PLK1 stabilizes β-catenin to drive colorectal carcinogenesis through NFKB2-mediated transcriptional activation of USP2a and site-specific phosphorylation.

The Wnt/β-catenin signaling pathway is crucial in driving colorectal cancer (CRC), but therapeutic targeting is difficult due to on-target toxicity and adaptive resistance. Polo-like kinase 1 (PLK1), an essential regulator of mitosis, is known to stabilize β-catenin in various cancers. However, its specific mechanistic role in CRC, especially regarding the regulation of β-catenin ubiquitination, remains unclear. We integrated RNA-seq co-expression analysis with functional studies in CRC models, employing coimmunoprecipitation, ubiquitination assays, luciferase reporter systems, site-directed mutagenesis, and xenograft experiments. We identified a dual-axis mechanism through which PLK1 posttranslationally and transcriptionally controls β-catenin stability. First, PLK1 directly phosphorylates β-catenin at Ser311, which facilitates its recruitment to the deubiquitinating enzyme USP2a, thereby shielding β-catenin from proteasomal degradation. Second, PLK1 activates the transcription factor NFKB2, which in turn transcriptionally upregulates USP2a, amplifying the deubiquitination capacity of cells. This coordinated regulation ensures robust β-catenin nuclear accumulation and activation of downstream targets such as c-Myc and Cyclin D1. Inhibiting PLK1, either genetically or pharmacologically, leads to β-catenin destabilization and reduces CRC proliferation and . Rescue experiments established a mechanistic hierarchy: USP2a overexpression cannot restore β-catenin stability when Ser311 phosphorylation is abolished, whereas NFKB2 restoration rescues USP2a expression but not the PLK1 activity-dependent β-catenin‒USP2a interaction. Our study identified PLK1 as a key regulator of β-catenin signaling flexibility in CRC, coordinating kinase-dependent and transcriptional mechanisms to sustain pathway activation. The discovery of the PLK1-NFKB2-USP2a-β-catenin axis provides a novel therapeutic rationale for targeting PLK1 to selectively disrupt Wnt-driven tumorigenesis, potentially overcoming the toxicity limitations of conventional Wnt inhibitors.