Postdoctoral Researcher

Dr Moto Ono

mono@dundee.ac.uk
Moto Ono
Moto Ono
Moto Ono

Project Description


Our project is aimed to understand novel human snoRNA (small nucleolar RNA) functions and also development of snoMEN(snoRNA Modulator of gene ExpressioN) vector. We are analysing RNA species that co-purify with nucleoli isolated from cultured human cells. This has identified novel snoRNAs. Based on these findings we have created a vector system for the targeted knock-down of one or more genes in mammalian cells [1](Ono M. and Lamond A.I., 2008, Patent, WO/2009/037490). The snoMENvector technology is based on the human Box C/D snoRNA we identified called HBII-180C, which contains an internal sequence that can be manipulated to make it complementary to RNA targets (Fig. 1)[6]. This approach allows for knock-down of two or more targeted genes simultaneously from a single vector and provides for parallel “transient/stable knock-in” of tagged or mutated replacement proteins from the same vector and potentially offers advantages over currently used strategies for both transient and stable protein knock-down. We foresee applications for snoMEN vectors in basic gene expression research, target validation and gene therapy in future. Furthermore, based on these findings, we have also investigated a relationship between snoRNAs and miRNA (microRNA) precursors using computational and biochemical analyses. This suggested that a subset of small regulatory RNAs may have evolved from snoRNAs [2,3,4,5]. Based on these findings, we will further develop snoMENvectors and analyse novel snoRNA mechanisms/functions in human cells.


Japanese

私達の研究目標はヒトの核小体RNA(snoRNA)の新規機能の解析及び、snoMEN™ (snoRNA Modulator of gene ExpressioN)ベクターの開発です。その手法として、ヒトの培養細胞より精製した核小体に含まれるRNA郡を抽出し、詳しく研究しています。これまでの解析結果から、新規snoRNAを同定し、その発見を基に、新規の遺伝子抑制ベクター、snoMEN™を開発しました[1] (Ono M. and Lamond A.I., 2008, 特許, WO/2009/037490)。snoMEN™ベクターはヒトsnoRNAのBox C/D郡に属するHBII-180Cを基に作られており、内部配列に標的遺伝子と相補的に結合できる領域をもっています(図1)。この技術を用いると1つのベクターで、同時に複数の標的遺伝子の遺伝子抑制、また人為的な遺伝子導入が、一過的にも恒常的にも可能であり、既存の遺伝子抑制系で困難であった問題を解決することができます(Ono et al., 2013)。更に私達はsnoMEN™ベクターを基礎生物学研究及び、将来的には遺伝子治療に役立てる事を目指しています。また、その他にも 核小体RNAの解析結果を基に、snoRNAとmicroRNA前駆体の関連について、生化学的及びバイオインフォマティクス的手法を用いて解析しています。これらの解析から、現在までに 一部のmicroRNA郡がsnoRNA郡から進化した痕跡を発見しました[2,3,4,5]。現在はこれらの発見をsnoMEN™技術の向上に役立て、ヒトsnoRNAの新たな機能解析に応用して研究を進めています。


Patents

Ono, M. and Lamond, A.I., (2008). Targeted moduration of gene expression. International patent specification, WO/2009/037490.

Yamada, K., Ono, M., Rocha, S., Lamond, A.I. (2010). Diagnosis and treatment of cell proliferation and differentiation disorders based on the fmn2 gene. International patent specification, WO/2010/103284.

Ono, M., Yamada, K., and Lamond, A.I., (2013). Targeting of miRNA precursors. WO/2013/054113.


References

1. Ono M, Yamada K, Avolio F, Scott MS, van Koningsbruggen S, et al. (2010) Analysis of human small nucleolar RNAs (snoRNA) and the development of snoRNA modulator of gene expression vectors. Mol Biol Cell 21: 1569-1584.

2. Ono M*, Scott MS*, Yamada K, Avolio F, Barton GJ, et al. (2011) Identification of human miRNA precursors that resemble box C/D snoRNAs. Nucleic Acids Res 39: 3879-3891.

3. Scott MS, Avolio F, Ono M, Lamond AI, Barton GJ (2009) Human miRNA precursors with box H/ACA snoRNA features. PLoS Comput Biol 5: e1000507.

4. Scott MS, Ono M (2011) From snoRNA to miRNA: Dual function regulatory non-coding RNAs. Biochimie 93: 1987-1992.

5. Scott MS*, Ono M*, Yamada K, Endo A, Barton GJ, et al. (2011) Human box C/D snoRNA processing conservation across multiple cell types. Nucleic Acids Res.

6. Ono M*, Yamada K*, Endo A*, Avolio F, Lamond AI (2013) Analysis of human protein replacement stable cell lines established using snoMEN-PR vector. PloS one 8: e62305. [Selected for the best poster prize at GRE symposium 2013]

7. Yamada K, Ono M, Bensaddek D, Lamond AI, Rocha S (2013) FMN2 is a novel regulator of the cyclin-dependent kinase inhibitor p21. Cell cycle 12: 2348-2354.

8. Yamada K*, Ono M*, Perkins ND, Rocha S, Lamond AI (2013) Identification and functional characterization of FMN2, a regulator of the cyclin-dependent kinase inhibitor p21. Molecular cell 49: 922-933. [Won the Haward Elder Prize 2013]

* Joint authorship.

Figure
Figure 1. Features of snoMEN technology
Schematic diagram showing differences between the siRNA/shRNA and snoMEN systems. Arrows show promoters for RNA polymerase III (shRNA) and RNA polymerase II (snoMEN), respectively. Red squares show the coding region, e.g. either mCherry cDNA, or endogenous genes. Striped squares show non-coding exon region. The bars show non-coding regions, e.g. introns.
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