A Cycle of Altered Proteasome and Reactive Oxygen Species Production in Renal Proximal Tubular Cells.

AIMS:An intricate relationship exists between the mitochondrial function and proteasome activity. Our recent report showed in a rat model of renal transplantation that mitochondrial dysfunction precedes compromised proteasome function and this results in a vicious cycle of mitochondrial injury and proteasome dysfunction. In this study, we studied whether reactive oxygen species (ROS) has a role in proteasome alteration in renal cells and vice versa.METHODS:We used the genomic and pharmacologic approach on rat normal kidney proximal tubular (NRK) cell lines. First, we knocked down β5 or Rpt6 subunit of the proteasome using small interfering RNA (siRNA) in NRK cells. We also treated NRK cells with Bortezomib, a proteasome inhibitor, and peroxynitrite (a potent ROS). RESULTS:Studies with RNA interference showed increased mitochondrial ROS following knockdown of β5 or Rpt6 subunit in NRK cells. Similarly, pharmacological inhibition of the proteasome in NRK cells using Bortezomib also showed an increase of mitochondrial ROS in a dose-dependent manner. Next, exposing NRK cells to different concentrations of peroxynitrite provided evidence that the higher levels of peroxynitrite exposure decreased the key subunits (β5 and α3) of the proteasome in NRK cells. CONCLUSION:Our results suggest that proteasome inhibition/downregulation increases ROS, which then impairs proteasome subunits in renal proximal tubular cells.