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.
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•The V-ATPase is required for mTOR-dependent tissue growth in Drosophila•V-ATPase/mTOR signaling controls apical endocytosis•Megalin activity and apical surface area is regulated by V-ATPase/mTOR signaling•Long-term mTOR inhibition causes Megalin downregulation and proteinuria in the kidney
Epithelial growth requires the endocytic uptake of proteins, which are broken down into amino acids. Amino acids recruit the growth-regulating mTORC1 complex to the lysosomal surface, thereby causing its activation. Gleixner et al. now find that in epithelial cells apical protein uptake is controlled by the lysosomal V-ATPase/mTORC1 complex in a feedforward manner. The multiligand receptor Megalin, which internalizes proteins on the apical surface, is crucial for this regulatory pathway.
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.
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Advanced systemic mastocytosis (SM) is an aggressive hematopoietic neoplasm with poor prognosis and short survival times. So far, no curative therapy is available for affected patients. We have ...identified the cell surface antigen CD52 (CAMPATH‐1) as a molecular target expressed abundantly on the surface of primary neoplastic mast cells (MCs) in patients with advanced SM. In contrast, neoplastic MCs of patients with indolent SM and normal MCs expressed only low levels or did not express CD52. To study the mechanisms of CD52 expression and the value of this antigen as a potential therapeutic target, we generated a human MC cell line, designated MCPV‐1, by lentiviral immortalization of cord blood‐derived MC progenitor cells. Functional studies revealed that activated RAS profoundly promotes surface expression of CD52. The CD52‐targeting antibody alemtuzumab induced cell death in CD52+ primary neoplastic MCs obtained from patients with SM as well as in MCPV‐1 cells. NSG mice xenotransplanted with MCPV‐1 cells survived significantly longer after treatment with alemtuzumab (median survival: 31 d untreated vs. 46 d treated; P=0.0012). We conclude that CD52 is a novel marker and potential therapeutic target in neoplastic MCs in patients with advanced SM.—Hoermann, G., Blatt, K., Greiner, G., Putz, E. M., Berger, A., Herrmann, H., Cerny‐Reiterer, S., Gleixner, K. V., Walz, C., Hoetzenecker, K., Müllauer, L., Reiter, A., Sotlar, K., Sexl, V., Valent, P., Mayerhofer, M. CD52 is a molecular target in advanced systemic mastocytosis. FASEB J. 28, 3540–3551 (2014). www.fasebj.org
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