2018 Fiscal Year Final Research Report
Development of a method for medium-length RNA, and functional analysis of SINE non-coding RNAs
| Project/Area Number |
16K14784
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| Research Category |
Grant-in-Aid for Challenging Exploratory Research
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| Allocation Type | Multi-year Fund |
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| Research Field |
Genetics/Chromosome dynamics
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| Research Institution | Nagoya University |
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Principal Investigator |
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| Research Collaborator |
Ichiyanagi Tomoko
Mori Yoshinobu
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Keywords | RNA / 大規模シーケンシング / SINE / tRNA / snRNA / rRNA |
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| Outline of Final Research Achievements |
No technique was available to comprehensively analyze the expression levels of RNAs of medium length, such as tRNA, 5S rRNA, spiceosomal RNA, and SINE RNA. In this study, we have developed a protocol to do this, which we designated as meRNA-seq. The protocol involves processing of the 5’ end of RNAs by the RppH enzyme to make a monophosphated end, specific adaptor ligation to the 5’ and 3’ ends, respectively, PCR, and deep sequencing on Illumina MiSeq in a single-end sequencing mode of 300 bp. We then focused on SINE RNA expression, and revealed that SINEs show tissue-specific expression pattern with very high expression in spermatogonia (male germ cells). Interestingly, even in spermatogonia, loss of DNA methylation or piRNA resulted in upregulation of SINEs, suggesting that SINE transcription by RNA polymerase III is epigenetically regulated.
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| Free Research Field |
エピジェネティクス
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| Academic Significance and Societal Importance of the Research Achievements |
本研究によって、これまで解析が不可能であったAlu, B1, B2などのSINEレトロトランスポゾンの発現量について、ローカスを区別しながら、しかも定量的に解析できるようになった。特にAluの発現は癌化や神経疾患などの病態と関連があることが知られている。Aluはゲノムに１００万コピーあり、その配列は少しずつ異なっているが、この解析方法を使えば、Aluが発現しているかどうかを全体で見るだけでなく、どんな配列をもったどのゲノムローカスにAluなのかというように個々に解析できる。また、細胞分化等に伴ってtRNAレパートリーがどのように変化するかなど、これまで不可能であった解析を可能にした。
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