The mitochondrial intracrine Stanniocalcin 1 (STC1) activates mitochondrial anti-oxidant defenses. LRP2 (megalin) shuttles STC1 to the mitochondria through retrograde early endosome-to-Golgi- and Rab32-mediated pathway, and LRP2 KO impairs mitochondrial respiration and glycolysis. We determined STC1-LRP2 interaction domains using HA- and FLAG-tagged fragments of STC1 and LRP2, respectively, co-expressed in HEK293T cells. The trans-membrane domain of LRP2 is required for trafficking to the mitochondria. STC1-FLAG expressed in LRP2 KO cells fails to reach the mitochondria; thus, mitochondrial STC1 is extracellularly-derived via LRP2-mediated trafficking. Tri-leucines L12-14 in LRP2's signal peptide interact with STC1's signal peptide. Mutant LRP2 (L(12-14)A) does not bind STC1, while hSTC1 lacking signal peptide or Leucines L8/9/11 does not bind LRP2. STC1 fails to induce respiration or glycolysis in megalin KO mouse embryonal fibroblasts (MEF) expressing mutant LRP2, while mutant hSTC1 (L8/L9/L11 - > A8/A9/A11) fails to reach the mitochondria or induce respiration and glycolysis in WT MEF. Our data suggest direct regulation of mitochondrial metabolism by extracellular cues and reveal an important role for signal peptides' leucines in protein-protein interactions and mitochondrial biology.
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