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 November 1, 2019; First 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 6

Atr gene depletion in RPTECs results in increased profibrotic changes after IRI.

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Atr gene depletion in RPTECs results in increased profibrotic changes a...
(A) Representative images of ATR-stained sections of kidneys 48 hours after IRI or sham operation. Scale bar: 50 μm. (B) Real-time PCR analysis of Atr mRNA levels in ATRCtrl and ATRRPTC–/– kidneys on day 28 after IRI. ATRCtrl (n = 6), ATRRPTC–/– (n = 6). (C and D) Changes in serum creatinine and BUN following IRI. Sham operation: ATRCtrl (n = 3), ATRRPTC–/– (n = 3); IRI: ATRCtrl (n = 8), ATRRPTC–/– (n = 8). (E) Representative images of PAS-stained kidneys from ATRCtrl and ATRRPTC–/– mice on day 7 after IRI. Scale bars: 300 μm (top) and 50 μm (bottom). (F) Representative images of MT-stained kidneys from ATRCtrl and ATRRPTC–/– mice on day 28 after IRI or sham operation. Dot plot shows the quantification of the MT+ area. Scale bar: 100 μm. (G) Representative images of KSP- and α-SMA–stained kidney sections from ATRCtrl and ATRRPTC–/– mice 28 days after IRI or sham operation. Dot plots show the quantification of KSP+ and α-SMA+ areas. Scale bar: 50 μm. (F and G) Sham: ATRCtrl (n = 3 mice, 6 kidneys), ATRRPTC–/– (n = 3 mice, 6 kidneys); IRI: ATRCtrl (n = 8 mice, 16 kidneys), ATRRPTC–/– (n = 8 mice, 16 kidneys). (H–K) RT-PCR analysis of TGF-β, α-SMA, Col1a1, and p21 mRNA levels in ATRCtrl and ATRRPTC–/– kidneys on day 28 following IRI. Sham operation: ATRCtrl (n = 3), ATRRPTC–/– (n = 3); IRI: ATRCtrl (n = 8), ATRRPTC–/– (n = 8). Data are presented as the mean ± SEM. Statistical significance was determined by 2-tailed, unpaired t test (IRI ATRCtrl vs. IRI ATRRPTC–/–). *P < 0.05 and **P < 0.01.