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Abstract: PO0718

mTORC2 Is Essential for Sodium-Glucose Cotransporter 2

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Shabbir, Waheed, University of California San Francisco, San Francisco, California, United States
  • Demko, John E., University of California San Francisco, San Francisco, California, United States
  • Takagi, Enzo, University of California San Francisco, San Francisco, California, United States
  • Saha, Bidisha, University of California San Francisco, San Francisco, California, United States
  • Leite-Dellova, Deise C A, University of California San Francisco, San Francisco, California, United States
  • Sambandam, Bharathi, University of California San Francisco, San Francisco, California, United States
  • Pearce, David, University of California San Francisco, San Francisco, California, United States

Group or Team Name

  • David Pearce group
Background

The role of mammalian target of rapamycin (mTOR) complexes mTORC1 and mTORC2 in renal tubule ion transport has been well characterized. We and others have shown that mTORC2 is a key regulatory kinase for serum and glucocorticoid kinase 1 (SGK1) and that its activity is required for epithelial Na+ channel (ENaC)-dependent sodium reabsorption in the aldosterone-sensitive distal nephron (ASDN). Also, it has been shown that mTORC1 activity is increased in renal proximal tubule cells (RPTCs) in diabetes, which was prevented by the inhibition of sodium-glucose co-transporter 2 (SGLT2), and that mTORC1 KO in mice causes a Fanconi’s syndrome-like phenotype. However, the roles of mTORC2 in the regulation of RPTC transporters, particularly as it pertains to glucose reabsorption remain obscure. In this study we explored the relationship between mTORC2 and SGLT2 in CRISPR-modified HEK-293Tcells and in mice, using patch clamp and membrane expression studies

Methods

We used CRISPR-Cas9 to generate Sin1 (an essential component of mTORC2)-deficient HEK-293T cells, which were compared with wild-type cells. The cells were transiently transfected with SGLT2. We recorded in WT HEK-293T cells the Dapa-sensitive SGLT2 sodium current. We used an inducible Cre-Lox system (Pax8-LC1) to KO Rictor (another key component of mTORC2) in mice. Dapagliflozin-sensitive whole-cell SGLT2 sodium current was measured in the microdissected proximal tubules and HEK-293T cells

Results

Strikingly, in mTORC2-knockout HEK-293T cells the Dapa-sensitive SGLT2 sodium current was significantly reduced versus WT HEK-293T cells. In mice, mTORC2 KO caused glycosuria without hyperglycemia, and patch-clamp studies showed decreased glucose-induced, dapagliflozin-inhibited Na+ current.

Conclusion

Knockout of mTORC2 in the HEK-293T cells or in mice inhibits SGLT2-sodium current. Our study delineates the essential role of mTORC2 in SGLT2 function. These observations may explain the broad role of SGLT2 inhibition therapy and variable resistance to their effects.

Funding

  • NIDDK Support