Kidney Week

Abstract: SA-PO646

Characterization and Regulation of the Disease-Causing Mucin 1 Frameshift Protein in Patient-Derived Cells Affected by Autosomal Dominant Tubulointerstitial Kidney Disease (ADTKD)-MUC1

Session Information

• Genetic Kidney Diseases: Models, Mechanisms, and Therapies
  October 26, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Genetic Diseases of the Kidneys

• 1202 Genetic Diseases of the Kidneys: Non-Cystic

Authors

• Schwarz, Hannah, Universitatsklinikum Erlangen Medizinische Klinik 4 Nephrologie und Hypertensiologie, Erlangen, Bayern, Germany

Hannah Schwarz, MD

• Knaup, Karl Xaver, Universitatsklinikum Erlangen Medizinische Klinik 4 Nephrologie und Hypertensiologie, Erlangen, Bayern, Germany

Karl Xaver Knaup, PhD

• Wopperer, Florian Josef, Universitatsklinikum Erlangen Medizinische Klinik 4 Nephrologie und Hypertensiologie, Erlangen, Bayern, Germany

Florian Josef Wopperer, MD

• Schiffer, Mario, Universitatsklinikum Erlangen Medizinische Klinik 4 Nephrologie und Hypertensiologie, Erlangen, Bayern, Germany

Mario Schiffer, MD, DrMed, MBA

• Buettner, Maike Julia, Friedrich-Alexander-Universitat Erlangen-Nurnberg, Erlangen, Bayern, Germany

Maike Julia Buettner, MD

• Amann, Kerstin U., Friedrich-Alexander-Universitat Erlangen-Nurnberg, Erlangen, Bayern, Germany

Kerstin U. Amann, MD

• Wiesener, Michael Sean, Universitatsklinikum Erlangen Medizinische Klinik 4 Nephrologie und Hypertensiologie, Erlangen, Bayern, Germany

Michael Sean Wiesener, MD, DrMed

Background

Autosomal Dominant Tubulointerstitial Kidney Diseases (ADTKD) are rare monogenic diseases of the kidney, which are mainly caused by germline mutations of UMOD and MUC1, leading to end-stage renal disease (ESRD) usually in mid adulthood. ADTKD-MUC1 is caused by a characteristic frameshift mutation, leading to the de novo protein MUC1-fs. MUC1-fs is believed to play a distinct role in terms of a “toxic proteinopathy”, mainly accumulating in TMED9-containing vesicles (Dvela-Levitt et al., Cell 2019). The substance BRD4780 is reported to re-route MUC1-fs towards lysosomal degradation, with interventional trials being in preparation. Therefore, we aimed to gain more insights into MUC1-fs temporal and spacial regulative characteristics with respect to (pharmacological) intervention with TMED9.

Methods

To analyze MUC1-fs protein in more detail, we generated iTCs (immortalized Tubular Cells) from huPTC (human urinary Primary Tubular Cells) of several patients with ADTKD-MUC1. Clonal selection of cells was performed to retrieve immortalized clones with MUC1-fs expression.
From these cells functional studies including immunoblotting, immunofluorescence and electron microscopy (EM) were performed to analyze the temporal and special expression of MUC1-fs and TMED9 in more detail.

Results

Immunofluorescence studies confirmed that MUC1-fs accumulates in the cytoplasm, possibly the secretory pathway. Co-localization was only partially observed with TMED9 (which localizes to COP vesicles), being involved in protein trafficking within the early secretory pathway. TMED9 negative components of this pathway also showed MUC1-fs staining, such as the Golgi apparatus and Early Endosomes. Immunogold EM of iTCs reveals MUC1-fs expression within the ER and vesicular structures, compatible with the secretory pathway.
BRD4780 led to profound reduction of MUC1-fs. However, this effect was not specific. Protein half-life and reconstitution of MUC1-fs after blockade comprise a few hours.

Conclusion

Our data confirm and extend previously published information on intracellular MUC1-fs localization and regulation. The small molecule BRD4780 is effective, but not specific.

Funding

• Government Support – Non-U.S.