title: Transcription activator-like effector-mediated regulation of gene expression based on the inducible packaging and delivery via designed extracellular vesicles.
abstract: Transcription activator-like effector (TALE) proteins present a powerful tool for genome editing and engineering, enabling introduction of site-specific mutations, gene knockouts or regulation of the transcription levels of selected genes. TALE nucleases or TALE-based transcription regulators are introduced into mammalian cells mainly via delivery of the coding genes. Here we report an extracellular vesicle-mediated delivery of TALE transcription regulators and their ability to upregulate the reporter gene in target cells. Designed transcriptional activator TALE-VP16 fused to the appropriate dimerization domain was enriched as a cargo protein within extracellular vesicles produced by mammalian HEK293 cells stimulated by Ca-ionophore and using blue light- or rapamycin-inducible dimerization systems. Blue light illumination or rapamycin increased the amount of the TALE-VP16 activator in extracellular vesicles and their addition to the target cells resulted in an increased expression of the reporter gene upon addition of extracellular vesicles to the target cells. This technology therefore represents an efficient delivery for the TALE-based transcriptional regulators.
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry, Ljubljana, Hajdrihova 19, 1000 Ljubljana, Slovenia; Graduate School of Biomedicine, University of Ljubljana, 1000 Ljubljana, Slovenia.
authors.forename: Duško
authors.initials: D
authors.lastname: Lainšček
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry, Ljubljana, Hajdrihova 19, 1000 Ljubljana, Slovenia; Graduate School of Biomedicine, University of Ljubljana, 1000 Ljubljana, Slovenia.
authors.forename: Tina
authors.initials: T
authors.lastname: Lebar
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry, Ljubljana, Hajdrihova 19, 1000 Ljubljana, Slovenia; EN-FIST, Centre of Excellence, Trg osvobodilne fronte 13, 1000 Ljubljana, Slovenia. Electronic address: roman.jerala@ki.si.
authors.forename: Roman
authors.initials: R
authors.lastname: Jerala
mesh_terms: D019281:Dimerization; D000067128:Extracellular Vesicles / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D054876:Prenylation; D000071336:Transcription Activator-Like Effectors / Q000502:physiology*; D015854:Up-Regulation
chemical_list: D000071336:Transcription Activator-Like Effectors
keywords: Dimerization; Extracellular vesicles; TALE; Transcriptional regulation