Ultrastructural analysis of crystalloid endoplasmic reticulum in UT-1 cells and its disappearance in response to cholesterol.
The crystalloid endoplasmic reticulum (ER) consists of hexagonally packed membrane tubules that contain 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG CoA reductase), an intrinsic membrane protein that catalyses the rate-limiting step in cholesterol synthesis. The crystalloid ER appears in a clone of Chinese hamster ovary cells, designated UT-1, that contain high levels of HMG CoA reductase as a result of growth in the presence of compactin, a competitive inhibitor of the reductase. In the present studies, we have used ultrastructural morphometry to estimate that the crystalloid ER: (1) occupies about 15% of the volume of UT-1 cells; (2) contains 3.4-fold more membrane area than the plasma membrane; and (3) contains less than 700 subunits of HMG CoA reductase per micrometer2 of membrane surface. The crystalloid ER tubules contain 2000 intramembrane particles per micrometer2 with a mean diameter of 10.4 nm, as determined by freeze-fracture. The crystalloid ER membranes are low in cholesterol, as indicated by the small number of filipin-cholesterol complexes in freeze-fracture images after treatment with filipin. The addition of cholesterol or related sterols to UT-1 cells promoted a rapid and stepwise disappearance of the crystalloid ER. Initially, the crystalloid ER fragmented into randomly arranged vesicles and tubules. Subsequently, membrane-bound structures disappeared from the cell so that after incubation with cholesterol for 24-72 h, the cells appeared completely normal. We found no morphological evidence that autophagic vacuoles participate in the degradation. We conclude: (1) that the crystalloid ER is more extensive than necessary merely to support HMG CoA reductase; and (2) that upon exposure to cholesterol the crystalloid ER is degraded by a process that does not involve autophagy.
D000818:Animals; D002478:Cells, Cultured; D002784:Cholesterol / Q000494:pharmacology*; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure*; D005260:Female; D005372:Filipin; D005614:Freeze Fracturing; D006801:Humans; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism; D007425:Intracellular Membranes / Q000648:ultrastructure; D008854:Microscopy, Electron; D008870:Microtubules / Q000648:ultrastructure; D010053:Ovary / Q000648:ultrastructure
D005372:Filipin; D002784:Cholesterol; D006903:Hydroxymethylglutaryl CoA Reductases