EXOCYST INACTIVATION IN UROTHELIAL CELLS LEADS TO FN14 UPREGULATION AND CELL DEATH IN A MOUSE MODEL OF PRENATAL URETER OBSTRUCTIONS

Villiger, Ross

EXOCYST INACTIVATION IN UROTHELIAL CELLS LEADS TO FN14 UPREGULATION AND CELL DEATH IN A MOUSE MODEL OF PRENATAL URETER OBSTRUCTIONS

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Villiger_hawii_0085A_11377.pdf (5.17 MB)

Date

2022

Authors

Villiger, Ross

Advisor

Fogelgren, Ben

Department

Cell and Molecular Biology

Abstract

Congenital obstructive nephropathy (CON) is the leading cause of chronic kidney disease(CKD) and end-stage renal disease in children. This disorder is usually detected as hydronephrosis by prenatal ultrasounds. The most common cause of CON is obstruction at the ureteropelvic junction (UPJ), where the renal pelvis transitions to the upper ureter. EXOC5 is the central stabilizing subunit of the exocyst, an octameric protein complex that regulates exocytosis of secretory vesicles to the plasma membrane. The Fogelgren laboratory previously generated a novel EXOC5-conditional knockout (EXOC5-CKO) mouse using the Cre/loxP technology to inactivate EXOC5 in ureteric bud-derived epithelial cells. The EXOC5-CKO mice had severe bilateral UPJ obstructions, hydronephrosis, and succumbed to neonatal death. In these mice, UPJ obstruction was preceded by failure of urothelial stratification and cell death at E17.5, followed by a fibroproliferative response by the underlying mesenchyme. By transcriptomic microarray, we identified the Fn14 gene as being robustly upregulated in the EXOC5-CKO embryonic ureter at E16.5. Fn14 is a transmembrane tumor necrosis factor (TNF) receptor superfamily member with a single known ligand, TWEAK, which can activate different pathways depending on cell context, including programmed cell death. We demonstrated that ureter obstructions can be rescued in EXOC5-CKO mice by administering a pan-caspase inhibitor zVAD-fmk at E16.5. We also found inhibited urothelial stratification alone did not drive cell death during ureter development. Instead, exocyst-perturbed urothelial cells in vivo and in vitro have disrupted autophagy which activated NF-κB signaling and cell death. To study the role of Fn14 in renal and urological diseases, we generated a global Fn14-knockout (KO) mouse using CRISPR-Cas9 gene editing. Fn14-KO mice had ameliorated renal pathology in a cisplatininduced CKD model. Our findings support the hypothesis that urothelial cell death is a critical event leading to UPJO pathogenesis and that Fn14 signaling contributes to cell death in renal and urological pathologies.

Keywords

Cellular biology, Developmental biology, Molecular biology, exocyst, Fn14, ureter, urothelium

URI

https://hdl.handle.net/10125/102128

Extent

135 pages

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Ph.D. - Cell and Molecular Biology

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