RNA-SEQ REVEALS HSP90 AS A REGULATOR FOR INTERLEUKIN 6-MEDIATED ACTIVATION OF NCC VIA THE MINERALOCORTICOID RECEPTOR

Abstract

Objective: Hypertension (HTN) is characterized by increased Na+ reabsorption and systemic inflammation including increased cytokines such as IL6. Clinically, IL6 is increased in hypertensive patients. Previous data from our laboratory has revealed a mechanism for IL6-mediated activation of the mineralocorticoid receptor (MR), leading to increased Na+ transporter activation and HTN via reactive oxygen species (ROS) and Rac1. We hypothesized that IL6 induces a specific transcriptome in the late distal nephron (DCT2) and used RNA-seq to investigate. Design and method: Mouse DCT2 (mDCT15) cells were treated with IL6 (100ng/mL, 24hrs) or vehicle (n = 4, each), then total RNA isolated. Data from the Illumina-sequenced transcriptome underwent comparative analysis to identify significant differentially expressed transcripts between vehicle and IL6 treated cells. Results: Differential gene expressed revealed multiple targets, including HSP90 (3.03x10-6, p-adj) encoding for heat shock protein (HSP) 90. HSP90 has been previously shown as crucial for ligand binding, MR nuclear translocation and activation. To investigate whether inhibition of HSP90 would affect IL6-mediated MR activation of distal Na+ transporters, such as the sodium chloride cotransporter (NCC), a thiazide-sensitive Na+ uptake was performed on transwells in vehicle, IL6 (100ng/mL) or IL6-Gel (geldanamycin, HSP90 inhibitor; 5uM) in mDCT15 cells (n = 3). IL6 treatment increased Na+ uptake compared (1666 ± 12 nmol/mg/20 min vs.1449 ± 7; p < 0.001). With HSP90 inhibition, Na+ uptake was reduced (1499 ± 12; p < 0.001). Conclusions: Using RNA-seq, we have uncovered specific transcripts activated within the late distal nephron (DCT2) following IL6, including HSP90, which we confirmed to be an important and novel player in IL6-mediated activation of NCC.