Much of what is known about mammalian cell regulation has been achieved with the aid of transiently transfected cells. However, overexpression can violate balanced gene dosage, affecting protein folding, complex assembly and downstream regulation. To avoid these problems, genome engineering technologies now enable the generation of stable cell lines expressing modified proteins at (almost) native levels.
This is a preview of subscription content, access via your institution
Relevant articles
Open Access articles citing this article.
-
Branched actin networks are organized for asymmetric force production during clathrin-mediated endocytosis in mammalian cells
Nature Communications Open Access 22 June 2022
-
Identifying nucleic acid-associated proteins in Mycobacterium smegmatis by mass spectrometry-based proteomics
BMC Molecular and Cell Biology Open Access 23 March 2020
-
A CRISPR-based base-editing screen for the functional assessment of BRCA1 variants
Oncogene Open Access 29 August 2019
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
References
Gingras, A.C., Gstaiger, M., Raught, B. & Aebersold, R. Nat. Rev. Mol. Cell Biol. 8, 645–654 (2007).
Van Roey, K., Dinkel, H., Weatheritt, R.J., Gibson, T.J. & Davey, N.E. Sci. Signal. 6, rs7 (2013).
Martin, K.C. & Ephrussi, A. Cell 136, 719–730 (2009).
Beck, M. et al. Mol. Syst. Biol. 7, 549 (2011).
Schwanhäusser, B. et al. Nature 473, 337–342 (2011).
Gibson, T.J. Trends Biochem. Sci. 34, 471–482 (2009).
Dumont, J.E., Dremier, S., Pirson, I. & Maenhaut, C. Am. J. Physiol. Cell Physiol. 283, C2–C28 (2002).
Nooren, I.M. & Thornton, J.M. J. Mol. Biol. 325, 991–1018 (2003).
Perkins, J.R., Diboun, I., Dessailly, B.H., Lees, J.G. & Orengo, C. Structure 18, 1233–1243 (2010).
Volonté, C., D'Ambrosi, N. & Amadio, S. Prog. Neurobiol. 86, 61–71 (2008).
Georges, A.B., Benayoun, B.A., Caburet, S. & Veitia, R.A. FASEB J. 24, 346–356 (2010).
Ettwiller, L. & Veitia, R.A. Comp. Funct. Genomics 2007, 58721 (2007).
Birchler, J.A. & Veitia, R.A. Proc. Natl. Acad. Sci. USA 109, 14746–14753 (2012).
Makino, T. & McLysaght, A. Proc. Natl. Acad. Sci. USA 107, 9270–9274 (2010).
Okamoto, I. et al. Nature 472, 370–374 (2011).
Cacace, A.M. et al. Mol. Cell. Biol. 19, 229–240 (1999).
Levchenko, A., Bruck, J. & Sternberg, P.W. Proc. Natl. Acad. Sci. USA 97, 5818–5823 (2000).
Calabrese, E.J. & Baldwin, L.A. Trends Pharmacol. Sci. 22, 285–291 (2001).
Vavouri, T., Semple, J.I., Garcia-Verdugo, R. & Lehner, B. Cell 138, 198–208 (2009).
Rizzo, M. A., Davidson, M. W. & Piston, D. W. Cold Spring Harb. Protoc. 2009, pdb.top64 (2009).
Lehmann, O.J. et al. Am. J. Hum. Genet. 67, 1129–1135 (2000).
Liu, Y. & Lehmann, M. Fly (Austin) 2, 92–98 (2008).
Natesan, S., Rivera, V.M., Molinari, E. & Gilman, M. Nature 390, 349–350 (1997).
Peters, J.M. Nat. Rev. Mol. Cell Biol. 7, 644–656 (2006).
Nguyen, H.G., Chinnappan, D., Urano, T. & Ravid, K. Mol. Cell. Biol. 25, 4977–4992 (2005).
Stewart, S. & Fang, G. Cancer Res. 65, 8730–8735 (2005).
Dobson, C.M. Nature 426, 884–890 (2003).
Kutay, U. & Güttinger, S. Trends Cell Biol. 15, 121–124 (2005).
Diella, F. et al. Front. Biosci. 13, 6580–6603 (2008).
Hantschel, O. et al. Mol. Cell 19, 461–473 (2005).
Kadlec, J., Izaurralde, E. & Cusack, S. Nat. Struct. Mol. Biol. 11, 330–337 (2004).
Stirnimann, C.U., Ptchelkine, D., Grimm, C. & Müller, C.W. J. Mol. Biol. 400, 71–81 (2010).
Shen, D. et al. Cell Biochem. Biophys. 60, 173–185 (2011).
Roberti, M.J., Jovin, T.M. & Jares-Erijman, E. PLoS ONE 6, e23338 (2011).
Ciotta, G. et al. Methods 53, 113–119 (2011).
Poser, I. et al. Nat. Methods 5, 409–415 (2008).
Ding, L., Poser, I., Paszkowski-Rogacz, M. & Buchholz, F. Stem Cell Rev. 8, 32–42 (2012).
