Year-end Sale % OFF 
Abstract
Two genes that encode enzymes in cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase and HMG-CoA synthase, and the gene encoding the low density lipoprotein (LDL) receptor are repressed when sterols accumulate in animal cells. Their 5'-flanking regions contain a common element, designated sterol regulatory element-1 (SRE-1). In the HMG-CoA synthase and LDL receptor promoters, the SRE-1 enhances transcription in the absence of sterols and is inactivated in the presence of sterols. In the HMG-CoA reductase promoter, the region containing the SRE-1 represses transcription when sterols are present. In the current studies, we show that the SRE-1 retains enhancer function but loses sterol sensitivity in mutant Chinese hamster ovary cells that are resistant to the repressor, 25-hydroxycholesterol. In the absence of sterols, the mutant cells produced high levels of all three sterol-regulated mRNAs, and there was no repression by 25-hydroxycholesterol. When transfected with plasmids containing each of the regulated promoters fused to a bacterial reporter gene, the mutant cells showed high levels of transcription in the absence of sterols and no significant repression by sterols. When the SRE-1 in the LDL receptor and HMG-CoA synthase promoters was mutated prior to transfection into the mutant cells, transcription was markedly reduced. Thus, the 25-hydroxycholesterol-resistant cells retain a protein that enhances transcription by binding to the SRE-1 in the absence of sterols, but they have lost the function of a protein that abolishes this enhancement in the presence of sterols. Mutation of a 30-base pair segment of the HMG-CoA reductase promoter that contains the SRE-1 did not reduce transcription in the mutant cells, indicating that this promoter is driven by elements other than the SRE-1. Nevertheless, this promoter failed to be repressed by sterols in the mutant cells. These data suggest that a common factor mediates the effects of sterols on the SRE-1 in all three promoters and that this factor has been functionally lost in the 25-hydroxycholesterol-resistant cells.
Highlights
1).In the HMG-CoA synthase and LDL receptor promoters, the SRE-1 enhances transcription in the absence of sterols and is inactivated in the presence of sterols
Two lines of mutant 25-hy- when incubated with LDLs,uggesting that 25-hydroxycholesdroxycholesterol-resistantcells were isolated independently terol did not suppress LDL receptors in these mutant cells as described under “ExperimentalProcedures.” One line,des
The 1and SRD-2cells was resistant to suppression25b-yhydroxyother cell line, SRD-2, was mutagenized with y-irradiation, cholesterol.Enzyme activity was not which favors deletions [26].Following mutagenesis, cells were repressed to a normalextent by LDL-derivedcholesterol isolatedbasedupon theirabilityto survivegrowth in the
Summary
LDL receptorsin these wild-type cells (see below). IgnatedSRD-1, was mutagenized withnitrosoethylurea, Fig. 3 shows that the HMG-CoA reductase activofitSyRD-. The 1and SRD-2cells was resistant to suppression25b-yhydroxyother cell line, SRD-2, was mutagenized with y-irradiation, cholesterol (panels A and D).Enzyme activity was not which favors deletions [26].Following mutagenesis, cells were repressed to a normalextent by LDL-derivedcholesterol isolatedbasedupon theirabilityto survivegrowth in the (panel B and E). Absence of LDL and in the presence of 1 pg/ml 25-hydroxy- CoA reductase activity in thewild-type CHO cells but not in cholesterol, a concentration that kills 100% of the wild-type the mutant cells (panels C and F ).
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
