Abstract
Few regulators of K-Ras plasma membrane organization are known. We combined TurboID-based proximity proteomics with a BRET screen to identify eight potential K-Ras G-domain interactors. We focused on APLP2, which indirectly engages with C-Raf in immediate proximity to K-Ras, and SPRY2, which exhibits properties of an effector. Co-immunoprecipitation and BRET assays revealed that the SPRY2 C-terminal half binds oncogenic RasG12V more than full-length SPRY2. Both forms localize to the plasma membrane, but this localization and binding to K-Ras are disrupted by inhibitors of K-Ras membrane anchorage or activity. Mutations at the predicted interface of K-Ras and SPRY2's C-terminal region affect the interaction. Both full-length SPRY2 and its C-terminal fragment promote the differentiation of C2C12 muscle cells, a process requiring MAPK pathway inhibition. Finally, SPRY2 homo- and hetero-oligomerizes with SPRY4. We propose that active K-Ras recruits SPRY2 dimers to the membrane, where they block Ras effector access.