Josh Tycko

Systematic discovery of protein functions in human cells to understand gene regulation and enable genetic medicine

Tunable, self-contained gene dosage control via proteolytic cleavage of CRISPR-Cas systems


Journal article


Noa Katz, Connie An, Yu-Ju Lee, Josh Tycko, Meng Zhang, Jeewoo Kang, Lacramiora Bintu, Mike C Bassik, Wei-Hsiang Huang, Xiaojing J. Gao
bioRxiv, 2024

Semantic Scholar DOI PubMedCentral PubMed
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Cite

APA   Click to copy
Katz, N., An, C., Lee, Y.-J., Tycko, J., Zhang, M., Kang, J., … Gao, X. J. (2024). Tunable, self-contained gene dosage control via proteolytic cleavage of CRISPR-Cas systems. BioRxiv.


Chicago/Turabian   Click to copy
Katz, Noa, Connie An, Yu-Ju Lee, Josh Tycko, Meng Zhang, Jeewoo Kang, Lacramiora Bintu, Mike C Bassik, Wei-Hsiang Huang, and Xiaojing J. Gao. “Tunable, Self-Contained Gene Dosage Control via Proteolytic Cleavage of CRISPR-Cas Systems.” bioRxiv (2024).


MLA   Click to copy
Katz, Noa, et al. “Tunable, Self-Contained Gene Dosage Control via Proteolytic Cleavage of CRISPR-Cas Systems.” BioRxiv, 2024.


BibTeX   Click to copy

@article{noa2024a,
  title = {Tunable, self-contained gene dosage control via proteolytic cleavage of CRISPR-Cas systems},
  year = {2024},
  journal = {bioRxiv},
  author = {Katz, Noa and An, Connie and Lee, Yu-Ju and Tycko, Josh and Zhang, Meng and Kang, Jeewoo and Bintu, Lacramiora and Bassik, Mike C and Huang, Wei-Hsiang and Gao, Xiaojing J.}
}

Abstract

Gene therapy holds great therapeutic potential. Yet, controlling cargo expression in single cells is limited due to the variability of delivery methods. We implement an incoherent feedforward loop based on proteolytic cleavage of CRISPR-Cas activation or inhibition systems to reduce gene expression variability against the variability of vector delivery. We demonstrate dosage control for activation and inhibition, post-delivery tuning, and RNA-based delivery, for a genome-integrated marker. We then target the RAI1 gene, the haploinsufficiency and triplosensitivity of which cause two autism-related syndromes, Smith-Magenis-Syndrome (SMS) and Potocki-Lupski-Syndrome, respectively. We demonstrate dosage control for RAI1 activation in HEK293s, Neuro-2As, and mouse cortical neurons via AAVs and lentiviruses. Finally, we activate the intact RAI1 copy in SMS patient-derived cells to an estimated two-copy healthy range, avoiding the harmful three-copy regime. Our circuit paves the way for viable therapy in dosage-sensitive disorders, creating precise and tunable gene regulation systems for basic and translational research.




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