Scalable control of stem cell fate by riboswitch-regulated RNA viral vector without genomic integration

Narae Kim; Yohei Yokobayashi

doi:10.1016/j.ymthe.2025.01.005

Scalable control of stem cell fate by riboswitch-regulated RNA viral vector without genomic integration

Mol Ther. 2025 Jan 9:S1525-0016(25)00009-7. doi: 10.1016/j.ymthe.2025.01.005. Online ahead of print.

Authors

Narae Kim¹, Yohei Yokobayashi²

Affiliations

¹ Nucleic Acid Chemistry and Engineering Unit, Okinawa Institute of Science and Technology Graduate University, Onna, Okinawa 904 0495, Japan.
² Nucleic Acid Chemistry and Engineering Unit, Okinawa Institute of Science and Technology Graduate University, Onna, Okinawa 904 0495, Japan. Electronic address: [email protected].

PMID: 39797398
DOI: 10.1016/j.ymthe.2025.01.005

Abstract

Transgene expression in stem cells is a powerful means of regulating cellular properties and differentiation into various cell types. However, existing vectors for transgene expression in stem cells suffer from limitations such as the need for genomic integration, the transient nature of gene expression, and the inability to temporally regulate transgene expression, which hinder biomedical and clinical applications. Here we report a new class of RNA virus-based vectors for scalable and integration-free transgene expression in mouse embryonic stem cells (mESCs). The vector is equipped with a small molecule-regulated riboswitch and a drug selection marker that allow temporal regulation of transgene expression and stable maintenance of the vector in proliferating stem cells. We demonstrated the utility of the vector by maintaining the pluripotency of mESCs in a differentiation induction medium by expressing Nanog and inducing myogenic differentiation by triggering Myod1 expression, without altering the mESC genome.