Abstract: FR-PO161

Megalin-Mediated Shuttling of Angiotensin II, TGF-ß, and Stanniocalcin-1 to the Mitochondria

Session Information

  • Mitochondriacs and More
    November 03, 2017 | Location: Hall H, Morial Convention Center
    Abstract Time: 10:00 AM - 10:00 AM

Category: Cell Biology

  • 201 Cell Signaling, Oxidative Stress

Authors

  • Li, Qingtian, Baylor College of Medicine, Houston, Texas, United States
  • Lei, Fan, Baylor College of Medcine, Houston, Texas, United States
  • Tang, Yi, West China Hospital of Sichuan University, Chengdu, China
  • Pan, Jenny S., Baylor College of Medicine, Houston, Texas, United States
  • Tong, Qiang, Baylor College of Medicine, Houston, Texas, United States
  • Sheikh-Hamad, David, Baylor College of Medicine, Houston, Texas, United States
Background

Some extracellular signaling molecules are detected in the mitochondria, including angiotensin II, insulin, stanniocalcin-1 (STC1), TGF-ß and erythropoietin; these are known as mitochondrial intracrines. The mechanism of mitochondrial targeting of these proteins is unknown. Megalin/LRP2 is highly expressed on the apical surface of kidney proximal tubule cells, where it is involved in the uptake/reclamation of filtered vitamins, uptake and lysosomal degradation of filtered proteins, uptake of hormones including angiotensin II and angiotensin 1-7. Megalin mutations are linked to the pathogenesis of Donnai-Barrow and Lowe syndromes, characterized by developmental brain abnormalities and kidney dysfunction. Megalin has not been shown to reside in the mitochondria.

Results

Mass spectroscopy analysis of mitochondrial acetyl proteome from kidneys of stanniocalcin-1 transgenic mice identified megalin as a resident mitochondrial protein. Megalin is also present in the mitochondria of a number of cell lines, including 293T, C2C12 and Raw267.4 cells, and associates stanniocalcin-1 and the mitochondrial longevity protein SIRT3. These observations were validated using immunoprecipitation, confocal immunofluorescence, electron microscopy and co-expression experiments.
Because megalin, stanniocalcin-1, angiotensin II and TGF-ß are found at the plasma membrane and mitochondria, we hypothesized that megalin serves as a shuttle for mitochondrial intracrines from the cell surface to the mitochondria. CRISPR-Cas9-mediated knockdown of megalin in C2C12 cells diminishes whole-cell and mitochondrial content of megalin. Upon the addition of recombinant hSTC1-FLAG, hSTC1-FITC, angiotensin II-FITC or TGF-ß-FITC to the medium of WT C2C12 cells, we detect the corresponding FLAG or FITC within the mitochondria; however, we observe diminished presence of STC1-FLAG, STC1-FITC, angiotensin II-FITC or TGF-ß-FITC in the mitochondria of megalin knockdown cells.

Conclusion

The data suggest that megalin is present in the mitochondria and exists in complex with proteins involved in anti-oxidant defenses. Megalin serves as a shuttle for signaling molecules from the cell surface to the mitochondria, including angiotensin II, TGF-ß and stanniocalcin-1.

Funding

  • Veterans Affairs Support