Succinate and its carrier Sfc1 mediate metabolic control of mitochondrial protein import by the TIM23 translocase

Abstract Tim23 is an essential component of the mitochondrial inner membrane translocase and Sfc1 is a carrier that exchanges succinate for fumarate across that membrane. Sfc1 and succinic acid availability regulate dual targeting of fumarase and aconitase by facilitating mitochondrial import of their newly synthesized precursors, as shown by pulse-chase experiments. Here we show that Sfc1 directly interacts with the Tim23 and succinate affects this interaction, which in turn affects mitochondrial protein import. Physical interaction between Tim23 and Sfc1 was proven by co immunoprecipitation, Bimolecular Fluorescence Complementation (BiFC) and Biotin-based proximity labeling (TurboID). Proximity labeling-informed mutagenesis allowed us to dissect the carrier activity of Sfc1 from its function as a TIM23 regulator. We performed Rosetta-MP docking of Sfc1 and Tim23 to envisage the interface. Thus, our findings show that metabolites can regulate mitochondrial import and adjust the segregation of key metabolic enzymes between the cytosol and mitochondria. Competing Interest Statement The authors have declared no competing interest. Footnotes ↵5 Lead contact Data availability The data supporting the findings of this work are available within the paper and/ or its supplementary information or on request from the corresponding author [O.P].