Dephosphorylation of C20orf112 by PPP3CA/PPIA drives its nuclear translocation to promote colorectal cancer stemness.

Proteins with incomplete functional characterization represent a major opportunity to uncover novel mechanisms underlying tumor progression. C20orf112 (also known as NOL4L) has been implicated in tumor biology, yet its regulatory mechanisms and role in colorectal cancer (CRC) remain largely undefined. Here, using a sphere formation-colony formation swapping culture model, we identified C20orf112 as a potent regulator of CRC stemness and elucidated its underlying mechanism. Immunohistochemical (IHC) analysis revealed that C20orf112 is significantly overexpressed in CRC tissues compared with adjacent noncancerous tissues. Functional studies demonstrated that C20orf112 enhances cancer stemness in CRC by activating ERK signaling. Mechanistically, C20orf112 is dephosphorylated at serine 295 (S295) by the phosphatase complex PPP3CA/PPIA, promoting its nuclear translocation. This dephosphorylation increases the binding affinity between C20orf112 and karyopherin KPNA2, facilitating nuclear import. Subsequent nuclear accumulation of C20orf112 is associated with sustained ERK activation and promotes a stem-like phenotype. Collectively, these findings reveal a previously unrecognized PPP3CA/PPIA-KPNA2-ERK signaling axis that regulates C20orf112 function in CRC, highlighting its context-specific regulatory role and therapeutic potential.