TFEB-mediated lysosomal exocytosis alleviates high-fat diet–induced lipotoxicity in the kidney
Jun Nakamura, Takeshi Yamamoto, Yoshitsugu Takabatake, Tomoko Namba-Hamano, Satoshi Minami, Atsushi Takahashi, Jun Matsuda, Shinsuke Sakai, Hiroaki Yonishi, Shihomi Maeda, Sho Matsui, Isao Matsui, Takayuki Hamano, Masatomo Takahashi, Maiko Goto, Yoshihiro Izumi, Takeshi Bamba, Miwa Sasai, Masahiro Yamamoto, Taiji Matsusaka, Fumio Niimura, Motoko Yanagita, Shuhei Nakamura, Tamotsu Yoshimori, Andrea Ballabio, Yoshitaka Isaka
Jun Nakamura, Takeshi Yamamoto, Yoshitsugu Takabatake, Tomoko Namba-Hamano, Satoshi Minami, Atsushi Takahashi, Jun Matsuda, Shinsuke Sakai, Hiroaki Yonishi, Shihomi Maeda, Sho Matsui, Isao Matsui, Takayuki Hamano, Masatomo Takahashi, Maiko Goto, Yoshihiro Izumi, Takeshi Bamba, Miwa Sasai, Masahiro Yamamoto, Taiji Matsusaka, Fumio Niimura, Motoko Yanagita, Shuhei Nakamura, Tamotsu Yoshimori, Andrea Ballabio, Yoshitaka Isaka
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Research Article Metabolism Nephrology

TFEB-mediated lysosomal exocytosis alleviates high-fat diet–induced lipotoxicity in the kidney

Abstract

Obesity is a major risk factor for end-stage kidney disease. We previously found that lysosomal dysfunction and impaired autophagic flux contribute to lipotoxicity in obesity-related kidney disease, in both humans and experimental animal models. However, the regulatory factors involved in countering renal lipotoxicity are largely unknown. Here, we found that palmitic acid strongly promoted dephosphorylation and nuclear translocation of transcription factor EB (TFEB) by inhibiting the mechanistic target of rapamycin kinase complex 1 pathway in a Rag GTPase–dependent manner, though these effects gradually diminished after extended treatment. We then investigated the role of TFEB in the pathogenesis of obesity-related kidney disease. Proximal tubular epithelial cell–specific (PTEC-specific) Tfeb-deficient mice fed a high-fat diet (HFD) exhibited greater phospholipid accumulation in enlarged lysosomes, which manifested as multilamellar bodies (MLBs). Activated TFEB mediated lysosomal exocytosis of phospholipids, which helped reduce MLB accumulation in PTECs. Furthermore, HFD-fed, PTEC-specific Tfeb-deficient mice showed autophagic stagnation and exacerbated injury upon renal ischemia/reperfusion. Finally, higher body mass index was associated with increased vacuolation and decreased nuclear TFEB in the proximal tubules of patients with chronic kidney disease. These results indicate a critical role of TFEB-mediated lysosomal exocytosis in counteracting renal lipotoxicity.

Authors

Jun Nakamura, Takeshi Yamamoto, Yoshitsugu Takabatake, Tomoko Namba-Hamano, Satoshi Minami, Atsushi Takahashi, Jun Matsuda, Shinsuke Sakai, Hiroaki Yonishi, Shihomi Maeda, Sho Matsui, Isao Matsui, Takayuki Hamano, Masatomo Takahashi, Maiko Goto, Yoshihiro Izumi, Takeshi Bamba, Miwa Sasai, Masahiro Yamamoto, Taiji Matsusaka, Fumio Niimura, Motoko Yanagita, Shuhei Nakamura, Tamotsu Yoshimori, Andrea Ballabio, Yoshitaka Isaka

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Figure 4

PA-induced TFEB activation promotes lysosomal exocytosis of phospholipids to prevent MLB accumulation in PTECs.

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PA-induced TFEB activation promotes lysosomal exocytosis of phospholipid...
(A–C) To investigate the role of TFEB on lysosomal exocytosis, wild-type and Tfeb-deficient PTECs were treated with BSA (0.25%, A, or 0.125%, B and C) or PA (0.25 mM, A, or 0.125 mM, B and C) for 6 hours. (A) Electron micrographs of wild-type PTECs (n = 2). (B) β-Hexosaminidase activity in the culture supernatant of PTECs relative to the total activity (n = 3). (C) Immunofluorescence images of nonpermeabilized PTECs show only LAMP1 that is exposed on the plasma membrane (n = 3). (D and E) To investigate the trafficking of phospholipids, a fluorescent fatty acid pulse-chase assay was performed. (D) Schematic illustration of pulse-chase assay. FL HPC–loaded wild-type and Tfeb-deficient PTECs were chased after treatment with either 0.125% BSA or 0.125 mM PA for 6 hours, and the subcellular localization of FL HPC was determined by staining with LysoTracker Red for 0, 12, or 24 hours after PA washout. (E) To measure phospholipid accumulation in lysosomes, the number of dots indicating staining for both phospholipids and lysosomes was counted (n = 3). Bars: 5 μm (A) and 10 μm (C and E). Data are provided as means ± SEM. Statistically significant differences: *P < 0.05 versus wild-type PTECs with the same treatment; #P < 0.05 versus BSA-treated PTECs (B, 1-way ANOVA followed by the Tukey-Kramer test; E, 2-tailed Student’s t test). All images are representative of multiple experiments. Ly, lysosome; WT, wild-type PTECs; KO, Tfeb-deficient PTECs; DIC, differential interference contrast.