Peroxiredoxin‐4 Interacts with Dopamine D5 Receptor to Reduce Endoplasmic Reticulum Stress

Peroxiredoxin-4 (PRDX4), an endoplasmic reticulum (ER)-localized antioxidant enzyme, plays an essential role in cellular redox homeostasis by reducing hydrogen peroxide from thiol-containing compounds. The dopamine D5 Receptor (D5R) also plays a protective role against oxidative stress. We hypothesize that D5 R interacts with the PRDX4 to reduce oxidative stress in the kidney. In D5 R-HEK 293 cells, fenoldopam (FEN, 25 nM/12 hr, n=4), a D1 R and D5 R receptor agonist, increased PRDX4 protein expression (1.92±0.12-fold over basal level, n=4), mainly in non-lipid raft (LR) fractions (LRs: 24.9±11.4%, non-LRs: 75.1±11.4%, baseline; LRs: 30.9±13.9%, non-LRs: 174.1±16.7%, FEN). By contrast, fenoldopam did not affect PRDX4 protein expression in D1 R-HEK 293 cells, indicating a D5 R-specific effect on PRDX4 protein expression. In human renal proximal tubule cells (hRPTCs) and D5 R HEK293 cells, fenoldopam increased the co-immunoprecipitation between PRDX4 and D5 R and their co-localization in the ER. SiRNA-mediated silencing of PRDX4 increased hydrogen peroxide production in both the vehicle (Veh)- and fenoldopam-treated hRPTCs [(scrambled siRNA: 100 ±15.1% and 55.2± 7.2% with Veh and FEN, respectively; PRDX4 siRNA: 161.8±15.3% and 145.1±14.6 % with Veh and FEN, respectively, n=4)]. The D5 R protects against inflammation and siRNA silencing of PRDX4 increased the production of the pro-inflammatory cytokines, interleukin-1β [26.88±3.8 and 46.40±4.2 pg/mL (n=3, D5 R-HEK 293); 15.87±1.2 and 37.9±1.4 pg/mL (n=3, hRPTCs), and tumor necrosis factors [131.7±6.5 and 271.2±18.1 pg/mL (n=4, D5 R-HEK 293); 108.8±11.8 and 240.1±13.7 pg/mL (n=4, hRPTCs)]. In D5 R-HEK293 and hRPTCs, the fenoldopam-mediated decrease in ER-resident caspase-12 was also impaired by gene silencing of PRDX4. Furthermore, PRDX4 protein expression was reduced in the kidney cortices of Drd5-/- mice, relative to Drd5 WT mice (WT: 1.00±0.12, n=4; Drd5-/- : 0.64±0.13, n=4; P<0.05). We conclude that D5 R positively interacts with PRDX4 to reduce ER stress in the kidney.