ORCID Profile
0000-0001-9036-8180
Current Organisation
Nara Medical University
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Publisher: Elsevier BV
Date: 11-2009
DOI: 10.1016/J.YMETH.2009.04.010
Abstract: The laboratory rat is an invaluable animal model for biomedical research. However, mutant rat resource is still limited, and development of methods for large-scale generation of mutants is anticipated. We recently utilized the Sleeping Beauty (SB) transposon system to develop a rapid method for generating insertional mutant rats. Firstly, transgenic rats carrying single transgenes, namely the SB transposon vector and SB transposase, were generated. Secondly, these single transgenic rats were interbred to obtain doubly-transgenic rats carrying both transgenes. The SB transposon was mobilized in the germline of these doubly-transgenic rats, reinserted into another location in the genome and heterozygous mutant rats were obtained in the progeny. Gene insertion events were rapidly and non-invasively identified by the green fluorescence protein (GFP) reporter incorporated in the transposon vector, which utilizes a polyA-trap approach. Mutated genes were confirmed by either linker ligation-mediated PCR or 3'-rapid lification of cDNA ends (3'RACE). Endogenous expression profile of the mutated gene can also be visualized using the LacZ gene incorporated as a promoter-trap unit in the transposon vector. This method is straightforward, readily applicable to other transposon systems, and will be a valuable mutant rat resource to the biomedical research community.
Publisher: Elsevier BV
Date: 08-2012
DOI: 10.1016/J.IMMUNI.2012.06.009
Abstract: Interferon-γ (IFN-γ) is essential for host defense against intracellular pathogens. Stimulation of innate immune cells by IFN-γ upregulates ∼2,000 effector genes such as immunity-related GTPases including p65 guanylate-binding protein (Gbp) family genes. We show that a cluster of Gbp genes was required for host cellular immunity against the intracellular parasite Toxoplasma gondii. We generated mice deficient for all six Gbp genes located on chromosome 3 (Gbp(chr3)) by targeted chromosome engineering. Mice lacking Gbp(chr3) were highly susceptible to T. gondii infection, resulting in increased parasite burden in immune organs. Furthermore, Gbp(chr3)-deleted macrophages were defective in IFN-γ-mediated suppression of T. gondii intracellular growth and recruitment of IFN-γ-inducible p47 GTPase Irgb6 to the parasitophorous vacuole. In addition, some members of Gbp(chr3) restored the protective response against T. gondii in Gbp(chr3)-deleted cells. Our results suggest that Gbp(chr3) play a pivotal role in anti-T. gondii host defense by controlling IFN-γ-mediated Irgb6-dependent cellular innate immunity.
Publisher: Informa UK Limited
Date: 03-2007
DOI: 10.1128/MCB.01500-06
Publisher: Springer Science and Business Media LLC
Date: 14-01-2007
DOI: 10.1038/NMETH1002
Abstract: Although the laboratory rat (Rattus norvegicus) is an indispensable experimental animal for biomedical research and drug development, the lack of embryonic stem cell lines h ers gene-knockout studies. Here we report the successful generation of insertional mutant rats using the Sleeping Beauty (SB) transposon system. This would benefit a variety of biomedical research fields for which the rat model is better suited than the mouse model.
Publisher: Informa UK Limited
Date: 08-2006
DOI: 10.1128/MCB.00018-06
Publisher: Oxford University Press (OUP)
Date: 06-2007
DOI: 10.1534/GENETICS.107.071647
Abstract: Massive accumulation of retrotransposons, comprising & % of human and mouse genomes, is one of the major events in the evolution of the genome. However, most retrotransposons have lost retrotransposition competency, which makes studying their role in genome evolution elusive. Intracisternal A-particle (IAP) elements are long terminal repeat (LTR)-type mouse retrotransposons consisting of full-length and internally deleted types. Some are retrotransposition competent and their upregulated activity has been reported in mutant mice deficient in genome defense systems, suggesting that IAP elements provide a unique platform for studying the interaction between retrotransposons and mammalian genomes. Using the IAP element as a model case, here we show that mobilization of retrotransposons alters the mouse transcriptome. Retrotransposition assay in cultured cells demonstrated that a subset of internally deleted IAP elements, called IΔ1 type, retrotranspose efficiently when supplied with functional IAP proteins. Furthermore, the IΔ1 type IAP element exhibited substantial transcription-inducing activity in the flanking region. Genomewide transcript analysis of embryonic stem (ES) cells identified IAP-induced transcripts, including fusion transcripts between IAP sequence and endogenous genes. Unexpectedly, nearly half of these IAP elements obtained from ES cells derived from 129 mouse strain were absent in the C57BL/6 genome, suggesting that IAP-driven transcription contributes to the unique trait of the in idual mouse strain. On the basis of these data, we propose that retrotransposons are one of the drivers that shape the mammalian transcriptome.
Publisher: Informa UK Limited
Date: 12-2003
Publisher: Cold Spring Harbor Laboratory
Date: 02-05-2008
Abstract: Retrotransposons constitute a major component of the genome and their proliferation significantly impacts genome evolution. Retrotransposons can propagate autonomously or nonautonomously. Nonautonomous type transposition occurs through trans -complementation by autonomous type retrotransposons. While autonomous type retrotransposons have been studied extensively, the translation products from nonautonomous type retrotransposons are not well characterized. In a previous study, we isolated both autonomous and nonautonomous type intracisternal A particle (IAP) elements from the mouse genome and established a tissue culture assay to examine trans -complementation of nonautonomous type IAP element. Using this system in the present study, we determined an active role for the translation product from nonautonomous type IAP element. Point mutations that either eliminated or truncated the IAP protein were introduced and their effects on trans -complementation were examined. Trans -complementation efficiency correlated with the expression of nonautonomous type IAP protein. The effect of nonautonomous type IAP protein was observed only when it was provided in cis , suggesting an interaction of nonautonomous type IAP protein and its transcript immediately after transcription. Interaction of autonomous and nonautonomous type IAP proteins was demonstrated by immunostaining and coimmunoprecipitation assay. Based on these findings, we propose a model in which nonautonomous type IAP protein associates with its transcript, recruits autonomous type IAP protein, and promotes the assembly of transposition competent IAP particle. The active role of the nonautonomous type IAP protein revealed in this study may provide a new insight into retrotransposon proliferation within the genome.
Publisher: Springer Science and Business Media LLC
Date: 22-09-2005
DOI: 10.1038/NMETH795
Abstract: Recent consolidation of the whole-genome sequence with genome-wide transcriptome profiling revealed the existence of functional units within the genome in specific chromosomal regions, as seen in the coordinated expression of gene clusters and colocalization of functionally related genes. An efficient region-specific mutagenesis screen would greatly facilitate research in addressing the importance of these clusters. Here we use the 'local hopping' phenomenon of a DNA-type transposon, Sleeping Beauty (SB), for region-specific saturation mutagenesis. A transgenic mouse containing both transposon (acts as a mutagen) and transposase (recognizes and mobilizes the transposon) was bred for germ-cell transposition events, allowing us to generate many mutant mice. All genes within a 4-Mb region of the original donor site were mutated by SB, indicating the potential of this system for functional genomic studies within a specific chromosomal region.
Publisher: Springer Science and Business Media LLC
Date: 2007
No related grants have been discovered for Kyoji Horie.