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Watanabe Y.,RIKEN | Watanabe Y.,University of Tsukuba | Numata K.,RIKEN | Murata S.,Keio University | And 10 more authors.

The functionality of sense-antisense transcripts (SATs), although widespread throughout the mammalian genome, is largely unknown. Here, we analyzed the SATs expression and its associated promoter DNA methylation status by surveying 12 tissues of mice to gain insights into the relationship between expression and DNA methylation of SATs. We have found that sense and antisense expression positively correlate in most tissues. However, in some SATs with tissue-specific expression, the expression level of a transcript from a CpG island-bearing promoter is low when the promoter DNA methylation is present. In these circumstances, the expression level of its opposite-strand transcript, especially when it is poly(A)-negative was coincidentally higher. These observations suggest that, albeit the general tendency of sense-antisense simultaneous expression, some antisense transcripts have coordinated expression with its counterpart sense gene promoter methylation. This cross-strand relationship is not a privilege of imprinted genes but seems to occur widely in SATs. © 2010 Elsevier Inc. Source

Saito R.,Keio University | Kohno K.,University of Tsukuba | Okada Y.,Keio University | Osada Y.,Keio University | And 15 more authors.
BMC Medical Genomics

Background: Recent studies have identified thousands of sense-antisense gene pairs across different genomes by computational mapping of cDNA sequences. These studies have shown that approximately 25% of all transcriptional units in the human and mouse genomes are involved in cis-sense-antisense pairs. However, the number of known sense-antisense pairs remains limited because currently available cDNA sequences represent only a fraction of the total number of transcripts comprising the transcriptome of each cell type. Methods. To discover novel antisense transcripts encoded in the antisense strand of important genes, such as cancer-related genes, we conducted expression analyses of antisense transcripts using our custom microarray platform along with 2376 probes designed specifically to detect the potential antisense transcripts of 501 well-known genes suitable for cancer research. Results: Using colon cancer tissue and normal tissue surrounding the cancer tissue obtained from 6 patients, we found that antisense transcripts without poly(A) tails are expressed from approximately 80% of these well-known genes. This observation is consistent with our previous finding that many antisense transcripts expressed in a cell are poly(A)-. We also identified 101 and 71 antisense probes displaying a high level of expression specifically in normal and cancer tissues respectively. Conclusion: Our microarray analysis identified novel antisense transcripts with expression profiles specific to cancer tissue, some of which might play a role in the regulatory networks underlying oncogenesis and thus are potential targets for further experimental validation. Our microarray data are available at http://www.brc.riken.go.jp/ncrna2007/viewer-Saito-01/index.html. © 2011 Saito et al; licensee BioMed Central Ltd. Source

Akiyama H.,Toray Industries Inc | Ueda Y.,Toray Industries Inc | Nobumasa H.,Toray Industries Inc | Ooshima H.,Mitsubishi Group | And 13 more authors.
Analytical Biochemistry

RNA external standards, although important to ensure equivalence across many microarray platforms, have yet to be fully implemented in the research community. In this article, a set of unique RNA external standards (or RNA standards) and probe pairs that were added to total RNA in the samples before amplification and labeling are described. Concentration-response curves of RNA external standards were used across multiple commercial DNA microarray platforms and/or quantitative real-time polymerase chain reaction (RT-PCR) and next-generation sequencing to identify problematic assays and potential sources of variation in the analytical process. A variety of standards can be added in a range of concentrations spanning high and low abundances, thereby enabling the evaluation of assay performance across the expected range of concentrations found in a clinical sample. Using this approach, we show that we are able to confirm the dynamic range and the limit of detection for each DNA microarray platform, RT-PCR protocol, and next-generation sequencer. In addition, the combination of a series of standards and their probes was investigated on each platform, demonstrating that multiplatform calibration and validation is possible. © 2014 Elsevier Inc. All rights reserved. Source

Yamane T.,Hokkaido University | Yamane T.,Shiga University of Medical Science | Murao S.,Hokkaido University | Kato-Ose I.,Hokkaido University | And 8 more authors.
Biochemical and Biophysical Research Communications

Legumain (EC is an asparaginyl endopeptidase. Strong legumain activity was observed in the mouse kidney, and legumain was found to be highly expressed in tumors. We previously reported that bovine kidney annexin A2 was co-purified with legumain and that legumain cleaved the N-terminal region of annexin A2 at an Asn residue in vitro and in vivo. In this study, we found a p53-binding site in intron 1 of the human legumain gene using computational analysis. To determine whether transcription of the legumain gene is regulated by p53, HCT116 cells were transfected with p53 siRNA and the effect of knockdown of p53 expression on legumain expression was examined. The results showed that expression levels of both legumain mRNA and protein were decreased in the siRNA-treated cells. Furthermore, enzyme activity of legumain was also increased by doxorubicin and its activity was reduced by knockdown of p53 in HCT116 cells. These results suggest that legumain expression and its enzyme activity are regulated by p53. © 2013 Elsevier Inc. Source

Matsuura H.,Aichi University of Technology | Hokonohara H.,Osaka University | Sugita T.,Hokkaido System Science Co. | Takagi A.,Osaka University | And 3 more authors.
Journal of Applied Physics

This paper investigates the roles of a solution (decane) in deoxyribonucleic acid (DNA) observation with a scanning tunneling microscope. Our study indicates that decane prevents continuous water adsorption from air and subsequent ionization of the water to create specific conditions for DNA observation. Analysis of the tunneling current reveals that the current with decane became twice as stable in deviation and the current is sustained 1 nm further in the z-direction than without decane. The apparent barrier height with decane is also decreased by a factor of 0.18. These properties enable us to measure bulky DNA (4 nm) at the highest success ratio ever attained. © 2011 American Institute of Physics. Source

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