Cargando…

A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation

CRISPR-based technology has provided new avenues to interrogate gene function, but difficulties in transgene expression in post-mitotic neurons has delayed incorporation of these tools in the central nervous system (CNS). Here, we demonstrate a highly efficient, neuron-optimized dual lentiviral CRIS...

Descripción completa

Detalles Bibliográficos
Autores principales: Savell, Katherine E., Bach, Svitlana V., Zipperly, Morgan E., Revanna, Jasmin S., Goska, Nicholas A., Tuscher, Jennifer J., Duke, Corey G., Sultan, Faraz A., Burke, Julia N., Williams, Derek, Ianov, Lara, Day, Jeremy J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Society for Neuroscience 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6412672/
https://www.ncbi.nlm.nih.gov/pubmed/30863790
http://dx.doi.org/10.1523/ENEURO.0495-18.2019
_version_ 1783402660176592896
author Savell, Katherine E.
Bach, Svitlana V.
Zipperly, Morgan E.
Revanna, Jasmin S.
Goska, Nicholas A.
Tuscher, Jennifer J.
Duke, Corey G.
Sultan, Faraz A.
Burke, Julia N.
Williams, Derek
Ianov, Lara
Day, Jeremy J.
author_facet Savell, Katherine E.
Bach, Svitlana V.
Zipperly, Morgan E.
Revanna, Jasmin S.
Goska, Nicholas A.
Tuscher, Jennifer J.
Duke, Corey G.
Sultan, Faraz A.
Burke, Julia N.
Williams, Derek
Ianov, Lara
Day, Jeremy J.
author_sort Savell, Katherine E.
collection PubMed
description CRISPR-based technology has provided new avenues to interrogate gene function, but difficulties in transgene expression in post-mitotic neurons has delayed incorporation of these tools in the central nervous system (CNS). Here, we demonstrate a highly efficient, neuron-optimized dual lentiviral CRISPR-based transcriptional activation (CRISPRa) system capable of robust, modular, and tunable gene induction and multiplexed gene regulation across several primary rodent neuron culture systems. 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.
format Online
Article
Text
id pubmed-6412672
institution National Center for Biotechnology Information
language English
publishDate 2019
publisher Society for Neuroscience
record_format MEDLINE/PubMed
spelling pubmed-64126722019-03-12 A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation Savell, Katherine E. Bach, Svitlana V. Zipperly, Morgan E. Revanna, Jasmin S. Goska, Nicholas A. Tuscher, Jennifer J. Duke, Corey G. Sultan, Faraz A. Burke, Julia N. Williams, Derek Ianov, Lara Day, Jeremy J. eNeuro Methods/New Tools CRISPR-based technology has provided new avenues to interrogate gene function, but difficulties in transgene expression in post-mitotic neurons has delayed incorporation of these tools in the central nervous system (CNS). Here, we demonstrate a highly efficient, neuron-optimized dual lentiviral CRISPR-based transcriptional activation (CRISPRa) system capable of robust, modular, and tunable gene induction and multiplexed gene regulation across several primary rodent neuron culture systems. 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. Society for Neuroscience 2019-03-07 /pmc/articles/PMC6412672/ /pubmed/30863790 http://dx.doi.org/10.1523/ENEURO.0495-18.2019 Text en Copyright © 2019 Savell et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.
spellingShingle Methods/New Tools
Savell, Katherine E.
Bach, Svitlana V.
Zipperly, Morgan E.
Revanna, Jasmin S.
Goska, Nicholas A.
Tuscher, Jennifer J.
Duke, Corey G.
Sultan, Faraz A.
Burke, Julia N.
Williams, Derek
Ianov, Lara
Day, Jeremy J.
A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title_full A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title_fullStr A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title_full_unstemmed A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title_short A Neuron-Optimized CRISPR/dCas9 Activation System for Robust and Specific Gene Regulation
title_sort neuron-optimized crispr/dcas9 activation system for robust and specific gene regulation
topic Methods/New Tools
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6412672/
https://www.ncbi.nlm.nih.gov/pubmed/30863790
http://dx.doi.org/10.1523/ENEURO.0495-18.2019
work_keys_str_mv AT savellkatherinee aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT bachsvitlanav aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT zipperlymorgane aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT revannajasmins aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT goskanicholasa aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT tuscherjenniferj aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT dukecoreyg aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT sultanfaraza aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT burkejulian aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT williamsderek aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT ianovlara aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT dayjeremyj aneuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT savellkatherinee neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT bachsvitlanav neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT zipperlymorgane neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT revannajasmins neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT goskanicholasa neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT tuscherjenniferj neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT dukecoreyg neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT sultanfaraza neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT burkejulian neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT williamsderek neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT ianovlara neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation
AT dayjeremyj neuronoptimizedcrisprdcas9activationsystemforrobustandspecificgeneregulation