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 thro ugh post-transcriptional mechanisms that are mediated by nonsterol products of mevalonate metabolism. Previous reports have suggested that these media tors are derived from farnesyl pyrophosphate (FPP). Recent studies have imp licated FPP hydrolysis products (e.g., farnesol), the squalene synthetase ( SQS) reaction products presqualene pyrophosphate (PSQPP) and squalene, or t heir metabolites. To distinguish among these possible mediators, we evaluat ed the ability of HMGR and SQS inhibitors to induce compensatory increases in HMGR activity in cultured IM-9 cells. Mevinolin (HMGR inhibitor) produce d predicted increases in HMGR activity that were related to the degree of c holesterolgenesis inhibition (e.g., 4-fold, 9-fold, and 17-fold increases r elative to 50%, 76%, and 90% inhibition, respectively). By contrast, a vari ety of structurally distinct reversible, competitive, first half-reaction S QS inhibitors all reduced cholesterolgenesis by up to 90% with no appreciab le increases in HMGR activity. These observations strongly suggest that non sterol-mediated posttranscriptional mechanisms regulating HMGR activity rem ain intact after SQS first half-reaction inhibition, indicating that nonste rol regulator production is independent of SQS action and ruling out PSQPP, squalene and their metabolites as possible mediators. Unexpectedly, the SQ S mechanism-based irreversible inactivator, zaragozic acid A (ZGA) exhibite d 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 m agnitude of HMGR stimulation by ZGA versus mevinolin at similar levels of c holesterolgenesis inhibition suggests that ZGA may directly interfere with the production or action of the nonsterol regulator.