Active legumain promotes invasion and migration of neuroblastoma by regulating epithelial-mesenchymal transition

Abstract

Neuroblastoma (NB) is a commonly occurring malignancy in children. Epithelial-mesenchymal transition (EMT) is an adaptive change in promoting tumor metastasis. As an important factor in regulating tumor metastasis, whether legumain could promote metastasis of NB by EMT is still unexplored. Legumain is the active form of prolegumain, abundant in tumor plasma. So in the current study, different forms of legumain were identified in NB. Second, correlation analysis of N-cadherin and active legumain was identified by western blot analysis. Third, legumain gene amplification or gene knockdown were proceeded to examine the effect of legumain on EMT by scratch and transwell assay; meanwhile, active mature legumain or its asparagine endopeptidase (AEP) inhibitor was also added in. Finally, legumain can be detected differently in NB cells. Changes in legumain could influence NB metastasis by regulating EMT markers (e.g., N-cadherin, vimentin, and slug). Besides, the effect of legumain on EMT by its AEP activity was proved by intervention experiment of AEP gene transfection and gene knockdown experiments or adding recombinant human legumain suspension or specific inhibitor of AEP in NB cells (p < 0.05). These results suggest that legumain can promote invasion and migration of NB by regulating EMT, and EMT of NB is regulated by AEP activity of legumain, which can be inhibited by a specific AEP inhibitor.

Keywords

• legumain
• asparagine endopeptidase
• neuroblastoma
• epithelial-mesenchymal transition
• metastasis