Abstract: TH-PO382
Identifying Molecular Regulators of Kidney Collecting Duct Cell Type Plasticity
Session Information
- Development, Organoids, Injury, and Regeneration
October 24, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Development, Stem Cells, and Regenerative Medicine
- 600 Development, Stem Cells, and Regenerative Medicine
Authors
- Simmons, Alicia L., Sanford Research, Sioux Falls, South Dakota, United States
- Mukherjee, Malini, Sanford Research, Sioux Falls, South Dakota, United States
- Isaiah, Oduduabasi, Sanford Research, Sioux Falls, South Dakota, United States
- Kareta, Michael S., Sanford Research, Sioux Falls, South Dakota, United States
- Surendran, Kameswaran, Sanford Research, Sioux Falls, South Dakota, United States
Group or Team Name
- Surendran Lab.
Background
Remodeling of kidney collecting duct segments by altering the cell type composition may underlie adaptive capacity when it occurs in a limited manner but may also be part of pathological states. Deficiencies in the machinery and number of principal cells (PCs) are associated with acquired Nephrogenic Diabetes Insipidus (NDI). Conversion of PCs to intercalated cells (ICs) results in loss of PCs when Notch signaling is blocked in adult mouse kidneys resulting in a NDI-like phenotype. Here we use rodent Notch-signaling deficient kidneys to identify the molecular mediators of PC to IC conversion.
Methods
We profiled the wild type versus littermate mice with inactivation of Hes1 in adult kidney epithelia for 7 days by total kidney RNA-sequencing and by single cell RNA-sequencing of samples enriched for epithelial cells of the loop of Henle, distal nephron and collecting duct segments. We used the mpkccdc14, a principal cell line, with ectopic Hes1-FLAG expression to identify the genes directly regulated by Hes1 by performing Hes1-FLAG-chromatin immunoprecipitation (ChIP) followed by sequencing and qPCR. The spatial pattern of the differentially expressed genes in Hes1 deficient kidneys versus the control wild type littermates were analyzed by RNAscope, and immunohistochemistry.
Results
Hes1-deficiency altered expression of signaling components of Hedgehog, mTOR, and Wnt pathways. Sub-clustering cells expressing PC and/or IC markers as determined by scRNA-seq revealed two PC clusters, two IC clusters and a fifth transitional cell (TC) cluster with both PC and IC marker expression. Comparison of TC versus PC cluster revealed higher expression of IC specific genes and down-regulation of PC specific genes. Some of the upregulated genes in the TCs, such as irs1, were identified as potential direct targets of Hes1 by Hes1-FLAG-ChIP-qPCR.
Conclusion
Hes1-deficient kidneys are useful to identify transcriptional signature and potential mediators of PC to IC conversion that occurs in adult kidneys. Considering that lithium treatment increased PC to IC conversion as determined by lineage tracing, identifying the molecular mediators of PC to IC conversion will allow us to test if these mechanisms are conserved in different forms of acquired of NDI, such as lithium induced or low K+ diets.
Funding
- NIDDK Support