TFEB drives mTORC1 hyperactivation and kidney disease in Tuberous Sclerosis Complex
Tuberous Sclerosis Complex (TSC) is caused by germline TSC1 or TSC2 mutations, leading to hyperactivation of mechanistic target of rapamycin complex 1 (mTORC1) and tumors in multiple organs including the brain, heart, lung (lymphangioleiomyomatosis), and kidney (angiomyolipoma and renal cell carcinoma). Previously, we found that TFEB is constitutively active in models of TSC. To determine the impact of TFEB in vivo, we generated two novel mouse models of TSC, resulting in premature death, in which kidney pathology was the primary phenotype. RNA sequencing revealed that lysosomal and proteasomal gene pathways were the most highly upregulated in the TSC2-deficient kidneys. Knockout of TFEB rescued both kidney pathology and overall survival in both models, indicating that TFEB is the primary driver of renal disease in TSC. Importantly, mTORC1 activity, which was elevated in the TSC2 knockout kidneys, was normalized by TFEB knockout.  Knockdown of Rheb or treatment of TSC2-deficient cells with Rapamycin paradoxically increases TFEB phosphorylation at the mTORC1-site (S211) and relocalizes TFEB from the nucleus to the cytoplasm via a Rag-dependent mechanism. Accordingly, treatment of TSC2 knockout mice with Rapamycin normalized lysosomal gene expression, similar to TFEB knockout, suggesting that the beneficial effects of Rapamycin in TSC are TFEB-dependent. These results change the view of the mechanisms leading to mTORC1 hyperactivation in TSC and may lead to novel therapeutic avenues for the treatment of TSC. Overall design: To investigate the effects of Tfeb inactivation in the setting of Tsc2 deficiency in vivo we created two distinct mouse models: whole-body tamoxifen inducible CaggCre model and kidney-specific KSPCre, both in BL/6 background. Main comparison groups in both models included control mice (group 1), mice with inactivation of Tsc2 (group 2), mice with inactivation of Tfeb (group 3) and mice with inactivation of both Tsc2 and Tfeb (group 4). Gene expression changes were analyzed using RNA-sequencing in three biological replicates per experimental group in both models. Kidneys from CaggCre groups were harvested at P30; kidneys from KSPCre groups were harvested at P50. In addition, gene expression changes induced by pharmacologic inhibition of mTOR versus Tfeb inactivation were assesed in KSPCre model using 6 separate groups and 3 biological replicates per group: control mice treated with vehicle control (group 1); control mice treated with Rapamycin (1mg/kg/3 times per week, starting at P25 for a total of 11 injections) (group 2); mice with inactivation of Tsc2 treated with Vehicle control (group 3); mice with inactivation of Tsc2 treated with Rapamycin as in group 2 (group 4); mice with inactivation of both Tsc2 and Tfeb treated with Vehicle control (group 5); mice with inactivation of both Tsc2 and Tfeb treated with Rapamycin as in group 2 (group 6).