Computationally designed sensors for endogenous Ras activity reveal signaling effectors within oncogenic granules
bioRxiv (Cold Spring Harbor Laboratory)(2022)
摘要
Abstract Genetically encoded biosensors have accelerated biological discovery, however many important targets such as active Ras (Ras-GTP) are difficult to sense as strategies to match a sensor’s sensitivity to the physiological range of target are lacking. Here, we use computational protein design to generate and optimize intracellular sensors of Ras activity ( LOCKR -based S ensor for Ras activity: Ras-LOCKR-S) and proximity labelers of the signaling environment of Ras ( LOCKR -based, Ras activity-dependent P roximity L abeler: Ras-LOCKR-PL). We demonstrate that our tools can measure endogenous Ras activity and environment at subcellular resolution. We illustrate the application of these tools by using them to identify Ras effectors, notably Src-Associated in Mitosis 68 kDa protein (SAM68), enriched in oncogenic EML4-Alk granules. Localizing these sensors to these granules revealed that SAM68 enhances Ras activity specifically at the granules, and SAM68 inhibition sensitizes EML4-Alk-driven cancer cells to existing drug therapies, suggesting a possible therapeutic strategy.
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关键词
endomembranes,oncogenic ras,sensors
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