Maliga, Z. et al. Nat. Cell Biol. 15, 325–334 (2013).
Schebelle, L. et al. Nucleic Acids Res. 38, e106 (2010).
DeFrancesco, L. Nat. Biotechnol. 29, 681–684 (2011).
Dosztányi, Z., Csizmok, V., Tompa, P. & Simon, I. Bioinformatics 21, 3433–3434 (2005).
Letunic, I., Doerks, T. & Bork, P. Nucleic Acids Res. 37, D229–D232 (2009).
Waterhouse, A.M., Procter, J.B., Martin, D.M., Clamp, M. & Barton, G.J. Bioinformatics 25, 1189–1191 (2009).
D'Alise, A.M. et al. Mol. Cancer Ther. 7, 1140–1149 (2008).
Dong, X. et al. Nature 458, 1136–1141 (2009).
Taagepera, S. et al. Proc. Natl. Acad. Sci. USA 95, 7457–7462 (1998).
Fries, B. et al. J. Biol. Chem. 282, 4504–4515 (2007).
Giannini, A. et al. Exp. Cell Res. 295, 150–160 (2004).
Ossovskaya, V., Lim, S.T., Ota, N., Schlaepfer, D.D. & Ilic, D. FEBS Lett. 582, 2402–2406 (2008).
Roovers, K., Klein, E.A., Castagnino, P. & Assoian, R.K. Dev. Cell 5, 273–284 (2003).
Murai, N., Murakami, Y. & Matsufuji, S. J. Biol. Chem. 278, 44791–44798 (2003).
Xiao, Z., Watson, N., Rodriguez, C. & Lodish, H.F. J. Biol. Chem. 276, 39404–39410 (2001).
Begitt, A., Meyer, T., van Rossum, M. & Vinkemeier, U. Proc. Natl. Acad. Sci. USA 97, 10418–10423 (2000).
Kulisz, A. & Simon, H.G. Mol. Cell. Biol. 28, 1553–1564 (2008).
Shirley, R.L., Ford, A.S., Richards, M.R., Albertini, M. & Culbertson, M.R. Genetics 161, 1465–1482 (2002).
Turan, S. et al. J. Mol. Biol. 407, 193–221 (2011).
Osterwalder, M. et al. Nat. Methods 7, 893–895 (2010).
Kim, Y.G., Cha, J. & Chandrasegaran, S. Proc. Natl. Acad. Sci. USA 93, 1156–1160 (1996).
Zhang, F. et al. Nat. Biotechnol. 29, 149–153 (2011).
Boch, J. Nat. Biotechnol. 29, 135–136 (2011).
Cong, L. et al. Science 339, 819–823 (2013).
Mali, P. et al. Science 339, 823–826 (2013).
Casola, S. Methods Mol. Biol. 667, 145–163 (2010).
Soriano, P. Nat. Genet. 21, 70–71 (1999).
Nord, A.S. et al. Nucleic Acids Res. 34, D642–D648 (2006).
Skarnes, W.C. et al. Nature 474, 337–342 (2011).
Huh, W.K. et al. Nature 425, 686–691 (2003).
Winzeler, E.A. et al. Science 285, 901–906 (1999).
Dietzl, G. et al. Nature 448, 151–156 (2007).
Acknowledgements
We thank many colleagues at the European Molecular Biology Laboratory, Monod Institute and two research consortia, the German National Genome Research Network–funded DiGtoP and European Union–funded SyBoSS—which are focused on developing and applying genome engineered cell lines—for useful discussions. We apologize for not citing many important references because of space limitations. M.S. is funded by DiGtoP. R.A.V. is supported by Centre National de la Recherche Scientifique, La Ligue contre le Cancer (Comité de Paris), l'Université Paris Diderot–Paris 7 and Institut Universitaire de France. Special thanks to I. Poser, M. Augsburg and A. Nitzsche (Max Planck Institute of Molecular Cell Biology and Genetics, Dresden) for providing the images of stably engineered cell lines.
Author information
Authors and Affiliations
Corresponding authors
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Rights and permissions
About this article
Cite this article
Gibson, T., Seiler, M. & Veitia, R. The transience of transient overexpression. Nat Methods 10, 715–721 (2013). https://doi.org/10.1038/nmeth.2534
Published:
Issue Date:
DOI: https://doi.org/10.1038/nmeth.2534
This article is cited by
-
Gene augmentation for autosomal dominant retinitis pigmentosa using rhodopsin genomic loci nanoparticles in the P23H+/− knock-in murine model
Gene Therapy (2023)
-
Branched actin networks are organized for asymmetric force production during clathrin-mediated endocytosis in mammalian cells
Nature Communications (2022)
-
RTG-TOF, a rainbow trout (Oncorhynchus mykiss) cell line with an inducible gene expression system
In Vitro Cellular & Developmental Biology - Animal (2022)
-
A guide to the optogenetic regulation of endogenous molecules
Nature Methods (2021)
-
Identifying nucleic acid-associated proteins in Mycobacterium smegmatis by mass spectrometry-based proteomics
BMC Molecular and Cell Biology (2020)