Biomedical subjects

Shigeru Iwase

Publications and source records attributed to Shigeru Iwase.

3 recordsLinked to original sources

PubMed2006

Essential role for gene profiling analysis in the authentication of human cell lines.

Cross-contamination between cultured cell lines can result in the generation of erroneous scientific data. Hence, it is very important to eliminate cell lines that are of an origin different from that being claimed. Inter-species contamination can be detected by various established methods, such as karyotype and isozyme analyses. However, it has been impossible to detect intraspecies cross-contamination prior to the development of technology to detect differences between cell lines at the molecular level. Recently, profiling of short tandem repeat (STR) polymorphisms has been established as a method for the analyses of gene polymorphism. Gene profiling by STR polymorphism (STR profiling) is a simple and reliable method to identify individual cell lines. Each human cell line currently provided by the Cell Engineering Division of the RIKEN BioResource Center was analyzed by STR profiling to authenticate its identity. We found that more than 10 human cell lines out of approximately 400 were in fact identical to a different cell line deposited in the collection, and therefore had been misidentified. We conclude that STR profiling is a useful and powerful method for eliminating cell lines that have been misidentified by cross-contamination or by other causes. Hence, STR profiling of human cell lines used in published research will likely be a prerequisite for publication in the future, so that the problem of misidentification of cell lines can be eliminated.

Cell Line↗

PubMed2005

Transient strong reduction of PTEN expression by specific RNAi induces loss of adhesion of the cells.

The tumor suppressor gene pten encodes a lipid phosphatase that dephosphorylates D3 of phosphatidylinositol(3,4,5)trisphosphate, producing phosphatidylinositol(4,5)bisphosphate. Although PTEN has been implicated in cell adhesion and migration, the underlying molecular mechanism is unknown. To investigate the role of PTEN in cell adhesion, we designed three different siRNAs (siRNA PTEN-a, siRNA PTEN-b, and siRNA PTEN-c) and transfected into 293T cells. Two days later, only the cells transfected with siRNA PTEN-b became round and detached from the culture dishes, whereas cells transfected with a control siRNA against GFP or the two other siRNAs against PTEN did not. Evaluation of the RNAi effect revealed that siRNA PTEN-b inhibited >95% of PTEN expression, the most effective among the three siRNAs. To check for non-specific effects such as interferon response and inhibition of off-target genes, we then used quantitative PCR analysis and DNA microarray analysis. None was detected, indicating that the RNAi system was highly specific. Immunofluorescence studies using PTEN-knockdown HeLa cells revealed that the loss of adhesion was accompanied by a reduction in the number of focal adhesion plaques and disorganization of the actin cytoskeleton. Transient and near-complete loss of PTEN expression induces loss of adhesion of the cells.

Apoptosis↗

PubMed2005

Disclosing hidden transcripts: mouse natural sense-antisense transcripts tend to be poly(A) negative and nuclear localized.

Genome-wide in silico analysis identified thousands of natural sense-antisense transcript (SAT) pairs in the mouse transcriptome. We investigated their expression using strand-specific oligo-microarray that distinguishes expression of sense and antisense RNA from 1947 SAT pairs. The majority of the predicted SATs are expressed at various steady-state levels in various tissues, and cluster analysis of the array data demonstrated that the ratio of sense and antisense expression for some of the SATs fluctuated markedly among these tissues, while the rest was unchanged. Surprisingly, further analyses indicated that vast amounts of multiple-sized transcripts are expressed from the SAT loci, which tended to be poly(A) negative, and nuclear localized. The tendency that the SATs are often not polyadenylated is conserved, even in the randomly chosen SAT genes in the plant Arabidopsis thaliana. Such common characteristics imply general roles of the SATs in regulation of gene expression.

Animals↗