ROCK2 promotes osteosarcoma growth and metastasis by modifying PFKFB3 ubiquitination and degradation

Abstract

Rho-associated coiled-coil-containing protein kinase 2 (ROCK2) and 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) are widely involved in cell biological activities and play a key role in controlling various cell phenomena. However, the underlying mechanisms connecting ROCK2 and PFKFB3 in osteosarcoma growth and metastasis are poorly understood. In this study, we explored and analysed the role and molecular mechanism of ROCK2 and PFKFB3 in osteosarcoma. We analysed ROCK2 and PFKFB3 protein expression in 51 surgical specimens from osteosarcoma patients and determined the correlation between ROCK2 and PFKFB3. In addition, we used Transwell and wound-healing assays to detect cell invasion and migration and CCK8 and EdU assays to assess cell proliferation. Herein, we confirmed that ROCK2 and PFKFB3 proteins were significantly upregulated in osteosarcoma compared with adjacent normal tissues. Further studies revealed that knockdown of ROCK2 significantly decreased the expression levels of PFKFB3; moreover, growth and metastasis were decreased in shROCK2 osteosarcoma cells. Additionally, upregulation of PFKFB3 rescued the decreased proliferation and metastasis induced by ROCK2 knockdown, whereas knockdown of PFKFB3 decreased ROCK2-enhanced osteosarcoma proliferation and metastasis. These results suggest that PFKFB3 is essential for ROCK2-mediated proliferation and metastasis of osteosarcoma cells. Mechanistically, ROCK2 stabilizes PFKFB3 expression by modifying its ubiquitination and degradation. Taken together, our results link two drivers of proliferation and metastasis in osteosarcoma and identify a novel pathway for PFKFB3 regulation. Thus, we provide new evidence of the biological and clinical significance of PFKFB3 as a potential biomarker for osteosarcoma.

Keywords: Invasion; Migration; PFKFB3; Proliferation; ROCK2; Ubiquitination.