Abstract
The activity of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, an enzyme which plays a regulatory role in the synthesis of cholesterol, dolichol, and coenzyme Q, has been measured in the developing embryo of the sea urchin. Enzyme activity increased at least 200-fold during development from the unfertilized egg to the pluteus stage embryo. Mixing experiments suggested that the low level of enzyme activity found at early stages was not due to the presence of inhibitor(s) in the egg or zygote. The enzyme in the sea urchin embryo exhibited properties different from that found in mammals: only a fraction of the activity could be solubilized from microsomes, and mild trypsinization inactivated the enzyme without releasing any of it from the microsomes in soluble form. To further study the sea urchin HMG-CoA reductase, a genomic clone was identified by hybridization to a cDNA encoding hamster HMG-CoA reductase. Sequence analysis of this clone revealed a coding region that shares a high degree of homology with the carboxyl-terminal domain of hamster HMG-CoA reductase. Analysis of sea urchin embryo HMG-CoA reductase mRNA levels using a restriction fragment derived from the genomic clone revealed a 5.5-kilobase poly(A)+ mRNA that increased 15-fold during development from the egg to the gastrula stage and then decreased 1.5-fold at the pluteus stage. Since the relative increase in HMG-CoA reductase mRNA was less than the increase in enzyme activity (15-fold versus 200-fold) factors in addition to the level of mRNA may control the activity of this enzyme during embryogenesis.
Highlights
From the Departmentof Biochemistry and Molecular Biology, The University of Texas System Cancer Center, M
This catalytically activefragment can be released in soluble form from the endoplasmic reticulum by either endogenous or exogenous proteases (2, 8).The single carbohydrate attachment site islocated in the aminoterminal domain of the molecule and is oriented toward the the sea urchin HMG-CoA reductase, a genomic clone lumen of the endoplasmic reticulum (2, 3)
Analysis of sea urchin trolling enzyme of polyisoprenoid biosynthesis (9) and proembryo HMG-CoA reductase mRNA levels using a re- vides the precursor, mevalonic acid, common to cholesterol, striction fragment derived from thgenomic clone re- dolichol, and coenzyme Q
Summary
Sequence analysis of hG1 revealedother featuresof interest. The present study demonstrates thHatMG-CoA reductase activity is developmentally regulated during sea urchin embryogenesis. The results indicate that sea urchin embryo the hamster HMG-CoA reductase cDNA suggest that XG1. Embryo HMG-CoA reductase mRNA over the course of em- Another difference between the sea urchin embryo enzyme bryonic development suggests that de nouo transcription may and the mammalian reductase is thata variety of detergents contribute to theobserved increase in enzyme activity during solubilize only 20-30%of the sea urchin enzyme. Precedent for such a mode of regulation is found contrast to studies that hsahvowe n quantitative solubilization in the case of rats, where diets rich in cholestyramine and of enzyme activity from UT-1 cells with Triton X-100 and mevinolin result in a 20-fold increase in reductase transcripdeoxycholate detergents (1). We are indebted to Diana Welch for preparation of the manuscript
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