Fig 2: Summary of observed transcription factor EB (Tfeb)-related differences between normal and cystic renal epithelial cells. In normal renal epithelial cells (left), Tfeb is associated with the lysosomal compartment in the cytoplasm as a component of a large multiprotein complex (left, inset). In cystic epithelial cells (right) that develop due to the absence of polycystin-1, folliculin (Flcn), or the folliculin-interacting protein 1 and 2 (Fnip1/2) complex, there is a pronounced nuclear accumulation of Tfeb leading to the transcription of Tfeb target genes, such as glycoprotein nonmetastatic melanoma protein B (GPNMB) and cathepsin D (CTSD). In cells that lack polycystin-1 (Pkd1) expression, cytoplasmic light chain 3B (Lc3b) puncta increase in number, indicative of autophagy induction, and p62 puncta are decreased, indicative of increased degradation and supported by an observed increase in autophagic flux. Consistent with a role of the Tfeb pathway in lysosomal biogenesis and positioning, lysosomes increase in size and undergo perinuclear repositioning in the absence of Pkd1. In human ADPKD samples, upregulation and nuclear translocation of TFEB in renal cysts is associated with a concomitant increase in GPNMB and CTSD expression. This figure was created with BioRender.com.

Fig 3: Transcription factor EB (Tfeb) upregulation and nuclear translocation in renal epithelial cells in the folliculin (Flcn) knockout (KO) and folliculin-interacting protein 1 and 2 (Fnip1/2) double KO (dKO) models of cystic disease. Immunohistochemistry was performed on serial renal formalin-fixed paraffin-embedded sections derived from Flcnfl/fl:Ksp-Cre (Flcn KO; A–C), Fnipi1fl/fl:Fnip2fl/fl:Ksp-Cre (Fnip1/2 dKO; D-F) and aged-matched control (Flcnfl/+:Fnip1fl/+:Fnip2fl/+:Ksp-Cre; G–I) animals using antibodies raised against Tfeb (A, D, and G), glycoprotein nonmetastatic melanoma protein B (Gpnmb; B, E, and H), or cathepsin B (C, F, and I). A striking upregulation and nuclear translocation of Tfeb (A and D) were observed in renal cystic epithelial cells in kidneys from Flcn KO (A) and Fnip1/2 dKO (D) animals compared with the low-level cytoplasmic staining of Tfeb observed in a subset of renal tubules from control kidneys (G). This nuclear translocation of Tfeb in renal cystic epithelial cells corresponded with a significant upregulation of Gpnmb (B and E), a known downstream target of Tfeb, which exhibited strong apical membrane staining compared with control renal tubules (H). A similar staining pattern was observed for the lysosomal-specific enzyme cathepsin B, another Tfeb target, which was significantly upregulated (C and F) and exhibited a strong apical localization pattern compared with the low expression evident in control renal tubules (I). Scale bars = 100 µm; scale bars in insets = 20 µm. Images are representative of five ×10 images acquired from three separate animals for each genotype. P18, postnatal day 18.