title: A yeast-based rapid prototype platform for gene control elements in mammalian cells.
abstract: Programming genetic circuits in mammalian cells requires flexible, tunable, and user-tailored gene-control systems. However, most existing control systems are either mechanistically specific for microbial organisms or must be laboriously re-engineered to function in mammalian cells. Here, we demonstrate a ribozyme-based device platform that can be directly transported from yeast to mammalian cells in a "plug-and-play" manner. Ribozyme switches previously prototyped in yeast are shown to regulate gene expression in a predictable, ligand-responsive manner in human HEK 293, HeLa, and U2OS cell lines without any change to device sequence nor further optimization. The ribozyme-based devices, which exhibit activation ratios comparable to the best RNA-based regulatory devices demonstrated in mammalian cells to-date, retain their prescribed functions (ON or OFF switch), tunability of regulatory stringency, and responsiveness to different small-molecule inputs in mammalian hosts. Furthermore, we observe strong correlations of device performance between yeast and all mammalian cell lines tested (R(2)  = 0.63-0.97). Our unique device architecture can therefore act as a rapid prototyping platform (RPP) based on a yeast chassis, providing a well-developed and genetically tractable system that supports rapid and high-throughput screens for generating gene-controllers with a broad range of functions in mammalian cells. This platform will accelerate development of mammalian gene-controllers for diverse applications, including cell-based therapeutics and cell-fate reprogramming.
authors.affiliation: Department of Bioengineering, Stanford University, 473 Via Ortega, MC 4201, Stanford, CA 94305, USA.
authors.forename: Kathy Y
authors.initials: KY
authors.lastname: Wei
authors.forename: Yvonne Y
authors.initials: YY
authors.lastname: Chen
authors.forename: Christina D
authors.initials: CD
authors.lastname: Smolke
mesh_terms: D002460:Cell Line; D053263:Gene Regulatory Networks; D005800:Genes, Fungal; D006801:Humans; D016337:RNA, Catalytic / Q000378:metabolism; D015003:Yeasts / Q000235:genetics*
chemical_list: D016337:RNA, Catalytic