Quantitative Proteomics of All 14 Renal Tubule Segments in Rat
Kavee Limbutara, Chung‐Lin Chou, Mark A. Knepper
Abstract
Significance Statement The renal tubule’s 14 distinct segments consist of epithelial cells with different transport and metabolic functions. Identifying the proteins mediating each function is crucial to gaining an overall understanding of kidney physiology and pathophysiology. New developments in protein mass spectrometry have resulted in a marked increase in sensitivity of protein detection and quantification. In this study, the authors manually microdissected kidney tubules from rat kidneys and leveraged the advances in protein mass spectrometry to identify and quantify the proteins expressed in each of the 14 tubule segments. They used these data to create an online information resource, the Kidney Tubule Expression Atlas, to allow researchers throughout the world to browse segment-specific protein expression data and download them for their own investigations. Background Previous research has used RNA sequencing in microdissected kidney tubules or single cells isolated from the kidney to profile gene expression in each type of kidney tubule epithelial cell. However, because proteins, not mRNA molecules, mediate most cellular functions, it is desirable to know the identity and amounts of each protein species to understand function. Recent improvements in the sensitivity of mass spectrometers offered us the ability to quantify the proteins expressed in each of 14 different renal tubule segments from rat. Methods We manually dissected kidney tubules from rat kidneys and subjected samples to protein mass spectrometry. We used the “proteomic ruler” technique to estimate the number of molecules of each protein per cell. Results Over the 44 samples analyzed, the average number of quantified proteins per segment was 4234, accounting for at least 99% of protein molecules in each cell. We have made the data publicly available online at the Kidney Tubule Expression Atlas website (https://esbl.nhlbi.nih.gov/KTEA/). Protein abundance along the renal tubule for many commonly studied water and solute transport proteins and metabolic enzymes matched expectations from prior localization studies, demonstrating the overall reliability of the data. The site features a “correlated protein” function, which we used to identify cell type–specific transcription factors expressed along the renal tubule. Conclusions We identified and quantified proteins expressed in each of the 14 segments of rat kidney tubules and used the proteomic data that we obtained to create an online information resource, the Kidney Tubule Expression Atlas. This resource will allow users throughout the world to browse segment-specific protein expression data and download them for their own research.