The COPS3-FOXO3 positive feedback loop regulates autophagy to promote cisplatin resistance in osteosarcoma

Abstract

ABSTRACT Chemotherapy is an important treatment modality for osteosarcoma (OS), but the development of chemoresistance limits the therapeutic efficacy of OS and results in a poor prognosis. Thus, a better understanding of the mechanisms underlying chemoresistance in OS is essential. We previously demonstrated that COPS3/CSN3 (COP9 signalosome subunit 3) functions as an oncogene to promote OS cells lung metastasis, which is closely related to chemoresistance. Here, we showed that COPS3 was significantly upregulated in OS tissues with poor response to preoperative chemotherapy. Moreover, COPS3 depletion made OS cells more sensitive to cisplatin treatment in vitro and in vivo, implicating COPS3 as a driver of cisplatin resistance. Mechanistic investigations showed that COPS3 induced a cytoprotective macroautophagy/autophagy in response to cisplatin. Specifically, we identified FOXO3 as a critical target of COPS3, as high expression of COPS3 enhanced the nuclear abundance of FOXO3 and increased the expression of FOXO3-responsive genes, promoting autophagosome formation and maturation. In turn, FOXO3 regulated COPS3 levels by inhibiting ubiquitin-mediated degradation and attenuating SKP2-mediated COPS3 inhibition, cooperatively maintaining a high level of COPS3. In both COPS3-expressing OS cells and a murine xenograft model, inhibition of autophagy could also overcome resistance to cisplatin. Collectively, our results offer insights into the mechanisms of cisplatin resistance and suggest that targeting COPS3-mediated autophagy is a promising therapeutic strategy for overcoming the cisplatin resistance of OS. Abbreviations: 3-MA: 3-methyladenine; BECN1: beclin 1; ChIP: chromatin immunoprecipitation; CHX: cycloheximide; COPS3/CSN3: COP9 signalosome subunit 3; CQ: chloroquine; DEGs: differentially expressed genes; FOXO3: forkhead box O3; GFP: green fluorescent protein; IC50: 50% inhibitory concentration; LAMP1: lysosomal associated membrane protein 1; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; mRFP: monomeric red fluorescent protein; OS: osteosarcoma; PBS: phosphate-buffered saline; qRT-PCR: quantitative real-time PCR; RAB7: RAB7, member RAS oncogene family; RPS6KB1/p70S6K1: ribosomal protein S6 kinase B1; SEM: standard error of the mean; shRNA: short hairpin RNA; siRNA: small interfering RNA; SKP2: S-phase kinase associated protein 2; TEM: transmission electron microscopy; UPS: ubiquitin-proteasome system