Abstract
Urokinase plasminogen activator receptor (uPAR) is known to promote invasion, migration, and metastasis in cancer cells. In this report, we showed that ionizing radiation (IR)-induced uPAR has a role in WNT-β-catenin signaling and mediates induction of cancer stem cell (CSC)-like properties in medulloblastoma cell lines UW228 and D283. We observed that IR induced the expression of uPAR and CSC markers, such as Musashi-1 and CD44, and activated WNT-7a-β-catenin signaling molecules. Overexpression of uPAR alone or with IR treatment led to increased WNT-7a-β-catenin-TCF/LEF-mediated transactivation, thereby promoting cancer stemness. In contrast, treatment with shRNA specific for uPAR (pU) suppressed WNT-7a-β-catenin-TCF/LEF-mediated transactivation both in vitro and in vivo. Quercetin, a potent WNT/β-catenin inhibitor, suppressed uPAR and uPAR-mediated WNT/β-catenin activation, and furthermore, addition of recombinant human WNT-7a protein induced uPAR, indicating the existence of a mutual regulatory relationship between uPAR and WNT/β-catenin signaling. We showed that uPAR was physically associated with the WNT effector molecule β-catenin on the membrane, cytoplasm, and nucleus of IR-treated cells and CSC. Most interestingly, we demonstrated for the first time that localization of uPAR in the nucleus was associated with transcription factors (TF) and their specific response elements. We observed from uPAR-ChIP, TF protein, and protein/DNA array analyses that uPAR associates with activating enhancer-binding protein 2α (AP2a) and mediates β-catenin gene transcription. Moreover, association of uPAR with the β-catenin·TCF/LEF complex and various other TF involved during embryonic development and cancer indicates that uPAR is a potent activator of stemness, and targeting of uPAR in combination with radiation has significant therapeutic implications.
Highlights
UPAR is a multifunctional protein, overexpressed in all human cancers
We showed that ionizing radiation (IR)-induced Urokinase plasminogen activator receptor (uPAR) has a role in WNT--catenin signaling and mediates induction of cancer stem cell (CSC)-like properties in medulloblastoma cell lines UW228 and D283
To determine whether IR-induced uPAR has a role in transcriptional activation of WNT/-catenin signaling necessary for CSC, we performed real time PCR in control, Fl-uPAR, CSCs alone, and in combination with IR treatments
Summary
Results: uPAR overexpression with radiation enhances -catenin stabilization, -catenin gene transcription, and WNT-7a-catenin-TCF/LEF-mediated transactivation. Conclusion: Association of uPAR with -catenin and various transcription factors involved in embryonic development suggests that uPAR is a potent activator of cancer stemness. We showed that ionizing radiation (IR)-induced uPAR has a role in WNT--catenin signaling and mediates induction of cancer stem cell (CSC)-like properties in medulloblastoma cell lines UW228 and D283. Overexpression of uPAR alone or with IR treatment led to increased WNT-7a--catenin-TCF/LEF-mediated transactivation, thereby promoting cancer stemness. Treatment with shRNA specific for uPAR (pU) suppressed WNT-7a--catenin-TCF/LEF-mediated transactivation both in vitro and in vivo. Association of uPAR with the -catenin1⁄7TCF/LEF complex and various other TF involved during embryonic development and cancer indicates that uPAR is a potent activator of stemness, Grant CA138409 R.). □S This article contains supplemental Figs. 1–3, Tables 1– 4, and Materials and
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