Sulforaphane downregulated fatty acid synthase and inhibited microtubule-mediated mitophagy leading to apoptosis.

We previously demonstrated that sulforaphane (SFN) inhibited autophagy leading to apoptosis in human non-small cell lung cancer (NSCLC) cells, but the underlying subcellular mechanisms were unknown. Hereby, high-performance liquid chromatography-tandem mass spectrometry uncovered that SFN regulated the production of lipoproteins, and microtubule- and autophagy-associated proteins. Further, highly expressed fatty acid synthase (FASN) contributed to cancer malignancy and poor prognosis. Results showed that SFN depolymerized microtubules, downregulated FASN, and decreased its binding to α-tubulin; SFN downregulated FASN, acetyl CoA carboxylase (ACACA), and ATP citrate lyase (ACLY) via activating proteasomes and downregulating transcriptional factor SREBP1; SFN inhibited the interactions among α-tubulin and FASN, ACACA, and ACLY; SFN decreased the amount of intracellular fatty acid (FA) and mitochondrial phospholipids; and knockdown of FASN decreased mitochondrial membrane potential (ΔΨm) and increased reactive oxygen species, mitochondrial abnormality, and apoptosis. Further, SFN downregulated mitophagy-associated proteins Bnip3 and NIX, and upregulated mitochondrial LC3 II/I. Transmission electron microscopy showed mitochondrial abnormality and accumulation of mitophagosomes in response to SFN. Combined with mitophagy inducer CCCP or autophagosome-lysosome fusion inhibitor Bafilomycin A1, we found that SFN inhibited mitophagosome-lysosome fusion leading to mitophagosome accumulation. SFN reduced the interaction between NIX and LC3 II/I, and reversed CCCP-caused FA increase. Furthermore, knockdown of α-tubulin downregulated NIX and BNIP3 production, and upregulated LC3 II/I. Besides, SFN reduced the interaction and colocalization between α-tubulin and NIX. Thus, SFN might cause apoptosis via inhibiting microtubule-mediated mitophagy. These results might give us a new insight into the mechanisms of SFN-caused apoptosis in the subcellular level.