genome stability by suppressing the activities of transposons guided by these small RNAs. Finally, Northern blotting was also used to confirm the Bombyx 5.8 s rRNA-derived small RNAs, demonstrating that various novel small RNAs exist in the silkworm.

ConclusionsUsing an RIP-seq method in combination with Northern blotting, we identified various types of small RNAs associated with the BmAgo2 protein, including tRNA-, TE-, rRNA-, snoRNA- and snRNA-derived small RNAs as well as miRNAs and piRNAs. Our findings provide new clues for future functional studies of the role of small RNAs in insect development and evolution.

KeywordsBombyx mori BmAgo2-associated small RNAs RIP-seq tRFs BmNPV AbbreviationsncRNAsnon-coding RNAs

BmAgo2Bombyx mori argonaute2

tRFstRNA-derived fragments

miRNAsmicroRNAs

siRNAssmall-interfering RNAs

piRNAspiwi-interacting RNAs

IPImmunoprecipitation

RIPRNA immunoprecipitation

TETransposable element

RISCRNA-induced silencing complex

AcNPVAutographa Californica nucleopolyhedrovirus

BmNPVBombyx mori nucleopolyhedrovirus

PAGEPolyacrylamide gel.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2164-14-661 contains supplementary material, which is available to authorized users.

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Author: Zuoming Nie - Fang Zhou - Dan Li - Zhengbing Lv - Jian Chen - Yue Liu - Jianhong Shu - Qing Sheng - Wei Yu - Wenping Zha

Source: https://link.springer.com/







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