title: Development of a synthetic gene network to modulate gene expression by mechanical forces.
abstract: The majority of (mammalian) cells in our body are sensitive to mechanical forces, but little work has been done to develop assays to monitor mechanosensor activity. Furthermore, it is currently impossible to use mechanosensor activity to drive gene expression. To address these needs, we developed the first mammalian mechanosensitive synthetic gene network to monitor endothelial cell shear stress levels and directly modulate expression of an atheroprotective transcription factor by shear stress. The technique is highly modular, easily scalable and allows graded control of gene expression by mechanical stimuli in hard-to-transfect mammalian cells. We call this new approach mechanosyngenetics. To insert the gene network into a high proportion of cells, a hybrid transfection procedure was developed that involves electroporation, plasmids replication in mammalian cells, mammalian antibiotic selection, a second electroporation and gene network activation. This procedure takes 1 week and yielded over 60% of cells with a functional gene network. To test gene network functionality, we developed a flow setup that exposes cells to linearly increasing shear stress along the length of the flow channel floor. Activation of the gene network varied logarithmically as a function of shear stress magnitude.
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Zoltán
authors.initials: Z
authors.lastname: Kis
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Tania
authors.initials: T
authors.lastname: Rodin
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.|Government College University, GC University, Katchery Road, Lahore, 54000, Pakistan.
authors.forename: Asma
authors.initials: A
authors.lastname: Zafar
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.|Department of Mechanical Engineering, National University of Singapore, 9 Engineering Drive 1, 117575, Singapore.|Department of Biomedical Engineering, National University of Singapore, 4 Engineering Drive 3, 117583, Singapore.
authors.forename: Zhangxing
authors.initials: Z
authors.lastname: Lai
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.|Institute of Child Health, University College London, 30 Guilford Street, London, WC1N 1EH, United Kingdom.
authors.forename: Grace
authors.initials: G
authors.lastname: Freke
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.|Department of Engineering, University of Cambridge, Trumpington Street, Cambridge, CB2 1PZ, United Kingdom.
authors.forename: Oliver
authors.initials: O
authors.lastname: Fleck
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Armando
authors.initials: A
authors.lastname: Del Rio Hernandez
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Leila
authors.initials: L
authors.lastname: Towhidi
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Ryan M
authors.initials: RM
authors.lastname: Pedrigi
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.|Institute for Integrated Cell-Material Sciences, Kyoto University, Yoshida Ushinomiya-cho, Sakyo-ku, Kyoto, 606-8501, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Homma
authors.affiliation: Department of Bioengineering, Imperial College London, Exhibition Road, London, SW7 2AZ, United Kingdom.
authors.forename: Rob
authors.initials: R
authors.lastname: Krams
mesh_terms: D002470:Cell Survival; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism; D015870:Gene Expression; D005786:Gene Expression Regulation; D053263:Gene Regulatory Networks; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000235:genetics; D040542:Mechanotransduction, Cellular; D013314:Stress, Mechanical; D015533:Transcriptional Activation; D014162:Transfection
chemical_list: C096161:KLF2 protein, human; D051741:Kruppel-Like Transcription Factors