Multiplexed mammalian gene control via modular splicing riboswitches
- 1Nanjing University of Chinese Medicine
Abstract
Precise, multiplexed gene regulation in mammalian cells remains constrained by a heavy reliance on heterologous protein regulators, whose associated immunogenicity and strict cargo-size limits impede clinical translation. Here, we introduce SpliceRACER (Splice-based Riboswitch Array for Combinatorial Expression Regulation), a multiplexed RNA switch platform that achieves scalable, orthogonal, and combinatorial control of gene expression in mammalian cells through alternative splicing, eliminating the need for exogenous protein co-expression. SpliceRACER independently orchestrates three orthogonal channels to regulate fluorescent reporter outputs, yielding 27 discrete combinatorial expression states, vastly exceeding the capacity of conventional binary gene switches. Furthermore, we engineered three generalizable strategies to effectively suppress the basal leakage inherent to synthetic riboswitches, thereby enabling tight OFF-state control of the potent pyroptotic effector GSDMD-NT for precise cell-death induction. This work offers broad utility in synthetic biology and therapeutic applications.
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BibTeX
@misc{jiang2026multiplexed,
title = {{Multiplexed mammalian gene control via modular splicing riboswitches}},
author = {J. Jiang and T. Ma},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.16.759771},
url = {https://www.biorxiv.org/content/10.64898/2026.10.16.759771v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Multiplexed mammalian gene control via modular splicing riboswitches
AU - J. Jiang
AU - T. Ma
PY - 2026
DA - 2026/10/16
PB - bioRxiv
DO - 10.64898/2026.10.16.759771
UR - https://www.biorxiv.org/content/10.64898/2026.10.16.759771v1
AB - Precise, multiplexed gene regulation in mammalian cells remains constrained by a heavy reliance on heterologous protein regulators, whose associated immunogenicity and strict cargo-size limits impede clinical translation. Here, we introduce SpliceRACER (Splice-based Riboswitch Array for Combinatorial Expression Regulation), a multiplexed RNA switch platform that achieves scalable, orthogonal, and combinatorial control of gene expression in mammalian cells through alternative splicing, eliminating the need for exogenous protein co-expression. SpliceRACER independently orchestrates three orthogonal channels to regulate fluorescent reporter outputs, yielding 27 discrete combinatorial expression states, vastly exceeding the capacity of conventional binary gene switches. Furthermore, we engineered three generalizable strategies to effectively suppress the basal leakage inherent to synthetic riboswitches, thereby enabling tight OFF-state control of the potent pyroptotic effector GSDMD-NT for precise cell-death induction. This work offers broad utility in synthetic biology and therapeutic applications.
ER -