Backgroud: Tumor heterogeneity complicates treatment of colorectal cancer (CRC) patients, as it leads to different responses to therapies. Polyploid giant cancer cells (PGCCs) are a rare special subpopulation of heterogeneous cancer cells that are a source of solid tumor heterogeneity. Daughter cells derived from PGCCs through asymmetric cell division exhibit strong migration, invasion and proliferation abilities. However, the underlying molecular mechanism concerning the malignant biological properties in daughter cells remains unclear. Methods: In this study, PGCCs were successfully induced in CRC cells via chemotherapy, using 5-fluorouracil and oxaliplatin. Oncomine, UALCAN, GEPIA, and HPA databases were used to analyze the expression of PTPRN2 and its effect on the survival rate of CRC patients was evaluated on the univariate and multivariate analysis of the GSE39582 dataset. The cellular aggressive phenotype and cell cycle were detected by flow cytometry, transwell, wound healing, and colony formation assays. Transient transfection, western blotting, and co-immunoprecipitation assays were used to analyze molecular mechanism. Results: PTPRN2 is significantly up-regulated compared to that in normal colorectal lesions at the mRNA and protein levels. Patients with CRC exhibiting higher PTPRN2 expression had an unfavorable overall survival. The expression of PTPRN2 was signally elevated in PGCCs with daughter cells whose migration, invasion, and proliferation abilities were significantly weakened after PTPRN2 knockdown. Mechanistic investigations suggested that the expression of the proteins in the MAPK pathway, epithelial-mesenchymal transition (EMT) process, and STAT3/cyclin D1 signaling decreased markedly after PTPRN2 knockdown in PGCCs and their daughter cells. In addition, PTPRN2 exerted these effects through interactions with tumor necrosis factor receptor-associated factor 2 (TRAF2), twist1, and cyclin D1. In PGCCs and their daughter cells, TRAF2 is mainly K63-ubiquitinated, which is necessary for the activation of the MAPK pathway. After PTPRN2 was inhibited, the K63-linked ubiquitination of TRAF2 reduced. Conclusions: PTPRN2 enhanced the malignant transformation in PGCCs and their daughter cells by mediating the EMT process, STAT3/cyclin D1, and MAPK signaling pathways, interacting with twist1, cyclin D1, and TRAF2, and especially accelerating the K63-linked ubiquitination chain attached to TRAF2. Overall, our results indicate that PTPRN2 may be a novel prognostic and theraeputic target for CRC treatment.