Abstract: TH-OR071
Single-Cell RNA-Seq Reveals Distribution of Na, K, and Cl Transporters among the Major Cell Types of the Collecting Duct
Session Information
- Na+, K+, Cl- Transport: Regulation and Molecular Mechanisms
November 02, 2017 | Location: Room 285, Morial Convention Center
Abstract Time: 04:30 PM - 04:42 PM
Category: Fluid, Electrolytes, and Acid-Base
- 703 Na+, K+, Cl- Basic
Authors
- Chen, Lihe, National Heart Lung and Blood Institute, Bethesda, Maryland, United States
- Chou, Chung-Lin, National Heart Lung and Blood Institute, Bethesda, Maryland, United States
- Burg, Maurice B., National Heart Lung and Blood Institute, Bethesda, Maryland, United States
- Verlander, Jill W., University of Florida, Gainesville, Florida, United States
- Wall, Susan M., Emory University School of Medicine, Atlanta, Georgia, United States
- Knepper, Mark A., National Heart Lung and Blood Institute, Bethesda, Maryland, United States
Background
Fine control of water, ion, and acid-base homeostasis is maintained by transport processes in the collecting duct. Collecting ducts are made up of at least 3 cell types: type A intercalated cells (A-ICs), type B intercalated cells (B-ICs) and principal cells (PCs). To identify transcriptomes for A-ICs, B-ICs and PCs, it is necessary to carry out RNA-Seq at a single-cell level.
Methods
We used cell-surface markers for A-ICs (c-Kit), B-ICs (PNA) and PCs (DBA), allowing these cell types to be enriched from kidney cell suspensions from untreated mice using fluorescence-activated cell sorting (FACS). The enriched fractions were used for microfluidic-based single-cell RNA-Seq. We performed a t-distributed stochastic neighbor embedding (t-SNE) analysis and resolved cell clusters. Based on the gene expression patterns, three major clusters are identified as A-ICs, B-ICs and PCs.
Results
In the table below, we present transcript abundances in TPM values for major Na, K and Cl transporters and associated regulators in the three major cell types of the collecting duct (largest value in bold). Our data confirm the selective expression of Hsd11b2, Scnn1b and Scnn1g, and major potassium channels (Kcnj1, Kcne1, Kcnj10, Kcnj16) in PCs. We also found AE1 (Slc4a1) and pendrin (Slc26a4) are mutually exclusive in A-IC and B-IC, respectively. Interestingly, we found that ENaC alpha (Scnn1a) and Na-K-ATPase (Atp1a1) transcripts are more abundant in B-ICs than in PCs or A-ICs. In addition, the mineralocorticoid receptor is predominantly expressed in B-ICs, while glucocorticoid receptor is expressed in all three cell types. AE4 (Slc4a9) was expressed in both IC cell types (confirmed by immunohistochemistry).
Conclusion
In conclusion, the single-cell RNA-Seq profiling results revealed a distinct distribution pattern of transporters and their regulators in A-ICs, B-ICs and PCs. The identification of transcripts related to Na+ transport and its regulation in B-ICs is consistent with a direct role of B-ICs in regulated Na reabsorption.
Funding
- Other NIH Support