Hydroxyurea increases eNOS protein levels through inhibition of proteasome activity.
Recent reports have identified the proteasome as the primary degradation pathway for inducible, neuronal and endothelial nitric oxide synthase (NOS). We have demonstrated that hydroxyurea increased nitric oxide (NO) production in endothelial cells through phosphorylation of eNOS as a short-term effect. We find now that NO production in endothelial cells is dose-dependently stimulated by hydroxyurea, as well as both specific and non-specific proteasome inhibitors, as a long term effect. Prolonged treatment of primary human umbilical vein endothelial cells (HUVEC) with hydroxyurea was found to increase eNOS protein levels without an effect on eNOS mRNA levels, suggesting posttranscriptional control. We observed that the inhibitors of proteasomes that we tested also increased eNOS protein levels in HUVEC. In a proteasome assay, we showed that hydroxyurea inhibited protein degradation in a dose-dependent manner, in both purified 20S proteasome and HUVEC lysates. The NO production induced by hydroxyurea in endothelial cells appears to be mediated by long term posttranscriptional augmentation in eNOS levels via inhibition of the proteasome activity.
Cokic
Vladan P
VP
Laboratory of Experimental Hematology, Institute for Medical Research, Dr. Subotica 4, P.O. Box 102, 11129 Belgrade, Serbia. vl@imi.bg.ac.yu
Beleslin-Cokic
Bojana B
BB
Noguchi
Constance T
CT
Schechter
Alan N
AN
D000970:Antineoplastic Agents / Q000494:pharmacology; D002460:Cell Line; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology*; D007783:Lactones / Q000494:pharmacology; D007976:Leupeptins / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D007783:Lactones; D007976:Leupeptins; D061988:Proteasome Inhibitors; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D006918:Hydroxyurea