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A brain-sparing diphtheria toxin for chemical genetic ablation of peripheral cell lineages
Mafalda M A Pereira1, Inês Mahú1, Elsa Seixas1
1Obesity Laboratory, Instituto Gulbenkian de Ciência, Oeiras 2780-156, Portugal.
Abstract:
Conditional expression of diphtheria toxin receptor (DTR) is widely used for tissue-specific ablation of cells. However, diphtheria toxin (DT) crosses the blood-brain barrier, which limits its utility for ablating peripheral cells using Cre drivers that are also expressed in the central nervous system (CNS). Here we report the development of a brain-sparing DT, termed BRAINSPAReDT, for tissue-specific genetic ablation of cells outside the CNS. We prevent blood-brain barrier passage of DT through PEGylation, which polarizes the molecule and increases its size. We validate BRAINSPAReDT with regional genetic sympathectomy: BRAINSPAReDT ablates peripheral but not central catecholaminergic neurons, thus avoiding the Parkinson-like phenotype associated with full dopaminergic depletion. Regional sympathectomy compromises adipose tissue thermogenesis, and renders mice susceptible to obesity. We provide a proof of principle that BRAINSPAReDT can be used for Cre/DTR tissue-specific ablation outside the brain using CNS drivers, while consolidating the link between adiposity and the sympathetic nervous system.
Insights
Researchers developed BRAINSPAReDT, a modified diphtheria toxin (DT), to selectively ablate peripheral cells without affecting the central nervous system. This innovation enables precise genetic targeting outside the brain, crucial for studying conditions like obesity.
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- Diphtheria toxin receptor (DTR)-mediated cell ablation is vital for tissue-specific studies.
- Standard diphtheria toxin (DT) crosses the blood-brain barrier, limiting its use with CNS-expressed Cre drivers for peripheral ablation.
- This limitation hinders research on peripheral systems when central nervous system (CNS) drivers are involved.
Purpose of the Study:
- To develop a brain-sparing diphtheria toxin (DT) for selective ablation of peripheral cells.
- To enable tissue-specific genetic ablation outside the CNS using Cre drivers also present in the CNS.
- To investigate the role of the sympathetic nervous system in adiposity and thermogenesis.
Main Methods:
- Development of BRAINSPAReDT, a PEGylated diphtheria toxin (DT), to prevent blood-brain barrier penetration.
- Validation using regional genetic sympathectomy in mice.
- Assessment of catecholaminergic neuron ablation in peripheral and central nervous systems.
- Evaluation of effects on adipose tissue thermogenesis and susceptibility to obesity.
Main Results:
- BRAINSPAReDT successfully ablated peripheral catecholaminergic neurons while sparing central neurons.
- This selective ablation avoided the Parkinson-like phenotype associated with complete dopaminergic depletion.
- Regional sympathectomy led to compromised adipose tissue thermogenesis and increased obesity susceptibility.
- Established a link between adiposity and the sympathetic nervous system.
Conclusions:
- BRAINSPAReDT is an effective tool for Cre/DTR-mediated tissue-specific ablation outside the brain, even with CNS drivers.
- This method overcomes the limitations of standard DT in studies involving both CNS and peripheral tissues.
- The findings reinforce the critical role of the sympathetic nervous system in regulating body weight and metabolism.