Disruption of Robo2-Baiap2 integrated signaling drives cystic disease
Qinggang Li, Shaoyuan Cui, Qian Ma, Ying Liu, Hongyu Yu, GuangRui Geng, Ewud Agborbesong, Chongyu Ren, Kai Wei, Yingjie Zhang, Jurong Yang, Xueyuan Bai, Guangyan Cai, Yuansheng Xie, Xiaogang Li, Xiangmei Chen
Qinggang Li, Shaoyuan Cui, Qian Ma, Ying Liu, Hongyu Yu, GuangRui Geng, Ewud Agborbesong, Chongyu Ren, Kai Wei, Yingjie Zhang, Jurong Yang, Xueyuan Bai, Guangyan Cai, Yuansheng Xie, Xiaogang Li, Xiangmei Chen
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Research Article Development Nephrology

Disruption of Robo2-Baiap2 integrated signaling drives cystic disease

Abstract

Hereditary renal cystic diseases are characterized by defects in primary cilia of renal tubular epithelial cells and abnormality of tubular epithelium, which ultimately result in the development of renal cysts. However, the mechanism leading from abnormality of the tubular epithelium to cystogenesis is not well understood. In this report, we demonstrate a critical role for Robo2 in regulating epithelial development, including ciliogenesis, polarization, and differentiation. We found that Robo2 deficiency results in cystic kidneys, and the cyst cells showed defective cilia and polarity defects in tubular epithelium. The cyst cells, less than terminally differentiated, continue to proliferate. We further established that Robo2 works with p53 as well as polarity and ciliary proteins (Par3, PKCς, ZO-2, and Claudin-2) to regulate these processes. Robo2 binds to Baiap2 (also known as IRSp53) through the IRSp53/MIM homology domain in renal epithelial cells. This binding allows Robo2 to phosphorylate MDM2 at Ser166 via Baiap2 and maintain p53 homeostasis. Disruption of the Robo2-Baiap2 complex causes MDM2 to be subjected to dephosphorylation, leading to a high level of active p53, and initiated p53-mediated cellular senescence via p21 and decreased the expression of ZO-1, ZO-2, PKCς, Par3, and Claudin-2 proteins, resulting in defects in epithelial development, including ciliogenesis, polarization, and differentiation. Importantly, double knockout of Robo2 and p53 rescued all the epithelial defects in kidneys compared with those in Robo2-knockout kidneys. Taken together, the present results demonstrate that Robo2 deficiency causes renal cystic disease, which is largely dependent on defective Robo2-Baiap2 integrated signaling in kidneys.

Authors

Qinggang Li, Shaoyuan Cui, Qian Ma, Ying Liu, Hongyu Yu, GuangRui Geng, Ewud Agborbesong, Chongyu Ren, Kai Wei, Yingjie Zhang, Jurong Yang, Xueyuan Bai, Guangyan Cai, Yuansheng Xie, Xiaogang Li, Xiangmei Chen

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Figure 1

Robo2 is required for progenitor cells to differentiate into ciliated and polarized epithelium.

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Robo2 is required for progenitor cells to differentiate into ciliated an...
(A) Representative H&E-stained image showing renal cysts (*) in the Robo2–/– mice. Scale bars: 50 μm. (B) Immunofluorescence (IF) staining of Robo2 and Pax8 in renal tubular epithelial cells in E15.5 wild-type kidneys (n = 5). Scale bars: 50 μm. (C) IF staining of Pax8 and Lotus tetragonolobus lectin (LTL) in renal tubular epithelial cells in E17 wild-type and Robo2-knockout kidneys. Pax8 was expressed in cystic segments with reduced or absent LTL staining (white arrow and arrowhead). The representative image shows cysts (*) in the kidney. Scale bars: 50 μm. (D) IF staining of Aqp1 and LTL in the cryosections of E17 wild-type and Robo2–/– kidneys. The misexpression of LTL and Aqp1 was found in the Robo2–/– kidneys. Scale bars: 10 μm. (E) IF staining of LTL and Na+/K+-ATPase in the Robo2–/– kidneys showed a lack of polarity in the renal tubule epithelial cells. Scale bars: 10 μm. Biological replicates: n = 10. (F) IF staining of α-tubulin and LTL in the cryosections of kidneys showed the defects of ciliogenesis in the Robo2–/– kidneys (n = 7). Scale bars: 10 μm. (G) Transmission electron micrographs show that primary cilia (white arrow) defects (white arrowhead) occurred in the Robo2–/– kidney. Scale bars: 200 μm. (H) The tight junction (white arrowhead) is disrupted in the Robo2–/– kidney. Scale bars: 500 μm.