A neuron-optimized CRISPR/dCas9 activation system for robust and specific gene regulation
| Autor: | Jennifer J. Tuscher, Jasmin S. Revanna, Jeremy J. Day, Svitlana V. Bach, Katherine E. Savell, Nicholas A. Goska, Corey G. Duke, Julia N. Burke, Morgan E. Zipperly, Faraz A. Sultan, Lara Ianov, Derek M. Williams |
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| Rok vydání: | 2018 |
| Předmět: |
Male
Transcription Genetic Cell Adhesion Molecules Neuronal Transgene Primary Cell Culture Nerve Tissue Proteins Computational biology Biology Novel Tools and Methods Rats Sprague-Dawley Random Allocation 03 medical and health sciences Bdnf 0302 clinical medicine Genome editing Neurotrophic factors Cell Line Tumor Gene expression Animals CRISPR Epigenetics Gene Methods/New Tools 030304 developmental biology Neurons Regulation of gene expression Extracellular Matrix Proteins 0303 health sciences epigenetics General Neuroscience Brain-Derived Neurotrophic Factor Serine Endopeptidases Brain Promoter General Medicine 3. Good health Reelin Protein Gene Expression Regulation Genetic Techniques 7.2 gene expression CRISPR-Cas Systems transcription Transcriptome 030217 neurology & neurosurgery |
| Zdroj: | eNeuro |
| Popis: | Recent developments in CRISPR-based gene editing have provided new avenues to interrogate gene function. However, application of these tools in the central nervous system has been delayed due to difficulties in transgene expression in post-mitotic neurons. Here, we present a highly efficient, neuron-optimized dual lentiviral CRISPR-based transcriptional activation (CRISPRa) system to drive gene expression in primary neuronal cultures and the adult brain of rodent model systems. We demonstrate robust, modular, and tunable induction of endogenous target genes as well as multiplexed gene regulation necessary for investigation of complex transcriptional programs. CRISPRa targeting unique promoters in the complex multi-transcript gene Brain-derived neurotrophic factor (Bdnf) revealed both transcript- and genome-level selectivity of this approach, in addition to highlighting downstream transcriptional and physiological consequences of Bdnf regulation. Finally, we illustrate that CRISPRa is highly efficient in vivo, resulting in increased protein levels of a target gene in diverse brain structures. Taken together, these results demonstrate that CRISPRa is an efficient and selective method to study gene expression programs in brain health and disease. |
| Databáze: | OpenAIRE |
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