Kidney Week

Abstract: FR-PO519

Distal Convoluted Tubule-Specific Disruption of the COP9 Signalosome and CUL3 Activates the WNK4-SPAK-NCC Pathway

Session Information

• Fluid, Electrolyte, Acid-Base Disorders: Basic
  November 03, 2023 | Location: Exhibit Hall, Pennsylvania Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Fluid, Electrolytes, and Acid-Base Disorders

• 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

• Cornelius, Ryan J., Oregon Health & Science University, Portland, Oregon, United States

Ryan J. Cornelius,

• Maeoka, Yujiro, Oregon Health & Science University, Portland, Oregon, United States

Yujiro Maeoka,

• Bradford, Tanner, Oregon Health & Science University, Portland, Oregon, United States

Tanner Bradford,

• Su, Xiao-Tong, Oregon Health & Science University, Portland, Oregon, United States

Xiao-Tong Su,

• Yang, Chao-Ling, Oregon Health & Science University, Portland, Oregon, United States

Chao-Ling Yang,

• Ellison, David H., Oregon Health & Science University, Portland, Oregon, United States

David H. Ellison,

• McCormick, James A., Oregon Health & Science University, Portland, Oregon, United States

James A. McCormick,

Background

The disease Familial Hyperkalemic Hypertension (FHHt; also known as Gordon Syndrome) is caused by aberrant accumulation of WNK4 activating the NaCl cotransporter (NCC) in the distal convoluted tubule (DCT) of the kidney. Mutations in cullin 3 (CUL3) cause FHHt by disrupting interaction with its regulator, the COP9 signalosome (CSN). Deletion of Cul3 and Jab1 (the catalytically active CSN subunit) along the entire nephron causes an FHHt phenotype with activation of the WNK4-SPAK-NCC pathway. However, only a partial FHHt phenotype develops due to kidney injury. We hypothesized that DCT-specific deletion would more accurately model the disease.

Methods

We utilized NCC-Cre-ERT2 mice to delete Cul3 (DCT-Cul3^-/-) and Jab1 (DCT-Jab1^-/-) only in the DCT. Blood was collected for plasma electrolyte analysis, and kidneys were collected for Western blotting and immunofluorescent staining.

Results

Both DCT-Cul3^-/- and DCT-Jab1^-/- mice showed an FHHt-like phenotype, with increased WNK4, SPAK, and pNCC abundance. CUL3 inactivation in the DCT-Cul3^-/- mice caused an increase in both KLHL3 (the CUL3 substrate adaptor for WNK4) and WNK4 abundance. WNK4 accumulated in DCT-Jab1^-/- mice due to hyperactivation of CUL3, which caused decreased CUL3 and KLHL3 abundance. Both genotypes showed formation of pSPAK and WNK4 puncta, however, the number of WNK4 puncta was higher in DCT-Cul3^-/- mice. Contrasting the disease, both genotypes showed no change in plasma K⁺, Cl^-, or HCO₃^- at baseline, but the DCT-Jab1^-/- mice became hyperkalemic when challenged with a high K⁺ diet. Additionally, over time DCT-Jab1^-/- mice showed a large decrease in NCC abundance, which was not observed in the DCT-Cul3^-/- mice. Interestingly, although Jab1 was deleted only in the DCT, the mice had increased abundance of KIM-1, a marker of proximal tubule injury.

Conclusion

DCT-specific knockout of both CUL3 and the cullin-RING-ligase regulator JAB1 led to an FHHt-like phenotype. Chronically, only DCT-Jab1^-/- mice developed a large reduction in NCC abundance, suggesting the decrease is due to a direct JAB1 effect, or effects of JAB1 on other cullins.

Funding

• NIDDK Support