Engineered transcription factor-binding arrays for DNA-based gene expression control in mammalian cells
File(s)
Published version
Author(s)
Zouein, Annalise
Lende-Dorn, Brittany
Galloway, Kate E
Ellis, Tom
Ceroni, Francesca
Type
Journal Article
Abstract
Tools that manipulate gene expression in mammalian cells without any additional expression are critical for cell engineering applications. Here, we demonstrate the use of arrays of transcription factor (TF) recognition elements (REs) as DNA tools for controlling gene expression. We first demonstrate that TetR-based RE arrays can alter synthetic gene circuit performance. We then open the approach to any TF with a known binding site by developing a new technique called Cloning Troublesome Repeats in Loops (CTRL), which can assemble plasmids with up to 256 RE repeats. Transfection of custom RE array plasmids assembled by CTRL into mammalian cells modifies host cell gene regulation by sequestration of TFs of interest and can sequester both synthetic and native TFs, offering applications in the control of gene circuits and for directing cell fate. This work advances our ability to assemble repetitive DNA arrays and shows how TF-binding RE arrays expand possibilities in mammalian cell engineering.
Date Issued
2025-08-01
Date Acceptance
2025-05-08
Citation
Trends in Biotechnology, 2025, 43 (8), pp.2029-2048
ISSN
0167-7799
Publisher
Elsevier BV
Start Page
2029
End Page
2048
Journal / Book Title
Trends in Biotechnology
Volume
43
Issue
8
Copyright Statement
© 2025 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
License URL
Identifier
10.1016/j.tibtech.2025.05.006
Publication Status
Published
Date Publish Online
2025-06-09