ZEB2's Role in Renin-Expressing Cell Development and Hypotension

Heterozygous mutations or deletions in the ZEB2 gene cause Mowat-Wilson syndrome (MWS, OMIM #235730), a congenital disorder with multiple defects, including renal anomalies. Some MWS patients reported having hypotension. ZEB2 has been reported as a super-enhancer for the renin (REN) gene, and renin-null cells have upregulated ZEB2 expression. However, the role of ZEB2 in renin cell biology and the cellular and molecular mechanisms underlying the hypotension phenotype in MWS remain unclear. Renin precursor cells derive from FOXD1-positive kidney stromal progenitors. ZEB2 is highly expressed in FOXD1-positive kidney stromal progenitors and regulates their differentiation. We hypothesize that ZEB2 is required for the differentiation of FOXD1-positive kidney stromal progenitors into renin + cells. We deleted the Zeb2 gene in the kidney stroma using Foxd1Cre and analyzed the Zeb2 flox/flox ; Foxd1Cre + conditional knockout mice ( Zeb2 cKO) and their wild-type (WT) littermate controls for the renin cell number and arterial blood pressure. Single-cell RNA sequencing (scRNA-seq) was performed using the 10x Genomics Chromium platform on kidney tissues from 3-week-old Zeb2 cKO and WT mice. The data were analyzed using our in-house computational pipeline. Immunostaining was performed to validate scRNA-seq findings. We found that 4-week-old Zeb2 cKO mice had reduced renin-expressing cells and lower arterial blood pressure compared to WT controls. Our clustering analysis identified nine major cell types in both groups. The scRNA-seq and gene expression analysis revealed a significant increase in the proportion of podocytes and parietal epithelial cells (PECs) and ectopic expression of podocyte and PEC markers (WT1, TLE4, and CLDN1) within the vascular compartment (also expressing renin) of Zeb2 cKO kidneys. Trajectory and functional enrichment analyses identified significant alterations in angiogenesis and chromatin remodeling pathways in Zeb2 cKO mouse kidneys. In summary, ZEB2 is required for the differentiation of FOXD1-positive stromal progenitors in the kidney. In the absence of ZEB2, FOXD1-positive stromal progenitors adopt a podocyte—and PEC-like cell fate instead of renin-expressing cells, leading to a hypotension phenotype. Our results suggest that ZEB2 is a critical determinant of renin + cell lineage development.