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Conditional gene targeting in mouse pancreatic ß-Cells: analysis of ectopic Cre transgene expression in the brain
Barton Wicksteed1, Marcela Brissova, Wenbo Yan
1Section of Adult and Pediatric Endocrinology, Diabetes and Metabolism, Department of Medicine, University of Chicago, Chicago, llinois, USA. wicksteed@uchicago.edu
Objective:
Conditional gene targeting has been extensively used for in vivo analysis of gene function in β-cell biology. The objective of this study was to examine whether mouse transgenic Cre lines, used to mediate β-cell- or pancreas-specific recombination, also drive Cre expression in the brain.
Research Design And Methods:
Transgenic Cre lines driven by Ins1, Ins2, and Pdx1 promoters were bred to R26R reporter strains. Cre activity was assessed by β-galactosidase or yellow fluorescent protein expression in the pancreas and the brain. Endogenous Pdx1 gene expression was monitored using Pdx1(tm1Cvw) lacZ knock-in mice. Cre expression in β-cells and co-localization of Cre activity with orexin-expressing and leptin-responsive neurons within the brain was assessed by immunohistochemistry.
Results:
All transgenic Cre lines examined that used the Ins2 promoter to drive Cre expression showed widespread Cre activity in the brain, whereas Cre lines that used Pdx1 promoter fragments showed more restricted Cre activity primarily within the hypothalamus. Immunohistochemical analysis of the hypothalamus from Tg(Pdx1-cre)(89.1Dam) mice revealed Cre activity in neurons expressing orexin and in neurons activated by leptin. Tg(Ins1-Cre/ERT)(1Lphi) mice were the only line that lacked Cre activity in the brain.
Conclusions:
Cre-mediated gene manipulation using transgenic lines that express Cre under the control of the Ins2 and Pdx1 promoters are likely to alter gene expression in nutrient-sensing neurons. Therefore, data arising from the use of these transgenic Cre lines must be interpreted carefully to assess whether the resultant phenotype is solely attributable to alterations in the islet β-cells.
Insights
Certain mouse models used for studying pancreatic beta cells also show Cre expression in the brain. Researchers found that Ins2 and Pdx1 promoter-driven Cre lines affect brain neurons, necessitating careful interpretation of experimental results.
Area of Science:
- Neuroscience
- Endocrinology
- Genetics
Background:
- Conditional gene targeting is crucial for in vivo gene function analysis in beta-cell biology.
- Transgenic Cre lines are widely used to achieve pancreas-specific gene recombination.
Purpose of the Study:
- To investigate if mouse transgenic Cre lines, intended for beta-cell or pancreas-specific recombination, also exhibit Cre expression in the brain.
- To assess the specificity of Cre activity in different transgenic models.
Main Methods:
- Transgenic Cre lines utilizing Ins1, Ins2, and Pdx1 promoters were crossed with R26R reporter strains.
- Cre activity was evaluated via beta-galactosidase or yellow fluorescent protein expression in the pancreas and brain.
- Immunohistochemistry was employed to examine Cre expression in beta-cells and its co-localization with specific neuronal populations in the brain.
Main Results:
- Cre lines driven by the Ins2 promoter displayed widespread brain activity.
- Cre lines using Pdx1 promoter fragments showed restricted activity, mainly in the hypothalamus.
- Cre activity was detected in orexin-expressing and leptin-responsive neurons in the hypothalamus.
- The Tg(Ins1-Cre/ERT)(1Lphi) line was the only one lacking brain Cre activity.
Conclusions:
- Cre expression under the control of Ins2 and Pdx1 promoters can impact gene expression in brain nutrient-sensing neurons.
- Results from studies using these specific transgenic Cre lines require careful interpretation to distinguish between beta-cell specific effects and potential off-target effects in the brain.
