Abstract: TH-OR006
Stress Response Gene NUPR1 Protects Renal Tubular Cells from Proliferation-Induced Energy Exhaustion
Session Information
- AKI: New Players and New Mechanisms
October 25, 2018 | Location: 6D, San Diego Convention Center
Abstract Time: 05:30 PM - 05:42 PM
Category: Acute Kidney Injury
- 103 AKI: Mechanisms
Authors
- Galichon, Pierre, Brigham and Women's Hospital/Harvard Medical School , Boston, Massachusetts, United States
- Li, Li, Brigham and Women's Hospital/Harvard Medical School , Boston, Massachusetts, United States
- Valerius, M. Todd, Brigham and Women's Hospital/Harvard Medical School , Boston, Massachusetts, United States
- Bonventre, Joseph V., Brigham and Women's Hospital/Harvard Medical School , Boston, Massachusetts, United States
Background
Energy depletion in renal tubular cells is a hallmark of acute kidney injury associated with cell death. NUPR1 (Nuclear protein 1, transcriptional regulator) promotes survival, increases ATP and inhibits proliferation in renal tubular cells. Although proliferation of tubular cells is necessary for kidney repair after resolution of injury, it could increase vulnerability in the context of ongoing injury. The effect of proliferation on energy in cells is unknown. We hypothesized that NUPR1 protects renal tubular cells from proliferation-induced energy exhaustion.
Methods
Stable renal proximal tubular cell lines were generated from HK-2 cells with inducible expression of NUPR1 (gain-of-function) or of CRISPR-dCas9-KRAB repression of NUPR1 (loss-of-function). The cells were transfected with the fluorescent biosensor PercevalHR to visualize the intracellular ATP/ADP ratio and with the intracellular pH sensor pHRed to adjust the ATP/ADP measures. The RatioPlus plugin from Fiji was used to measure ATP/ADP and pH from dual wavelength acquisitions of PercevalHR and pHRed, and the TrackMate plugin was used for monitoring single cell trajectories. The effect of tenovin-1 (p53 agonist for G1/S arrest), rigosertib (PLK1 inhibitor for G2/M arrest), pifithrin-α (p53 inhibitor for G1/S passage) and KU55933 (ATM kinase inhibitor for G2/M passage) on the ATP/ADP ratio was assessed.
Results
NUPR1 overexpression increased the ATP/ADP ratio in tubular cells. Conversely, NUPR1 inhibition decreased the ATP/ADP ratio. Modeling proliferative repair after injury by a scratch assay or facilitation of cell cycle passage by pifithrin-α or KU55933 decreased ATP/ADP ratio in migrating and proliferating cells. In contrast, inhibition of proliferation by either contact inhibition, tenovin-1 or rigosertib recapitulated NUPR1's protective effect on ATP/ADP levels. Proliferating cells undergo cyclic variations of the ATP/ADP ratio, with a mitotic peak followed by a sudden drop of ATP/ADP ratio and a gradual recovery.
Conclusion
NUPR1 protects renal tubular cells from energy depletion caused by proliferation. Proliferation is associated with decreased energy, characterized by cyclic steep diminution of ATP/ADP ratio after mitosis. Delaying proliferation is a potential therapeutic goal to minimize damage during acute kidney injury and prevent maladaptive repair after injury.
Funding
- NIDDK Support