Motoharu Ono- PhD
- Research Associate at University of Dundee
Motoharu Ono
- PhD
- Research Associate at University of Dundee
About
40
Publications
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Introduction
I'm a researcher and also an inventor. I develop a vector system termed snoMEN based on small nucleolar RNAs (snoRNAs), which are a type of small non-coding RNAs, to modulate targeted gene expression in mammalian cells, especially in human cells. I also study new biological functions of snoRNA/snoMEN biogenesis using inducible cancer/tumor model human cell lines with SILAC proteomics method. We foresee future applications for snoMEN vectors in basic research and possibly also for gene therapy.
Current institution
Additional affiliations
June 2004 - present
April 1999 - March 2004
Education
April 1995 - March 1999
Publications
Publications (40)
The snoMEN (snoRNA Modulator of gene ExpressioN) vector technology was developed from a human box C/D snoRNA, HBII-180C, which contains an internal sequence that can be manipulated to make it complementary to RNA targets, allowing knock-down of targeted genes. Here we have screened additional human nucleolar snoRNAs and assessed their application f...
Supporting Table of SILAC analysis.
(XLSX)
Supporting Table of in silico analysis.
(DOCX)
We have previously reported an antisense technology, 'snoMEN vectors', for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the sp...
We have identified the human FMN2 gene as a novel target regulated by induction of p14ARF and by multiple other stress responses, including DNA damage and hypoxia, which have in common activation of cell cycle arrest. We showed that increased expression of the FMN2 gene following p14ARF induction is caused, at the transcriptional level, by relief o...
The study of the function of many human proteins is often hampered by technical limitations, such as cytotoxicity and phenotypes that result from overexpression of the protein of interest together with the endogenous version. Here we present the snoMEN (snoRNA Modulator of gene ExpressioN) vector technology for generating stable cell lines where ex...
Gene-expression profile of snoMEN replacement stable cell lines. (A) Distribution pattern of GFP-SMN1 signal intensity of U2OSGFP–SMN1-PR stable cell line. Cytoplasmic GFP signals were calculated from randomly selected cells (n = 42). Each signal was normalised by DAPI signal. (B) Expression level comparison of proteins detected by Mass spectrometr...
Characterisation of FP–protein complexes in replacement stable cell lines by Quantitative SILAC Proteomic analysis. (A) SILAC result of GFP–SMN1 complex pull-down assay visualised on a 2D logarithmic graph. On the x axis, log2 (H/L ratio) correlates with the enrichment in GFP–SMN1 IP with protein replacement versus control IP. On the y axis, log2 (...
The list of SILAC ratio values of labelled proteins described in
Figure 2E
.
(XLSX)
The list of SILAC ratio values of top 10% labelled proteins described in Figure S5A.
(XLSX)
SiRNA and shRNA knock-down targeted to endogenous SMN1 pre-mRNAs. (A–D) These are the same experiment as in Figure 3C–F except the target gene is SMN1 in U2OS cells. (A) Scrambled siRNA (Negative control siRNA) and SMN1 siRNA (Dharmacon) were transfected as a negative and a positive control, respectively. SMN1 Mbox siRNA-1 to -3 (siSM1–3) have the...
SnoRNA expression analysis after Fibrillarin knock-down treatment. qRT-PCR was performed to measure snoRNA expression level after treatment with scramble siRNA (Control) and Fibrillarin siRNA. Equal amounts of total RNA from U2OSGFP–SMN1-PR cells, extracted following siRNA treatment, was used for qRT-PCR reactions. Graph shows the snoRNA expression...
Optimisation of RNA polymerase I inhibition using low concentration Actinomycin-D treatment. (A) In vivo transcription assay with/without Actinomycin-D treatment in HeLa cells. HeLa cells were treated either with ethanol (EtOH) as a negative control, or with Actinomycin-D (0.01 µg/ml) for each time point: 30 min, 1 hr, and 2 hr. Transcription in th...
The list of SILAC ratio values of top 10% labelled proteins described in
Figure 5B
.
(XLSX)
The list of SILAC ratio values of labelled proteins described in Figure S1B.
