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Biophysical Constraints Arising from Compositional Context in Synthetic Gene Networks

Yeung, Enoch and Dy, Aaron J. and Martin, Kyle B. and Ng, Andrew H. and Del Vecchio, Domitilla and Beck, James L. and Collins, James J. and Murray, Richard M. (2017) Biophysical Constraints Arising from Compositional Context in Synthetic Gene Networks. Cell Systems, 5 (1). pp. 11-24. ISSN 2405-4712. https://resolver.caltech.edu/CaltechAUTHORS:20170731-080503943

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Abstract

Synthetic gene expression is highly sensitive to intragenic compositional context (promoter structure, spacing regions between promoter and coding sequences, and ribosome binding sites). However, much less is known about the effects of intergenic compositional context (spatial arrangement and orientation of entire genes on DNA) on expression levels in synthetic gene networks. We compare expression of induced genes arranged in convergent, divergent, or tandem orientations. Induction of convergent genes yielded up to 400% higher expression, greater ultrasensitivity, and dynamic range than divergent- or tandem-oriented genes. Orientation affects gene expression whether one or both genes are induced. We postulate that transcriptional interference in divergent and tandem genes, mediated by supercoiling, can explain differences in expression and validate this hypothesis through modeling and in vitro supercoiling relaxation experiments. Treatment with gyrase abrogated intergenic context effects, bringing expression levels within 30% of each other. We rebuilt the toggle switch with convergent genes, taking advantage of supercoiling effects to improve threshold detection and switch stability.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1016/j.cels.2017.06.001DOIArticle
http://www.sciencedirect.com/science/article/pii/S2405471217302296?via%3DihubPublisherArticle
ORCID:
AuthorORCID
Murray, Richard M.0000-0002-5785-7481
Additional Information:© 2017 Elsevier Inc. Received: December 24, 2015; Received in revised form: February 8, 2017; Accepted: June 5, 2017; Published: July 19, 2017. We thank Ophelia Venturelli for her invaluable inspiration and guidance in this project; Victoria Hsiao, Jin Park, Anu Thubagere, and Adam Rosenthal for great advice on imaging; David Younger, Ania Baetica, Vincent Noireaux, Clarmyra Hayes, and Zachary Z. Sun for guidance and assistance with TX-TL experiments; and Lea Goentoro, Johann Pauls-son, Long Cai, Jennifer Brophy, John Doyle, Eric Klavins, and Julius Lucks for insightful conversations. This work was supported in part by a Charles Lee Powell Foundation Fellowship, a Kanel Foundation Fellowship, a National Science Foundation Graduate Fellowship, a National Defense Science and Engineering Graduate Fellowship, Air Force Office of Scientific Research grant (AFOSR) FA9550-14-1-0060, Defense Threat Reduction Agency grant HDTRA1-14-1-0006, and Defense Advanced Research Projects Agency grant HR0011-12-C-0065. Author Contributions: E.Y. wrote the paper. E.Y. and A.J.D. designed the experiments. K.B.M., A.H.N., A.J.D., J.L.B., and D.D.V. reviewed and edited the paper. E.Y. and A.H.N. developed models and performed formal analysis. E.Y., A.J.D., K.B.M., and A.H.N. conducted the experiments. R.M.M., D.D.V., and J.J.C. secured funding. R.M.M., D.D.V., J.L.B., and J.J.C. supervised modeling and experimental research design.
Funders:
Funding AgencyGrant Number
Charles Lee Powell FoundationUNSPECIFIED
Kanel FoundationUNSPECIFIED
NSF Graduate Research FellowshipUNSPECIFIED
National Defense Science and Engineering Graduate Fellowship (NDSEG)UNSPECIFIED
Air Force Office of Scientific Research (AFOSR)FA9550-14-1-0060
Defense Threat Reduction AgencyHDTRA1-14-1-0006
Defense Advanced Research Projects Agency (DARPA)HR0011-12-C-0065
Subject Keywords:synthetic biology, genetic context, compositional context, transcriptional regulation, supercoiling
Issue or Number:1
Record Number:CaltechAUTHORS:20170731-080503943
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20170731-080503943
Official Citation:Enoch Yeung, Aaron J. Dy, Kyle B. Martin, Andrew H. Ng, Domitilla Del Vecchio, James L. Beck, James J. Collins, Richard M. Murray, Biophysical Constraints Arising from Compositional Context in Synthetic Gene Networks, Cell Systems, Volume 5, Issue 1, 2017, Pages 11-24.e12, ISSN 2405-4712, http://dx.doi.org/10.1016/j.cels.2017.06.001. (http://www.sciencedirect.com/science/article/pii/S2405471217302296)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:79576
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:31 Jul 2017 15:19
Last Modified:03 Oct 2019 18:21

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