Retrovirus-mediated gene transfer and expression cloning: powerful tools in functional genomics

Kitamura, Toshio; Koshino, Yuko; Shibata, Fumi; Oki, Toshihiko; Nakajima, Hitoshi; Nosaka, Tetsuya; Kumagai, Hidetoshi

doi:10.1016/s0301-472x(03)00260-1

Cited by 604 publications

(182 citation statements)

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“…In summary, we identified RasGRP4 as a candidate gene of class I mutations by our expression cloning strategy based on retrovirus-mediated gene transfer [31,32,39]. Although we did not find significant mutations in RasGRP4 derived from patients, overexpression of RasGRP4 confers factor independency on an IL-3 dependent cell line and induced T cell leukemia and myeloid leukemia in a mouse BMT model.…”

Section: Discussionmentioning

confidence: 76%

“…The point mutations of AML1 were not screened. Recombinant retroviruses were generated by transient transfection using an ecotropic packaging cell line PLAT-E as described with minor modifications [32]. Bone marrow or peripheral blood samples of patients were taken under the experimental procedure approved by the ethical committees of our institute (approve no.…”

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

“…We subcloned the PCR products into TOPO vector (Invitrogen, San Diego, CA). Then, the EcoRI fragment carrying Ras-GRP4 was inserted into the EcoRI sites of pMXs vector [32]. RasGRP4 sequences derived from patients and normal volunteers were not identical to those in the data bases as described in result section.…”

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

“…This mutant is hereafter referred to as AML1-S291fs. The AML1-S291fs was inserted upstream of the IRES-EGFP cassette of a retrovirus vector pMYs-IG [32] to generate pMYs-AML1-S291fs-IG.…”

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

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Possible involvement of RasGRP4 in leukemogenesis

et al. 2009

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It is now conceivable that leukemogenesis requires two types of mutations, class I and class II mutations. We previously established a mouse bone marrow-derived HF6, an IL-3-dependent cell line, that was immortalized by a class II mutation MLL/SEPT6 and can be fully transformed by class I mutations such as FLT3 mutants. To understand the molecular mechanism of leukemogenesis, particularly progression of myelodysplastic syndrome (MDS) to acute leukemia, we made cDNA libraries from the samples of patients and screened them by expression-cloning to detect class I mutations that render HF6 cells factor-independent. We identified RasGRP4, an activator of Ras, as a candidate for class I mutation from three of six patients (MDS/MPD = 1, MDS-RA = 1, MDS/AML = 2, CMMoL/AML = 1 and AML-M2 = 1). To investigate the potential roles of RasGRP4 in leukemogenesis, we tested its in vivo effect in a mouse bone marrow transplantation (BMT) model. C57BL/6J mice transplanted with RasGRP4-transduced primary bone marrow cells died of T cell leukemia, myeloid leukemia, or myeloid leukemia with T cell leukemia. To further examine if the combination of class I and class II mutations accelerated leukemic transformation, we performed a mouse BMT model in which both AML1 mutant (S291fsX300) and RasGRP4 were transduced into bone marrow cells. The double transduction led to early onset of T cell leukemia but not of AML in the transplanted mice when compared to transduction of RasGRP4 alone. Thus, we have identified RasGRP4 as a gene potentially involved in leukemogenesis and suggest that RasGRP4 cooperates with AML1 mutations in T cell leukemogenesis as a class I mutation.

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Section: Discussionmentioning

confidence: 76%

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

“…This mutant is hereafter referred to as AML1-S291fs. The AML1-S291fs was inserted upstream of the IRES-EGFP cassette of a retrovirus vector pMYs-IG [32] to generate pMYs-AML1-S291fs-IG.…”

Section: Screening Of Cdna Librariesmentioning

confidence: 99%

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Possible involvement of RasGRP4 in leukemogenesis

et al. 2009

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“…Hamster anti-mouse Tim4 monoclonal antibody (MAb) (clone Kat5-18) was described previously (10 (22) and pENV-IRES-puro (23) were provided by T. Kitamura (Institute of Medical Science, University of Tokyo). Lentiviral expression vectors (CSII-EF, pCAG-HIVgp, pENV-IRES-puro, and pRSV-Rev) were from H. Miyoshi, Riken Resource Center.…”

mentioning

confidence: 99%

Tim4- and MerTK-Mediated Engulfment of Apoptotic Cells by Mouse Resident Peritoneal Macrophages

Nishi

Toda

Segawa

et al. 2014

Molecular and Cellular Biology

122

125

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Hepatocyte nuclear factor 1 beta induces transformation and epithelial‐to‐mesenchymal transition

et al. 2016

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Gene amplification can be a cause of cancer, and driver oncogenes have been often identified in amplified regions. However, comprehensive analysis of other genes coamplified with an oncogene is rarely performed. We focused on the 17q12-21 amplicon, which contains ERBB2. We established a screening system for oncogenic activity with the NMuMG epithelial cell line. We identified a homeobox gene, HNF1B, as a novel cooperative transforming gene. HNF1B induced cancerous phenotypes, which were enhanced by the coexpression of ERBB2, and induced epithelial-to-mesenchymal transition and invasive phenotypes. These results suggest that HNF1B is a novel oncogene that can work cooperatively with ERBB2.

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Retrovirus-mediated gene transfer and expression cloning: powerful tools in functional genomics

Cited by 604 publications

References 0 publications

Possible involvement of RasGRP4 in leukemogenesis

Possible involvement of RasGRP4 in leukemogenesis

Tim4- and MerTK-Mediated Engulfment of Apoptotic Cells by Mouse Resident Peritoneal Macrophages

Hepatocyte nuclear factor 1 beta induces transformation and epithelial‐to‐mesenchymal transition

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