A Conserved Degron Containing an Amphipathic Helix Regulates the Cholesterol‐Mediated Turnover of Human Squalene Monooxygenase, a Rate‐Limiting Enzyme in Cholesterol Synthesis

Abstract

Cholesterol biosynthesis is a key process in cellular cholesterol homeostasis and is known to be tightly regulated by several mechanisms. Of note, squalene monooxygenase (SM), the second rate‐limiting enzyme in cholesterol biosynthesis, is regulated by transcriptional and posttranslational mechanisms. SM is degraded by the ubiquitin proteasome system when cholesterol is in excess. The cholesterol‐mediated degradation of SM depends on the first 100 amino acids (designated SM N100), and this represents the shortest cholesterol‐regulated degron identified to date. Despite having an important function, how these 100 amino acids grant SM the ability to be degraded by cholesterol remains elusive. From our mutational analyses, we have identified a 12‐residue region (Gln‐62 to Leu‐73) which is necessary for this degron to be successfully degraded by excess cholesterol. Using molecular dynamics simulation and circular dichroism, we identified the 12‐residue region contains an amphipathic helix. Our analyses also revealed that mutating SM residues Phe‐35, Ser‐37, Leu‐65 and Ile‐69 into alanine based on the key residues of the well‐studied Doa10 yeast degron, Deg1, also blunted the cholesterol‐mediated degradation of SM. In this study, we propose a model whereby the amphipathic helix of SM N100 is associated with the endoplasmic reticulum membrane when cholesterol is not in excess, and that excess cholesterol triggers the dissociation of this helix, unraveling it and exposing a hydrophobic patch which promotes degradation. Our current study reveals a key feature of how this rate‐limiting enzyme can regulate cholesterol levels, by sensing endoplasmic reticulum membrane cholesterol levels via a newly identified amphipathic helix. More importantly, we identified conservation of a key degron feature, namely an amphipathic helix, from yeast to humans.Support or Funding InformationNational Health and Medical Research Council (1060515)Australian Research Council (DP170101178)DST‐INSPIRE Faculty Grant LSBM‐45Department of Biotechnology, Government of India, Research associateship.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.