Figure 4.
Transcription factor EB (Tfeb) upregulation and nuclear translocation following deletion of the polycystin-1 (Pkd1) gene. A−H: formalin-fixed paraffin-embedded renal sections derived from Pkd1fl/fl:Pax8-rTTA:TetO-Cre (Pkd1 iKO) mice that had been induced with doxycycline (DOX) at postnatal days 10 and 11 and isolated 1 day (A; DPD1), 3 days (B; DPD3), 4 days (C; DPD4), and 9 days (D; DPD9) post-DOX were subjected to immunostaining with a Tfeb antibody and compared with age-matched control (DOX-induced Pkd1fl/fl:TetO-Cre without Pax8-rTTA) animals (E–H). Nuclear translocation of Tfeb was observed in renal epithelial cells as early as DPD1 (A, black arrows), and the overall frequency of nuclear Tfeb translocation increased with age compared with control samples. Significant cytoplasmic punctate accumulation of Tfeb was evident at the DPD1 (A) and DPD3 (B) time points compared with control samples (E and F, respectively). To improve visual contrast, color images were converted to 8-bit grayscale, and a lookup table (ICA) was used to represent the relative intensity of the pixel data (I–P). Scale bars = 100 µm; scale bars in insets = 20 µm. Images are representative of five ×40 images acquired from three separate animals for each genotype. Q−T: Tfeb staining was performed on renal sections derived from Pkd1 iKO mice harvested at day 25 (Q) or day 150 (R). There was a pronounced upregulation and nuclear translocation in renal cystic epithelial cells from mice that were 2 or 20 wk postinduction. In contrast, age-matched control samples (S and T) showed very low levels of Tfeb expression in normal renal tubules as well as little evidence of nuclear translocation at these two time points. Scale bars = 100 µm; scale bars in insets = 20 µm. Images are representative of five ×40 images acquired from three separate animals for each genotype.