ASN's Mission

ASN leads the fight to prevent, treat, and cure kidney diseases throughout the world by educating health professionals and scientists, advancing research and innovation, communicating new knowledge, and advocating for the highest quality care for patients.

learn more

Contact ASN

1401 H St, NW, Ste 900, Washington, DC 20005

email@asn-online.org

202-640-4660

The Latest on Twitter

Kidney Week

Abstract: FR-PO922

Role of IRE1α in Podocyte Proteostasis

Session Information

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Navarro-Betancourt, José R., McGill University, Montreal, Quebec, Canada
  • Papillon, Joan, McGill University, Montreal, Quebec, Canada
  • Guillemette, Julie, McGill University, Montreal, Quebec, Canada
  • Iwawaki, Takao, Kanazawa Medical University, Ishikawa, Japan
  • Cybulsky, Andrey V., McGill University, Montreal, Quebec, Canada
Background

Podocyte (glomerular epithelial cell; GEC) proteostasis is disrupted in glomerular diseases. To maintain proteostasis, the endoplasmic reticulum (ER) orchestrates the unfolded protein response (UPR), which includes upregulation of chaperones. Proteostasis also involves clearance of misfolded proteins via autophagy. Inositol requiring enzyme-1α (IRE1α) resides in the ER membrane and is a transducer of the UPR. In mice, podocyte-specific deletion of IRE1α leads to age-related podocyte injury, autophagy impairment, and disruption of glomerular permselectivity. This study characterizes mechanisms by which IRE1α regulates proteostasis in GECs.

Methods

GECs were isolated from transgenic mice with loxP sites flanking the ribonuclease domain of the IRE1α gene. IRE1α was deleted by transduction of Cre recombinase (IRE1α KO). GECs expressing full-length IRE1α served as control (IRE1α WT). GECs were exposed to tunicamycin (TM), rapamycin (R), and glutamine starvation (GS) during 24 h. ER chaperones and LC3 were monitored by immunoblotting. Mitochondria were visualized using MitoTracker Red CMXRos.

Results

IRE1α KO and WT GECs exhibited comparable proliferation rates and protein content, implying that IRE1α is not involved in cell cycle progression. Stimulation of GECs with the ER stressor TM upregulated total IRE1α in WT, but not IRE1α KO GECs. After TM treatment, the chaperones BiP, GRP94, and mesencephalic astrocyte-derived neurotrophic factor increased in WT GECs. Deletion of IRE1α, or chemical inhibition of the IRE1α RNase with 4μ8C, significantly attenuated upregulation of chaperones and enhanced ER stress-induced apoptosis (evidenced by caspase-3 cleavage). Neither R nor GS enhanced expression of ER chaperones. Compared with WT GECs, IRE1α KO and 4μ8C-treated WT GECs showed similar basal autophagy, but reduced LC3 lipidation after stimulation with TM, R, and GS, indicating deficient autophagosome formation. Under basal conditions and after exposure to TM, the percentage of cell area occupied by active mitochondria in IRE1α KO and 4μ8C-treated GECs was significantly lower than in WT GECs, suggesting collapse of the mitochondrial network.

Conclusion

Stress-induced chaperone production, autophagy and mitochondrial distribution are compromised by deletion of IRE1α. The IRE1α pathway may play an important protective role in glomerular diseases associated with podocyte ER stress.

Funding

  • Government Support - Non-U.S.