HUMAN RENAL RNA-SEQ IDENTIFIES RELATIONSHIP BETWEEN UMOD EXPRESSION AND MEDULLARY ION TRANSPORTERS

Abstract

Objective: Our previous genome wide association study (GWAS) highlighted a locus on the promoter of the human UMOD gene associated with hypertension. In this study, we sought to identify potential pathways which facilitate the association of UMOD with blood pressure through effects on gene expression. Design and method: From our bank of 98 human renal biopsy samples, we assessed UMOD and NKCC2 expression by qRT-PCR. Individuals were stratified by UMOD expression and RNA-sequencing using the NextSeq-500 system performed on a subset of high and low UMOD expression (n = 3/ group). Differential expression was assessed by DESeq2 to detect significant differences (p [adjusted] <0.05) in the transcriptome between high and low UMOD expressers and pathways visualised using Ingenuity Pathway Analysis (IPA). Results: RNA-Seq analysis identified 162 differentially expressed genes (138 protein coding genes,18 long non-coding RNAs and 8 additional transcriptomic elements), of these, 123 genes increased in expression with high UMOD expression. Using IPA, we detected enrichment for pathways related to immune system function, cell to cell communication and molecular ion transport. Notably, we show that an increase in UMOD expression associates with an increase in expression of the Na-K-Cl cotransporter (NKCC2) (p [adjusted] = 2.58e-9, fold-change = 10.89), the renal outer medullary potassium channel (ROMK) (p [adjusted] = 3.69e-2, fold-change = 2.79) and the gamma-subunit of the epithelial sodium channel (ENaC) (p [adjusted] = 1.83e-2, fold-change = 1.25). We further validated the relationship between UMOD and NKCC2, by Taqman qRT-PCR, showing a highly significant correlation of expression (p = 2.2e-16, R = 0.844) (n = 86). Using IPA we show this correlation may be facilitated by the transcription factor POU3F3, which can act at motifs found on promoters of both genes. Conclusions: Of several pathways identified by IPA, association of UMOD expression with expression of these three ion channels (NKCC2, ROMK and ENaC) may explain previously observed relationship between UMOD and blood pressure. These data suggest the UMOD: ion-transport pathway may be a drug target in the treatment of hypertension.