Abstract
Here we uncovered the involved subcellular mechanisms that sulforaphane-cysteine (SFN-Cys) inhibited invasion in human glioblastoma (GBM). SFN-Cys significantly upregulated 45 and downregulated 14 microtubule-, mitophagy-, and invasion-associated proteins in GBM cells via HPLC–MS/MS and GEO ontology analysis; SFN-Cys disrupted microtubule by ERK1/2 phosphorylation-mediated downregulation of α-tubulin and Stathmin-1 leading to the inhibition of cell migration and invasion; SFN-Cys downregulated invasion-associated Claudin-5 and S100A4, and decreased the interaction of α-tubulin to Claudin-5. Knockdown of Claudin-5 and S100A4 significantly reduced the migration and invasion. Besides, SFN-Cys lowered the expressions of α-tubulin-mediated mitophagy-associated proteins Bnip3 and Nix. Transmission electron microscopy showed more membrane-deficient mitochondria and accumulated mitophagosomes in GBM cells, and mitochondria fusion might be downregulated because that SFN-Cys downregulated mitochondrial fusion protein OPA1. SFN-Cys increased the colocalization and interplay of LC3 to lysosomal membrane-associated protein LAMP1, aggravating the fusion of mitophagosome to lysosome. Nevertheless, SFN-Cys inhibited the lysosomal proteolytic capacity causing LC3II/LC3I elevation but autophagy substrate SQSTM1/p62 was not changed, mitophagosome accumulation, and the inhibition of migration and invasion in GBM cells. These results will help us develop high-efficiency and low-toxicity anticancer drugs to inhibit migration and invasion in GBM.
Highlights
High invasion in human glioblastoma (GBM) is the key reason to cause low survival and bad outcome[1]
Gene expression analysis by GEPIA server showed that the expressions of TUBA1C, S100A4, CLDN5 (Claudin-5), MAPK1, LAMP1, LAMP2, BNIP3L (Nix), MAP1LC3B (LC3), PODXL, MMP-2, ITGB3, CD47, and FN1 were upregulated, while SEMA7A expression was downregulated in GBM tissues (Fig. 1b)
The data obtained from Gene Correlation Analysis showed that the expression of TUBA1C was positively correlated to the expression of CLDN5 and S100A4; MAP1LC3B expression was positively correlated to the expressions of Bnip[3], Nix and LAMP1; LAMP1 expression was positively correlated to S100A4 expression in GBM (Fig. 1e)
Summary
High invasion in human glioblastoma (GBM) is the key reason to cause low survival and bad outcome[1]. Plantderived sulforaphane (SFN) induced apoptosis and inhibited invasion by ERK1/2 phosphorylation in various cancers[2,3,4,5]. We previously reported that SFN-Cys inhibited migration and invasion via microtubulemediated Claudins dysfunction in human non-small cell. Cys suppressed invasion via downregulating Galectin-1 in human prostate cancer[8]. At subcellular levels we will determine how SFN-Cys inhibited migration and invasion in human GBM to establish a high-efficiency anticancer chemotherapy. Actins and tubulins form highly dynamic polymers that are capable of organizing cytoplasmic organelles, determining cell shape and polarity and promoting cell–cell and cell–matrix adhesions through their interactions with cadherins and integrins, respectively[10]. Studies showed that microtubule-associated protein Stathmin-1 binds to tubulins promoting the depolymerization of microtubule[11], while the inhibition of stathmin expression significantly reduced transendothelial
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