Transcriptomes of major renal collecting duct cell types in mouse identified by single-cell RNA-seq

Abstract

Prior RNA sequencing (RNA-seq) studies have identified complete transcriptomes for most renal epithelial cell types. The exceptions are the cell types that make up the renal collecting duct, namely intercalated cells (ICs) and principal cells (PCs), which account for only a small fraction of the kidney mass, but play critical physiological roles in the regulation of blood pressure, extracellular fluid volume, and extracellular fluid composition. To enrich these cell types, we used FACS that employed well-established lectin cell surface markers for PCs and type B ICs, as well as a newly identified cell surface marker for type A ICs, c-Kit. Single-cell RNA-seq using the IC- and PC-enriched populations as input enabled identification of complete transcriptomes of A-ICs, B-ICs, and PCs. The data were used to create a freely accessible online gene-expression database for collecting duct cells. This database allowed identification of genes that are selectively expressed in each cell type, including cell-surface receptors, transcription factors, transporters, and secreted proteins. The analysis also identified a small fraction of hybrid cells expressing aquaporin-2 and anion exchanger 1 or pendrin transcripts. In many cases, mRNAs for receptors and their ligands were identified in different cells (e.g., chiefly in PCs vs. chiefly in ICs), suggesting signaling cross-talk among the three cell types. The identified patterns of gene expression among the three types of collecting duct cells provide a foundation for understanding physiological regulation and pathophysiology in the renal collecting duct.

Single-cell transcriptomics of the mouse kidney reveals potential cellular targets of kidney disease

“…Our work complements previous efforts that have applied this technology to the kidney. Single cell sequencing has been used to study fetal mouse kidneys and sorted kidney segments (7, 28, 29) and a recent study identified kidney cell composition changes in patients with lupus nephritis (30). …”

“…3G and fig. S17) (7, 25). These results indicate that the collecting duct contains not only PC and IC cells but a third distinct transitional cell type; this raises the possibility that IC and PC cells represent two ends of a spectrum of cellular phenotypes and that the two cell types may undergo cellular transitions.…”

“…Such analyses have already been applied to other organs (2, 5–7) and even to whole multicellular organisms (8, 9). These experiments have identified novel cells as well as catalogued marker genes for previously defined cells, indicating that this approach has the power to redefine kidney cell types.…”

Single Cell Profiling Reveals Sex, Lineage and Regional Diversity in the Mouse Kidney

“…For example, female mice and humans are more resistant to ischemia or ER-stress-invoked renal injury (Hodeify, 2013;Park et al, 2004;Aufhauser et al, 2016). Single nucleus and single cell RNA-sequencing (scRNA-seq) have provided new insight into the cellular make-up of mammalian organ systems including the kidney (Wu et al, 2019;Habib et al, 2017;Macosko et al, 2015;Adam et al, 2017;Chen et al, 2017;Park et al, 2018;Karaiskos et al, 2018;Han et al,2018). However, kidney studies have been largely male-centered and lack insight into higher order organization of cell types critical to kidney function.…”

A Multimodal and Integrated Approach to Interrogate Human Kidney Biopsies with Rigor and Reproducibility: The Kidney Precision Medicine Project

“…We followed the concept of building an iterative marker list derived from published data and data generated from the KPMP to qualify the identity and composition of the tissue being interrogated, validate and optimize tissue processing pipelines and identify regions or cell types for integrative quality check and analysis to build the kidney atlas. Our initial list (made in 2018) of a subset of cell type markers mainly relied on rodent studies and bulk RNAseq data with corresponding evidence from the human protein atlas (Supplemental File 1) (33)(34)(35)(36)(37)(38)(39)(40)(41)(42). Later iterations of the potential cell types/states were heavily dependent on the data generated from the human kidney specimens derived in part from the pilot project (see below).…”