title: A computationally engineered RAS rheostat reveals RAS-ERK signaling dynamics.
abstract: Synthetic protein switches controlled with user-defined inputs are powerful tools for studying and controlling dynamic cellular processes. To date, these approaches have relied primarily on intermolecular regulation. Here we report a computationally guided framework for engineering intramolecular regulation of protein function. We utilize this framework to develop chemically inducible activator of RAS (CIAR), a single-component RAS rheostat that directly activates endogenous RAS in response to a small molecule. Using CIAR, we show that direct RAS activation elicits markedly different RAS-ERK signaling dynamics from growth factor stimulation, and that these dynamics differ among cell types. We also found that the clinically approved RAF inhibitor vemurafenib potently primes cells to respond to direct wild-type RAS activation. These results demonstrate the utility of CIAR for quantitatively interrogating RAS signaling. Finally, we demonstrate the general utility of our approach in design of intramolecularly regulated protein tools by applying it to the Rho family of guanine nucleotide exchange factors.
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington, USA.
authors.forename: John C
authors.initials: JC
authors.lastname: Rose
authors.affiliation: Department of Biochemistry, University of Washington, Seattle, Washington, USA.|Institute for Protein Design, University of Washington, Seattle, Washington, USA.
authors.forename: Po-Ssu
authors.initials: PS
authors.lastname: Huang
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, Washington, USA.
authors.forename: Nathan D
authors.identifier: http://orcid.org/0000-0002-1216-5262
authors.initials: ND
authors.lastname: Camp
authors.affiliation: Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, California, USA.
authors.forename: Jordan
authors.initials: J
authors.lastname: Ye
authors.affiliation: Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, California, USA.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Leidal
authors.affiliation: Department of Biochemistry, University of Washington, Seattle, Washington, USA.
authors.forename: Inna
authors.initials: I
authors.lastname: Goreshnik
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington, USA.
authors.forename: Bridget M
authors.initials: BM
authors.lastname: Trevillian
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington, USA.
authors.forename: Miles S
authors.initials: MS
authors.lastname: Dickinson
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Cunningham-Bryant
authors.affiliation: Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, California, USA.
authors.forename: Jayanta
authors.identifier: http://orcid.org/0000-0002-8745-4069
authors.initials: J
authors.lastname: Debnath
authors.affiliation: Department of Biochemistry, University of Washington, Seattle, Washington, USA.|Institute for Protein Design, University of Washington, Seattle, Washington, USA.|Howard Hughes Medical Institute, University of Washington, Seattle, Washington, USA.
authors.forename: David
authors.initials: D
authors.lastname: Baker
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, Washington, USA.
authors.forename: Alejandro
authors.identifier: http://orcid.org/0000-0003-2909-3880
authors.initials: A
authors.lastname: Wolf-Yadlin
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington, USA.|Department of Biochemistry, University of Washington, Seattle, Washington, USA.
authors.forename: Dustin J
authors.initials: DJ
authors.lastname: Maly
mesh_terms: D002460:Cell Line; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D006801:Humans; D020935:MAP Kinase Signaling System; D008958:Models, Molecular; D015202:Protein Engineering; D018631:ras Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D048049:Extracellular Signal-Regulated MAP Kinases; D018631:ras Proteins