(XLSX)
The present invention relates to a method of targeting miRNA and/or pre-miRNA molecules in order to treat diseases that are linked with miRNA expression, such as certain cancers. The present invention also provides modified snoRNA molecules for targeting mi RNA molecules for use in treating diseases that are linked with miRNA expression, such as ce...
The ARF tumor suppressor is a central component of the cellular defense against oncogene activation in mammals. p14ARF activates p53 by binding and inhibiting HDM2, resulting, inter alia, in increased transcription and expression of the cyclin-dependent kinase inhibitor p21 and consequent cell-cycle arrest. We analyzed the effect of p14ARF inductio...
Small nucleolar RNAs (snoRNAs) function mainly as guides for the post-transcriptional modification of ribosomal RNAs (rRNAs).
In recent years, several studies have identified a wealth of small fragments (<35 nt) derived from snoRNAs (termed sdRNAs)
that stably accumulate in the cell, some of which may regulate splicing or translation. A comparison...
The present invention provides methods for diagnosing cell proliferation and/or differentiation disorders, compounds and methods for treating the same and methods for identifying agents potentially useful in the treatment of cell proliferation and/or differentiation disorders.
Small nucleolar RNAs (snoRNAs) are an ancient class of small non-coding RNAs present in all eukaryotes and a subset of archaea that carry out a fundamental role in the modification and processing of ribosomal RNA. In recent years, however, a large proportion of snoRNAs have been found to be further processed into smaller molecules, some of which di...
There are two main classes of small nucleolar RNAs (snoRNAs): the box C/D snoRNAs and the box H/ACA snoRNAs that function
as guide RNAs to direct sequence-specific modification of rRNA precursors and other nucleolar RNA targets. A previous computational
and biochemical analysis revealed a possible evolutionary relationship between miRNA precursors...
Human small nucleolar RNAs (snoRNAs) that copurify with nucleoli isolated from HeLa cells have been characterized. Novel fibrillarin-associated snoRNAs were detected that allowed the creation of a new vector system for the targeted knockdown of one or more genes in mammalian cells. The snoMEN (snoRNA modulator of gene expressioN) vector technology...
MicroRNAs (miRNAs) and small nucleolar RNAs (snoRNAs) are two classes of small non-coding regulatory RNAs, which have been much investigated in recent years. While their respective functions in the cell are distinct, they share interesting genomic similarities, and recent sequencing projects have identified processed forms of snoRNAs that resemble...
The present invention relates to a method of modulating gene expression using snoRNA molecules or snoRNA like molecules or fragments, designed to target specific nucleic acid sequences.
The hibernation-specific HP-27 gene is expressed specifically in the liver of the chipmunk, a hibernating species of the squirrel family, and exists as a pseudogene in the tree squirrel, a nonhibernating species. In the promoter region, the chipmunk gene has a potential HNF-1 binding site, and the tree squirrel gene has two base substitutions in th...
The chipmunk hibernation-specific protein HP-27 is a component of the 140-kDa complex that decreases in the blood during hibernation. Although the HP-27 gene is detected in both the chipmunk, a hibernating species of the squirrel family, and the tree squirrel, a nonhibernating species, it is expressed only in the chipmunk, in a liver-specific manne...
The chipmunk hibernation-specific protein HP-20 is a component of the 140 kDa complex that drastically decreases in the blood during hibernation, and its gene is expressed specifically in the liver. To reveal molecular mechanisms underlying the liver-specific transcription of the HP-20 gene, we isolated chipmunk HP-20 genomic clones. The HP-20 gene...
Questions
Questions (2)
I’m trying to detect RNA modification of low abundant species in mammalian cells. I am wondering whether there are any protocols/kits for a high sensitivity and high throughput detection of RNA modifications, i.e. 2'O-ribose methylation and pseudouridylation. More specifically, I’m wondering whether there have been any qRT/RT-PCR methods to detect them in recent years.
I'm trying to take a TEM picture of my lenti-/adeno-virus using negative staining. However, we can only find a few viruses on a grid for some reason. I purified the virus and concentrated. I tested the virus titer of exactly the same batch and they are fine and very strong. I also tried to evaporate virus solution on a grid to increase attachment of virus, however, the result didn't change.
If I look at them on an SEM chip using the same methods I have seen a lot of ball shapes made by a lot of tiny virus size balls. Are they an aggregated virus or a just crystallized something?
