Direct Hematological Toxicity and Illegitimate Chromosomal Recombination Caused by the Systemic Activation of CreERT2

Higashi, Atsuko Y.; Ikawa, Tomokatsu; Muramatsu, Masamichi; Economides, Aris N.; Niwa, Akira; Okuda, Tomohiko; Murphy, Andrew; Rojas, José Luis Vázquez; Heike, Toshio; Nakahata, Tatsutoshi; Kawamoto, Hiroshi; Kita, Toru; Yanagita, Motoko

doi:10.4049/jimmunol.0802413

Cited by 128 publications

(130 citation statements)

References 43 publications

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“…Ectopic and continued expression of recombinases may be immunogenic, impair cassette exchange, and induce genotoxic off-target events (25). DNA-based approaches for delivery of recombinases, such as those relying on adenoviral vectors (26) or transient transfection (27), are popular but cannot avoid the risk of residual transgene integration.…”

Section: Discussionmentioning

confidence: 99%

Protein transduction from retroviral Gag precursors

Voelkel¹,

Galla²,

Maetzig³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

125

118

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Retroviral particles assemble a few thousand units of the Gag polyproteins. Proteolytic cleavage mediated by the retroviral protease forms the bioactive retroviral protein subunits before cell entry. We hypothesized that this process could be exploited for targeted, transient, and dose-controlled transduction of nonretroviral proteins into cultured cells. We demonstrate that gammaretroviral particles tolerate the incorporation of foreign protein at several positions of their Gag or Gag-Pol precursors. Receptor-mediated and thus potentially cell-specific uptake of engineered particles occurred within minutes after cell contact. Dose and kinetics of nonretroviral protein delivery were dependent upon the location within the polyprotein precursor. Proteins containing nuclear localization signals were incorporated into retroviral particles, and the proteins of interest were released from the precursor by the retroviral protease, recognizing engineered target sites. In contrast to integration-defective lentiviral vectors, protein transduction by retroviral polyprotein precursors was completely transient, as protein transducing retrovirus-like particles could be produced that did not transduce genes into target cells. Alternatively, bifunctional protein-delivering particle preparations were generated that maintained their ability to serve as vectors for retroviral transgenes. We show the potential of this approach for targeted genome engineering of induced pluripotent stem cells by delivering the site-specific DNA recombinase, Flp. Protein transduction of Flp after proteolytic release from the matrix position of Gag allowed excision of a lentivirally transduced cassette that concomitantly expresses the canonical reprogramming transcription factors (Oct4, Klf4, Sox2, c-Myc) and a fluorescent marker gene, thus generating induced pluripotent stem cells that are free of lentivirally transduced reprogramming genes.Flp recombinase | murine leukemia virus | pluripotent cells | protein transfer

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Section: Discussionmentioning

confidence: 99%

Protein transduction from retroviral Gag precursors

Voelkel¹,

Galla²,

Maetzig³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

125

118

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“…This review cannot possibly cover every Cre line that exhibits expression in the testis (for an expanded list, see Nagy et al (2009)), but will highlight the most commonly used Cre lines ( the same thing; the difference being whether analysis is focussed upon the gonad of the parent, or analysis is focussed upon the whole body of the offspring (for review, see Hammond & Matin (2009)). One real advantage of targeting specifically in gametes rather than simply driving Cre ubiquitously is that the floxed target gene and the Cre transgene can segregate independently, and thus it is possible to obtain recombined mice that have not inherited the Cre transgene, an important control, especially where cellular toxicity arising from systemic Cre expression is a concern (Higashi et al 2009, Jae Huh et al 2010. Many Cre lines have been developed that target GCs, albeit unwittingly.…”

Section: Targeting the Testismentioning

confidence: 99%

Good planning and serendipity: exploiting the Cre/Lox system in the testis

Smith¹

2011

Reproduction

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Over the past 20 years, genetic manipulation has revolutionised our understanding of male reproductive development and function. The advent of transgenic mouse lines has permitted elegant dissection of previously intractable issues. The development of the Cre/Lox system, which has permitted spatial and temporal localisation of genetic manipulation, has expanded upon this, and now makes up one of the primary approaches underpinning our increasing understanding of testis development and function. The success of conditional gene targeting is largely reliant upon the choice of Cre recombinase expressing mouse line, which is required to specifically target the correct cell type at the correct time. Presupposition that Cre lines will behave as expected has been one of the main oversights in the design of Cre/Lox experiments, as in practice, many Cre lines are prone to ectopic expression (both temporal and spatial), transgene silencing or genetic background effects. Empirical validation of the spatiotemporal profile of Cre expression prior to undertaking conditional gene targeting studies is essential and can be achieved through a combination of molecular and immunohistochemical approaches, along with in vivo examination of reporter gene expression in targeted tissues. This paper details the key considerations associated with exploitation of the Cre/Lox system and highlights a variety of validated Cre lines that have utility for conditional gene targeting within the testis.

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“…24 To obtain mosaic recombination, we administered 0.025, 0.05, and 0.15 mg/g body wt tamoxifen by intraperitoneal injection for 1 day.…”

Section: Administration Of Tamoxifenmentioning

confidence: 99%

Severity and Frequency of Proximal Tubule Injury Determines Renal Prognosis

Takaori

Nakamura

Yamamoto

et al. 2015

JASN

Self Cite

211

167

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AKI increases the risk of developing CKD, but the mechanisms linking AKI to CKD remain unclear. Because proximal tubule injury is the mainstay of AKI, we postulated that proximal tubule injury triggers features of CKD. We generated a novel mouse model to induce proximal tubule-specific adjustable injury by inducing the expression of diphtheria toxin (DT) receptor with variable prevalence in proximal tubules. Administration of high-dose DT in mice expressing the DT receptor consistently caused severe proximal tubule-specific injury associated with interstitial fibrosis and reduction of erythropoietin production. Mild proximal tubule injury from a single injection of low-dose DT triggered reversible fibrosis, whereas repeated mild injuries caused sustained interstitial fibrosis, inflammation, glomerulosclerosis, and atubular glomeruli. DT-induced proximal tubule-specific injury also triggered distal tubule injury. Furthermore, injured tubular cells cocultured with fibroblasts stimulated induction of extracellular matrix and inflammatory genes. These results support the existence of proximal-distal tubule crosstalk and crosstalk between tubular cells and fibroblasts. Overall, our data provide evidence that proximal tubule injury triggers several features of CKD and that the severity and frequency of proximal tubule injury determines the progression to CKD.

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Direct Hematological Toxicity and Illegitimate Chromosomal Recombination Caused by the Systemic Activation of CreERT2

Cited by 128 publications

References 43 publications

Protein transduction from retroviral Gag precursors

Protein transduction from retroviral Gag precursors

Good planning and serendipity: exploiting the Cre/Lox system in the testis

Severity and Frequency of Proximal Tubule Injury Determines Renal Prognosis

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