Preferential delivery of the Sleeping Beauty transposon system to livers of mice by hydrodynamic injection

Bell, Jason; Podetz-Pedersen, Kelly M.; Aronovich, Elena L.; Belur, Lalitha R.; McIvor, R. Scott; Hackett, Perry B.

doi:10.1038/nprot.2007.471

Cited by 131 publications

(139 citation statements)

References 73 publications

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“…2b). To further verify activation of the hepatic IRE1a-XBP1 pathway by chronic fasting, we introduced an Xbp1-splicing-directed luciferase reporter 23 into the livers of mice through hydrodynamic injection 24 . Consistently, in comparison with a 6-h short-term fast or ad libitum feeding, prolonged fasting for 18 h induced a marked elevation of liver luciferase reporter activity in flox/flox mice, but not in LKO mice (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The Luc reporter for the mouse Ppara promoter spanning the ARTICLE region from À 2,000 to þ 1 was constructed in pGL3 (Promega) utilizing a PCRbased cloning strategy. Reporter DNA constructs were introduced into livers of mice through hydrodynamic injection as described 24 . Briefly, a DNA solution (12.5 mg ml À 1 in sterile PBS) was injected at 10% of volume/body weight through the tail vein within 10 s, and mice were then allowed to recover for 24 h. For in vivo imaging, anesthetized mice were injected i.p.…”

Section: Methodsmentioning

confidence: 99%

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Hepatic IRE1α regulates fasting-induced metabolic adaptive programs through the XBP1s–PPARα axis signalling

et al. 2014

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Although the mammalian IRE1a-XBP1 branch of the cellular unfolded protein response has been implicated in glucose and lipid metabolism, the exact metabolic role of IRE1a signalling in vivo remains poorly understood. Here we show that hepatic IRE1a functions as a nutrient sensor that regulates the metabolic adaptation to fasting. We find that prolonged deprivation of food or consumption of a ketogenic diet activates the IRE1a-XBP1 pathway in mouse livers. Hepatocyte-specific abrogation of Ire1a results in impairment of fatty acid b-oxidation and ketogenesis in the liver under chronic fasting or ketogenic conditions, leading to hepatosteatosis; liver-specific restoration of XBP1s reverses the defects in IRE1a null mice. XBP1s directly binds to and activates the promoter of PPARa, the master regulator of starvation responses. Hence, our results demonstrate that hepatic IRE1a promotes the adaptive shift of fuel utilization during starvation by stimulating mitochondrial b-oxidation and ketogenesis through the XBP1s-PPARa axis.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Hepatic IRE1α regulates fasting-induced metabolic adaptive programs through the XBP1s–PPARα axis signalling

et al. 2014

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“…The reason for modified-cell attrition is not clear, although it may be that without selection, cells with a low level of transgene expression have a growth advantage over cells with higher levels of expression. Given that coselection was able to enrich and maintain modified cells, it seems unlikely that nuclease off-target activity or toxicity (20,32) is the cause for attrition. Instead, selection for antibiotic resistance is likely biased toward high levels of ectopic gene expression, thereby enriching for cells in which TALEN expression was also high, and cleavage more likely to have occurred.…”

Section: Discussionmentioning

confidence: 99%

“…Potentially, transposition mobilizes the same factors that are involved with repair after double-stranded DNA cleavage by nucleases. Because transposition occurs in only a low percentage of transfected cells, as well as cells in animal tissues (32)(33)(34), coselection might enrich for a subpopulation of cells that were amenable to NHEJ.…”

Section: Discussionmentioning

confidence: 99%

Efficient TALEN-mediated gene knockout in livestock

Carlson

Tan

Lillico

et al. 2012

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

531

404

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Transcription activator-like effector nucleases (TALENs) are programmable nucleases that join FokI endonuclease with the modular DNA-binding domain of TALEs. Although zinc-finger nucleases enable a variety of genome modifications, their application to genetic engineering of livestock has been slowed by technical limitations of embryo-injection, culture of primary cells, and difficulty in producing reliable reagents with a limited budget. In contrast, we found that TALENs could easily be manufactured and that over half (23/36, 64%) demonstrate high activity in primary cells. Cytoplasmic injections of TALEN mRNAs into livestock zygotes were capable of inducing gene KO in up to 75% of embryos analyzed, a portion of which harbored biallelic modification. We also developed a simple transposon coselection strategy for TALEN-mediated gene modification in primary fibroblasts that enabled both enrichment for modified cells and efficient isolation of modified colonies. Coselection after treatment with a single TALEN-pair enabled isolation of colonies with mono-and biallelic modification in up to 54% and 17% of colonies, respectively. Coselection after treatment with two TALEN-pairs directed against the same chromosome enabled the isolation of colonies harboring large chromosomal deletions and inversions (10% and 4% of colonies, respectively). TALEN-modified Ossabaw swine fetal fibroblasts were effective nuclear donors for cloning, resulting in the creation of miniature swine containing mono-and biallelic mutations of the LDL receptor gene as models of familial hypercholesterolemia. TALENs thus appear to represent a highly facile platform for the modification of livestock genomes for both biomedical and agricultural applications.Tal-effector nuclease | biotechnology | gene-editing

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“…Hydrodynamic tail-vein injections and Sleeping Beauty transposon-induced liver tumorigenesis were performed as described previously (25,35). Six-to 10-wk-old mice were injected with a plasmid mixture diluted in lactated Ringer's solution.…”

Section: Methodsmentioning

confidence: 99%

Cyclin-dependent kinase 1 (Cdk1) is essential for cell division and suppression of DNA re-replication but not for liver regeneration

Diril

Ratnacaram

Padmakumar

et al. 2012

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

330

324

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Cyclin-dependent kinase 1 (Cdk1) is an archetypical kinase and a central regulator that drives cells through G2 phase and mitosis. Knockouts of Cdk2, Cdk3, Cdk4, or Cdk6 have resulted in viable mice, but the in vivo functions of Cdk1 have not been fully explored in mammals. Here we have generated a conditional-knockout mouse model to study the functions of Cdk1 in vivo. Ablation of Cdk1 leads to arrest of embryonic development around the blastocyst stage. Interestingly, liver-specific deletion of Cdk1 is well tolerated, and liver regeneration after partial hepatectomy is not impaired, indicating that regeneration can be driven by cell growth without cell division. The loss of Cdk1 does not affect S phase progression but results in DNA re-replication because of an increase in Cdk2/cyclin A2 activity. Unlike other Cdks, loss of Cdk1 in the liver confers complete resistance against tumorigenesis induced by activated Ras and silencing of p53.cancer | knockout mice | cell cycle regulation | polyploidy

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Preferential delivery of the Sleeping Beauty transposon system to livers of mice by hydrodynamic injection

Cited by 131 publications

References 73 publications

Hepatic IRE1α regulates fasting-induced metabolic adaptive programs through the XBP1s–PPARα axis signalling

Hepatic IRE1α regulates fasting-induced metabolic adaptive programs through the XBP1s–PPARα axis signalling

Efficient TALEN-mediated gene knockout in livestock

Cyclin-dependent kinase 1 (Cdk1) is essential for cell division and suppression of DNA re-replication but not for liver regeneration

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