Transcriptional and Post-Transcriptional Regulation of Genes Critical for Sodium Reabsorption and Blood Pressure Control in the Kidney

Abstract

Serine/threonine kinases (WNK1, WNK4, SPAK, OSR1) and cation co-transporters (NKCC2, NCC) are members of a multi-kinase network that determines renal Na+ reabsorption and blood pressure (BP) regulation. The importance of these proteins is highlighted by their roles in monogenic forms of hyper- and hypotension, animal models, and by the efficacy of BP-lowering medications that target this pathway. While post-translational regulation of these proteins has been well established, regulation of these genes at the transcript level is not completely understood. In this study, we examined both human and mouse kidney transcriptomes to uncover novel transcriptional and post-transcriptional regulation of two genes in this pathway. First, STK39 encodes for Ste20-related proline alanine rich kinase (SPAK), which phosphorylates and activates cation co-transporters. Variants within STK39 are associated with susceptibility to essential hypertension, and SPAK null mice are hypotensive and mimic Gitelman syndrome, a rare monogenic salt-wasting human disorder. Mice exhibit nephron segment-specific expression of full length SPAK and N-terminally truncated SPAK isoforms with impaired kinase function. We established that while humans also express transcript isoforms similar to those found in mice, they differ in abundance and are transcribed from human-specific promoters. Second, SLC12A3 encodes for the thiazide-sensitive Na+-Cl- co-transporter (NCC), and rare mutations in this gene cause Gitelman syndrome. In humans and mice, alternative polyadenylation of NCC pre-mRNA results in a longer 3'UTR isoform, while alternative splicing within the final exon leads to an exon-exon junction downstream of the termination codon. Both of these events generate potential substrates for nonsense-mediated mRNA decay (NMD). By suppressing NMD, we demonstrated that NCC transcript abundance is partially determined by post-transcriptional processing of its final exon. Finally, dietary K+ manipulation differentially alters SPAK and NCC transcript and protein isoform abundance, demonstrating dynamic physiological regulation of gene expression in response to salt reabsorptive needs. In summary, genes in this pathway undergo complex transcriptional and post-transcriptional regulation, resulting in the differential expression of novel alternative transcripts that contribute to the fine-tuning of BP control.