Kidney Week

Abstract: SA-PO0487

Urinary Proteins Uromodulin and Mucin-1 Physically Interact and Synergistically Stimulate the Magnesium Channel TRPM6

Session Information

• Fluid, Electrolyte, and Acid-Base Disorders: Basic Research
  November 08, 2025 | Location: Exhibit Hall, 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

• Wolf, Matthias Tilmann, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States

Matthias Tilmann Wolf, MD, PhD, FASN

• Zhang, Jing, The University of Texas Southwestern Medical Center, Dallas, Texas, United States

Jing Zhang, MD, PhD

• Hoenderop, Joost, Radboud Universiteit, Nijmegen, GE, Netherlands

Joost Hoenderop, PhD

• An, Sung Wan, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States

Sung Wan An, PhD

Background

Uromodulin (UMOD) and Mucin-1 (MUC1) share characteristics: 1) They are both membrane proteins which are secreted into human urine; 2) They create a protective layer on the surface of tubular cells by forming polymers; 3) Pathogenic variants in UMOD and MUC1 cause autosomal dominant tubulo-interstitial kidney disease; 4) We have previously shown that UMOD and MUC1 stimulate the magnesium channel TRPM6. Given these similarities, we investigated if UMOD and MUC1 physically interact and if both proteins stimulate TRPM6 synergistically.

Methods

We applied whole-cell patch-clamp recording to determine TRPM6 current density and the half maximal effective concentration (EC₅₀) of UMOD and MUC1 to stimulate TRPM6 per six-well. Physical interaction was evaluated by co-immunoprecipitation (Co-IP).

Results

1. We found that 0.9 µg of wild-type (WT) UMOD-HA and 0.2 µg of MUC1 plasmid resulted in half maximal stimulation (EC₅₀) of TRPM6. While individual “subthreshold” doses of 0.6 µg of WT UMOD-HA and 0.2 µg of MUC1 plasmid did not enhance TRPM6, the combination of “subthreshold” UMOD plus MUC1 dosages stimulated TRPM6 indicating a synergistic effect. 2. Co-IP studies showed that WT UMOD and MUC1 physically interact. 3. We determined that the UMOD truncation plasmid R586X UMOD-HA stimulated TRPM6 but neither the L180X UMOD-HA nor C297X UMOD-HA constructs activated TRPM6 indicating that TRPM6 stimulation requires either UMOD’s D8C and/or zona pellucida domain. 4. After determining the EC₅₀ for R586X UMOD-HA at 0.8 µg, neither the “subthreshold” dose for R586X UMOD-HA (0.6 µg) nor MUC1 (0.2 µg) alone enhanced TRPM6 while combined they stimulated the magnesium channel indicating that the glycosylphosphatidylinositol (GPI) anchor is not required for the UMOD-MUC1 interaction. 5. No synergistic action was found for C297X UMOD-HA and MUC1 regarding TRPM6 stimulation indicating that the UMOD zona pellucida domain is required for the synergistic action. 6. In Co-IP studies, we found physical interaction between MUC1 and the truncation mutant R586X UMOD-HA but not the truncation variant C297X UMOD-HA, suggesting that the missing zona pellucida domain is needed for UMOD to bind to MUC1.

Conclusion

The UMOD zona pellucida domain is required for physical interaction between UMOD and MUC1. Both proteins stimulate synergistically TRPM6.

Funding

• NIDDK Support

Digital Object Identifier (DOI)

• doi: 10.1681/ASN.2025h1t41s5f