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Genome-wide prediction of matrix attachment regions that increase gene expression in mammalian cells

Abstract

Gene transfer in eukaryotic cells and organisms suffers from epigenetic effects that result in low or unstable transgene expression and high clonal variability. Use of epigenetic regulators such as matrix attachment regions (MARs) is a promising approach to alleviate such unwanted effects. Dissection of a known MAR allowed the identification of sequence motifs that mediate elevated transgene expression. Bioinformatics analysis implied that these motifs adopt a curved DNA structure that positions nucleosomes and binds specific transcription factors. From these observations, we computed putative MARs from the human genome. Cloning of several predicted MARs indicated that they are much more potent than the previously known element, boosting the expression of recombinant proteins from cultured cells as well as mediating high and sustained expression in mice. Thus we computationally identified potent epigenetic regulators, opening new strategies toward high and stable transgene expression for research, therapeutic production or gene-based therapies.

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Figure 1: Analysis of gene expression-activating portions of the chicken lysozyme MAR.
Figure 2: Computational analysis of the chicken lysozyme MAR element.
Figure 3: Use of SMARScan for the prediction of matrix attachment regions from genomic sequences.
Figure 4: Analysis of the effect of newly identified human MARs on gene transfer and expression.
Figure 5: MAR-driven expression of therapeutic genes in vitro and in vivo.

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Acknowledgements

We thank N. Besuchet-Schmutz, M. Wicht and other members of the Centre Hospitalier Universitaire Vaudois laboratory, A. Tamas and S. Meylan for skillful help and T. Forné for advice. This work was supported by grants from the Swiss NSF Priority Program in Biotechnology, the Swiss Commission for Technology and Innovation, Selexis SA, the Swiss Foundation for research on muscular dystrophies and by the Etat de Vaud.

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Authors

Contributions

P.-A.G., D.C., S.P., M.G., D.M., A.R. and D.S. performed experimental work, and D.-Q.C. carried out computational analysis. J.S.B. and P.B. supervised the genetic and computational work, and N.M. supervised experimental research and coordinated the project.

Corresponding author

Correspondence to Nicolas Mermod.

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Competing interests

Some of the genetic elements identified in this study are subject to a patent application, and their use is currently commercialized by a start-up biotechnology company, Selexis SA, of which N.M. is cofounder and shareholder. After the completion of this work, P.A.G., D.C. and A.R were hired by Selexis SA.

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Supplementary Figures 1–2, Supplementary Tables 1–3, Supplementary Methods (PDF 701 kb)

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Girod, PA., Nguyen, DQ., Calabrese, D. et al. Genome-wide prediction of matrix attachment regions that increase gene expression in mammalian cells. Nat Methods 4, 747–753 (2007). https://doi.org/10.1038/nmeth1076

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