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A fluorescent biosensor for measuring Ras activity in living cells

Weeks, Ryan, Zhou, Xin, Yuan, Tina and Zhang, Jin (2022) A fluorescent biosensor for measuring Ras activity in living cells. Journal of the American Chemical Society JACS.. ISSN 0002-78631520-5126


The small GTPase Ras is a critical regulator of cell growth and proliferation. Its activity is
frequently dysregulated in cancers, prompting decades of work to pharmacologically target Ras.
Understanding Ras biology and developing effective Ras therapeutics both require probing Ras
activity in its native context, yet tools to measure its activities in cellulo are limited. Here we
developed a ratiometric Ras activity reporter (RasAR) that provides quantitative measurement of
Ras activity in living cells with high spatiotemporal resolution. We demonstrated that RasAR can
probe live-cell activities of all the primary isoforms of Ras. Given that the functional roles of
different isoforms of Ras are intimately linked to their subcellular distribution and regulation, we
interrogated the spatiotemporal regulation of Ras utilizing subcellularly targeted RasAR and
uncovered the role of Src kinase as an upstream regulator to inhibit HRas. Furthermore, we
showed that RasAR enables capture of KRasG12C inhibition dynamics in living cells upon
treatment with KRasG12C covalent inhibitors, including ARS1620, Sotorasib, and Adagrasib.
We found in living cells a residual Ras activity lingers for hours in the presence of these
inhibitors. Together, RasAR represents a powerful molecular tool to enable live-cell
interrogation of Ras activity and facilitate the development of Ras inhibitors.

Item Type: Article
Date Deposited: 04 Oct 2022 00:45
Last Modified: 04 Oct 2022 00:46


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