RESUMEN
mTOR kinase is a master growth regulator that can be stimulated by multiple signals, including amino acids and the lysosomal small GTPase Rheb. Recent studies have proposed an important role for the V-ATPase in the sensing of amino acids in the lysosomal lumen. Using the Drosophila wing as a model epithelium, we show here that the V-ATPase is required for Rheb-dependent epithelial growth. We further uncover a positive feedback loop for the control of apical protein uptake that depends on V-ATPase/mTOR signaling. This feedback loop includes Rheb-dependent transcriptional regulation of the multiligand receptor Megalin, which itself is required for Rheb-induced endocytosis. In addition, we provide evidence that long-term mTOR inhibition with rapamycin in mice causes reduction of Megalin levels and proteinuria in the proximal tubular epithelium of the kidney. Thus, our findings unravel a homeostatic mechanism that allows epithelial cells to promote protein uptake under normal conditions and to prevent uptake in lysosomal stress conditions.
Asunto(s)
Endocitosis , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismo , Animales , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Epitelio/crecimiento & desarrollo , Epitelio/metabolismo , Retroalimentación Fisiológica , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas de Unión al GTP Monoméricas/genética , Proteínas de Unión al GTP Monoméricas/metabolismo , Neuropéptidos/genética , Neuropéptidos/metabolismo , Proteinuria/metabolismo , Proteína Homóloga de Ras Enriquecida en el Cerebro , Transducción de Señal , Serina-Treonina Quinasas TOR/genética , ATPasas de Translocación de Protón Vacuolares/genéticaRESUMEN
The C subunit of the vacuolar H(+)-ATPase or V-ATPase regulates the activity and assembly of the proton pump at cellular membranes. It has been shown to be strongly upregulated in oral squamous cell carcinoma, a highly metastatic epithelial cancer. In addition, increased V-ATPase activity appears to correlate with invasiveness of cancer cells, but the underlying mechanism is largely unknown. Using the Drosophila wing imaginal epithelium as an in vivo model system, we demonstrate that overexpression of Vha44, the Drosophila orthologue of the C subunit, causes a tumor-like tissue transformation in cells of the wing epithelium. Overexpressing cells are excluded from the epithelium and acquire invasive properties while displaying high apoptotic rates. Blocking apoptosis in these cells unmasks a strong proliferation stimulus, leading to overgrowth. Furthermore, we show that excess Vha44 greatly increases acidification of endocytic compartments and interferes with endosomal trafficking. As a result, cargoes such as GFP-Lamp1 and Notch accumulate in highly acidified enlarged endolysosomal compartments. Consistent with previous reports on the endocytic activation of Eiger/JNK signaling, we find that V-ATPase stimulation by Vha44 causes JNK signaling activation whereas downmodulation of JNK signaling rescues the invasive phenotypes. In summary, our in vivo-findings demonstrate that increased levels of V-ATPase C subunit induce a Eiger/JNK-dependent cell transformation within an epithelial organ that recapitulates early carcinoma stages.