Inhibition of mitochondrial protein import and proteostasis by a pro-apoptotic lipid

Abstract

AbstractMitochondria mediated cell death is critically regulated by bioactive lipids derived from sphingolipid metabolism. The lipid aldehyde trans-2-hexadecenal (t-2-hex) induces mitochondrial dysfunction in a conserved manner from yeast to humans. Here we apply unbiased transcriptomic, functional genomics and chemoproteomic approaches in the yeast model to uncover the principal mechanisms and biological targets underlying this lipid-induced mitochondrial inhibition. We find that loss of Hfd1 fatty aldehyde dehydrogenase function efficiently sensitizes cells for t-2-hex inhibition and apoptotic cell death. Excess of t-2-hex causes a profound transcriptomic response with characteristic hallmarks of impaired mitochondrial protein import like activation of mitochondrial and cytosolic chaperones or proteasomal function and severe repression of translation. We confirm that t-2-hex stress induces rapid accumulation of mitochondrial pre-proteins and protein aggregates and subsequent activation of Hsf1- and Rpn4-dependent gene expression. By saturated transposon mutagenesis we find that t-2-hex tolerance requires an efficient heat shock response and specific mitochondrial and ER functions and that mutations in ribosome, protein and amino acid biogenesis are beneficial upon t-2-hex stress. We further show that genetic and pharmacological inhibition of protein translation causes t-2-hex resistance indicating that loss of proteostasis is the predominant consequence of the pro-apoptotic lipid. Hfd1 associates with the Tom70 subunit of the TOM complex and t-2-hex covalently lipidates the central Tom40 channel, which altogether indicates that transport of mitochondrial precursor proteins through the outer mitochondrial membrane is directly inhibited by the pro-apoptotic lipid and thus represents a hotspot for pro- and anti-apoptotic signaling.