Proximal tubule ATR regulates DNA repair to prevent maladaptive renal injury responses
Seiji Kishi, … , Ryuji Morizane, Joseph V. Bonventre
Seiji Kishi, … , Ryuji Morizane, Joseph V. Bonventre
Published October 7, 2019
Citation Information: J Clin Invest. 2019;129(11):4797-4816. https://doi.org/10.1172/JCI122313.
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Research Article Nephrology

Proximal tubule ATR regulates DNA repair to prevent maladaptive renal injury responses

Abstract

Maladaptive proximal tubule (PT) repair has been implicated in kidney fibrosis through induction of cell-cycle arrest at G2/M. We explored the relative importance of the PT DNA damage response (DDR) in kidney fibrosis by genetically inactivating ataxia telangiectasia and Rad3-related (ATR), which is a sensor and upstream initiator of the DDR. In human chronic kidney disease, ATR expression inversely correlates with DNA damage. ATR was upregulated in approximately 70% of Lotus tetragonolobus lectin–positive (LTL+) PT cells in cisplatin-exposed human kidney organoids. Inhibition of ATR resulted in greater PT cell injury in organoids and cultured PT cells. PT-specific Atr-knockout (ATRRPTC–/–) mice exhibited greater kidney function impairment, DNA damage, and fibrosis than did WT mice in response to kidney injury induced by either cisplatin, bilateral ischemia-reperfusion, or unilateral ureteral obstruction. ATRRPTC–/– mice had more cells in the G2/M phase after injury than did WT mice after similar treatments. In conclusion, PT ATR activation is a key component of the DDR, which confers a protective effect mitigating the maladaptive repair and consequent fibrosis that follow kidney injury.

Authors

Seiji Kishi, Craig R. Brooks, Kensei Taguchi, Takaharu Ichimura, Yutaro Mori, Akinwande Akinfolarin, Navin Gupta, Pierre Galichon, Bertha C. Elias, Tomohisa Suzuki, Qian Wang, Leslie Gewin, Ryuji Morizane, Joseph V. Bonventre

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Figure 8

Deletion of RPTEC Atr results in more severe kidney injury and increased DNA damage and cleaved caspase 3 after UUO.

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Deletion of RPTEC Atr results in more severe kidney injury and increased...
(A) Representative kidney histological images of PAS-stained kidney sections 7 days after UUO. Scale bar: 50 μm. Dot plot shows quantified tubular injury score. ATRCtrl (n = 4), ATRRPTC–/– (n = 5). (B) Representative images of KIM-1–stained kidneys from ATRCtrl and ATRRPTC–/– mice 7 days after UUO. Scale bar: 50 μm. (C) Western blotting was performed on UUO-injured kidney lysates to determine KIM-1 expression. Each lane represents 1 sample from an individual mouse. Dot plot shows the corresponding quantification of band intensity. ATRCtrl (n = 3), ATRRPTC–/– (n = 5). (D) Representative images of γH2AX-stained sections of CLKs and injured kidneys from ATRCtrl and ATRRPTC–/– mice 7 days after UUO. Dot plots show the corresponding quantification of γH2AX+ cells and of nucleus-wide γH2AX+ and γH2AX+ foci. ATRCtrl (n = 4), ATRRPTC–/– (n = 5). Scale bars: 50 μm. (E) Representative images of cleaved caspase 3–stained sections of CLKs and injured kidneys from ATRCtrl and ATRRPTC–/– mice 7 days after UUO. Scale bar: 50 μm. Dot plot shows the corresponding quantification of cleaved caspase 3+ cells. ATRCtrl (n = 4), ATRRPTC–/– (n = 5). Data are presented as the mean ± SEM. Statistical significance was determined by 2-tailed, unpaired t test (UUO ATRCtrl vs. ATRRPTC–/–). *P < 0.05 and **P < 0.01. The complete unedited blots are provided in the supplemental material.