Shroom3 在 Wnt/平面细胞极性信号下游对乳腺癌细胞迁移的作用

The 5-year survival rate for patients diagnosed with metastatic breast cancer (BC) remains below 30%, highlighting the need to better understand the molecular mechanisms driving BC metastasis in order to reveal novel opportunities for therapeutic intervention. Wnt/planar cell polarity (Wnt/PCP) is a non-canonical Wnt signaling pathway that regulates epithelial tissue organization and cell migration during embryonic development. Core Wnt/PCP components are upregulated in more aggressive BCs, and their expression is associated with poor patient outcomes. Our group has demonstrated a role for Wnt/PCP in promoting BC cell dissemination; however, the mechanistic details linking core Wnt/PCP components to downstream actin cytoskeletal rearrangements that drive migration and invasion are not well understood. We therefore performed a phosphoproteomics screen using MDA-MB-231 triple negative BC cells to identify proteins differentially phosphorylated in response to pathway activation by the non-canonical Wnt ligand Wnt5a. Among the proteins identified was Shroom3 (Shrm3), in which two serine residues were significantly phosphorylated in response to Wnt5a treatment. Shrm3 is an F-actin-binding scaffolding protein known to recruit Rho kinase (ROCK), actin, and myosin to the apex of apically constricting cells to drive tissue morphogenesis during development and has previously been shown to function downstream of the Wnt/PCP pathway and to bind the effector Dvl2. Given its described functions, Shrm3 is a promising candidate to serve as a direct link between core Wnt/PCP components and the actin cytoskeleton. Knockdown (KD) of Shrm3 reduces both basal and Wnt/PCP-mediated BC cell migration, as well as the formation of actin-rich protrusions at the leading edge of migrating cells. Live-cell imaging confirms reduced lamellipodia formation and reveals that Shrm3 KD cell protrusions are shorter-lived, retract more quickly, and are less polarized to the leading edge. Additionally, loss of Shrm3 reduces activation of myosin light chain and myosin recruitment to the leading edge of BC cells. Co-localization of Shrm3, ROCK, and core Wnt/PCP component Vangl1 is observed at migratory protrusions, and we also observe Shrm3 localizing to focal adhesions (FAs). Loss of Shrm3 reduces FA number and size and may reduce cell-substrate adhesion. Whether phosphorylation of Shrm3 downstream of Wnt/PCP activation regulates these functions is currently under investigation. Based on these findings, we suggest a model in which core Wnt/PCP components recruit Shrm3 to the leading edge of migrating BC cells, where Shrm3 recruits ROCK, actin, and myosin to promote properly localized contractility that supports lamellipodia stabilization and focal adhesion maturation.