HMG-CoA reductase regulation: use of structurally diverse first half-reaction squalene synthetase inhibitors to characterize the site of mevalonate-derived nonsterol regulator production in cultured IM-9 cells.

The activity of HMG-CoA reductase (HMGR) is tightly regulated, in part through post-transcriptional mechanisms that are mediated by nonsterol products of mevalonate metabolism. Previous reports have suggested that these mediators are derived from farnesyl pyrophosphate (FPP). Recent studies have implicated FPP hydrolysis products (e.g., farnesol), the squalene synthetase (SQS) reaction products presqualene pyrophosphate (PSQPP) and squalene, or their metabolites. To distinguish among these possible mediators, we evaluated the ability of HMGR and SQS inhibitors to induce compensatory increases in HMGR activity in cultured IM-9 cells. Mevinolin (HMGR inhibitor) produced predicted increases in HMGR activity that were related to the degree of cholesterolgenesis inhibition (e.g., 4-fold, 9-fold, and 17-fold increases relative to 50%, 76%, and 90% inhibition, respectively). By contrast, a variety of structurally distinct reversible, competitive, first half-reaction SQS inhibitors all reduced cholesterolgenesis by up to 90% with no appreciable increases in HMGR activity. These observations strongly suggest that nonsterol-mediated post-transcriptional mechanisms regulating HMGR activity remain intact after SQS first half-reaction inhibition, indicating that nonsterol regulator production is independent of SQS action and ruling out PSQPP, squalene and their metabolites as possible mediators. Unexpectedly, the SQS mechanism-based irreversible inactivator, zaragozic acid A (ZGA) exhibited the greatest degree of HMGR modulation, producing 5-fold, 11-fold, and 40-fold increases in HMGR activity at concentrations that produced 25%, 50%, and 75% cholesterolgenesis inhibition, respectively. The markedly greater magnitude of HMGR stimulation by ZGA versus mevinolin at similar levels of cholesterolgenesis inhibition suggests that ZGA may directly interfere with the production or action of the nonsterol regulator.