PT  - JOURNAL ARTICLE
AU  - Jonathan P. Ling
AU  - Alexei M. Bygrave
AU  - Clayton P. Santiago
AU  - Rogger P. Carmen-Orozco
AU  - Vickie Trinh
AU  - Minzhong Yu
AU  - Yini Li
AU  - Jeong Han
AU  - Kamil Taneja
AU  - Ying Liu
AU  - Rochinelle Dongmo
AU  - Travis A. Babola
AU  - Patrick Parker
AU  - Lizhi Jiang
AU  - Patrick J. Leavey
AU  - Jennifer J. Smith
AU  - Rachel Vistein
AU  - Megan Y. Gimmen
AU  - Benjamin Dubner
AU  - Eric Helmenstine
AU  - Patric Teodorescu
AU  - Theodore Karantanos
AU  - Gabriel Ghiaur
AU  - Patrick O. Kanold
AU  - Dwight Bergles
AU  - Ben Langmead
AU  - Shuying Sun
AU  - Kristina J. Nielsen
AU  - Neal Peachey
AU  - Mandeep S. Singh
AU  - W. Brian Dalton
AU  - Fatemeh Rajaii
AU  - Richard L. Huganir
AU  - Seth Blackshaw
TI  - Cell-specific regulation of gene expression using splicing-dependent frameshifting
AID  - 10.1101/2022.03.02.481623
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2022.03.02.481623
4099  - http://biorxiv.org/content/early/2022/03/02/2022.03.02.481623.short
4100  - http://biorxiv.org/content/early/2022/03/02/2022.03.02.481623.full
AB  - Precise and reliable cell-specific gene delivery remains technically challenging. Here we report a splicing-based approach for controlling gene expression whereby separate translational reading frames are coupled to the inclusion or exclusion of cell-specific alternative exons. Candidate exons are identified by analyzing thousands of publicly available RNA sequencing datasets and filtering by cell specificity, sequence conservation, and local intron length. This method, which we denote splicing-linked expression design (SLED), can be combined in a Boolean manner with existing techniques such as minipromoters and viral capsids. SLED vectors can leverage the strong expression of constitutive promoters, without sacrificing precision, by decoupling the tradeoff between promoter strength and selectivity. We generated SLED vectors to selectively target all neurons, photoreceptors, or excitatory neurons, and demonstrated that specificity was retained in vivo when delivered using AAVs. We further demonstrated the utility of SLED by creating what would otherwise be unobtainable research tools, specifically a GluA2 flip/flop reporter and a dual excitatory/inhibitory neuronal calcium indicator. Finally, we show the translational potential of SLED by rescuing photoreceptor degeneration in Prph2rds/rds mice and by developing an oncolytic vector that can selectively induce apoptosis in SF3B1 mutant cancer cells. The flexibility of SLED technology enables new avenues for basic and translational research.Competing Interest StatementSB receives research support from Genentech, and is a co-founder and shareholder in CDI Labs, LLC. MSS is/was a paid advisor to Revision Therapeutics, Johnson & Johnson, Third Rock Ventures, Bayer Healthcare, Novartis Pharmaceuticals, W. L. Gore & Associates, Deerfield, Trinity Partners, Kala Pharmaceuticals, and Acucela. SB and JL have filed a patent application covering SLED technology (Provisional US Patent PCT/US20/56156).