Increase In Sterol Content Research Articles

Overexpression of HMG-CoA synthase promotes Arabidopsis root growth and adversely affects glucosinolate biosynthesis.

3-Hydroxy-3-methylglutaryl-CoA synthase (HMGS) catalyses the second step of the mevalonate (MVA) pathway. An HMGS inhibitor (F-244) has been reported to retard growth in wheat, tobacco, and Brassica juncea, but the mechanism remains unknown. Although the effects of HMGS on downstream isoprenoid metabolites have been extensively reported, not much is known on how it might affect non-isoprenoid metabolic pathways. Here, the mechanism of F-244-mediated inhibition of primary root growth in Arabidopsis and the relationship between HMGS and non-isoprenoid metabolic pathways were investigated by untargeted SWATH-MS quantitative proteomics, quantitative real-time PCR, and target metabolite analysis. Our results revealed that the inhibition of primary root growth caused by F-244 was a consequence of reduced stigmasterol, auxin, and cytokinin levels. Interestingly, proteomic analyses identified a relationship between HMGS and glucosinolate biosynthesis. Inhibition of HMGS activated glucosinolate biosynthesis, resulting from the induction of glucosinolate biosynthesis-related genes, suppression of sterol biosynthesis-related genes, and reduction in sterol levels. In contrast, HMGS overexpression inhibited glucosinolate biosynthesis, due to down-regulation of glucosinolate biosynthesis-related genes, up-regulation of sterol biosynthesis-related genes, and increase in sterol content. Thus, HMGS might represent a target for the manipulation of glucosinolate biosynthesis, given the regulatory relationship between HMGS in the MVA pathway and glucosinolate biosynthesis.

Hypoxia Is Present in Murine Atherosclerotic Plaques and Has Multiple Adverse Effects on Macrophage Lipid Metabolism

Human atherosclerotic plaques contain large numbers of cells deprived of O(2). In murine atherosclerosis, because the plaques are small, it is controversial whether hypoxia can occur. To examine if murine plaques contain hypoxic cells, and whether hypoxia regulates changes in cellular lipid metabolism and gene expression in macrophages. Aortic plaques from apolipoprotein-E-deficient mice were immunopositive for hypoxia-inducible transcription factor (HIF-1α) and some of its downstream targets. Murine J774 macrophages rendered hypoxic demonstrated significant increases in cellular sterol and triglycerides. The increase in sterol content in hypoxic macrophages correlated with elevated 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase activity and mRNA levels. In addition, when macrophages were incubated with cholesterol complexes, hypoxic cells accumulated 120% more cholesterol, predominately in the free form. Cholesterol-efflux assays showed that hypoxia significantly decreased efflux mediated by ATP-binding cassette subfamily A member 1 (ABCA1), whose sub cellular localization was altered in both J774 and primary macrophages. Furthermore, in vivo expression patterns of selected genes from cells in hypoxic regions of murine plaques were similar to those from J774 and primary macrophages incubated in hypoxia. The hypoxia-induced accumulation of sterol and decreased cholesterol efflux was substantially reversed in vitro by reducing the expression of the hypoxia-inducible transcription factor, HIF-1α. Hypoxic regions are present in murine plaques. Hypoxic macrophages have increased sterol content due to the induction of sterol synthesis and the suppression of cholesterol efflux, effects that are in part mediated by HIF-1α.

Cloning, functional expression and phylogenetic analysis of plant sterol 24C-methyltransferases involved in sitosterol biosynthesis

Sterol 24C-methyltransferases (SMTs) constitute a group of sequence-related proteins that catalyze the distinct patterns of 24-alkyl sterols that occur throughout nature. Two SMT cDNAs (SMT2-1 and SMT2-2) were cloned by homology based PCR methods from young leaves of Glycine max (soybean) and the corresponding enzymes were expressed functionally in Escherichia coli. The full-length cDNA for SMT2-1 and SMT2-2 have open reading frames of 1086 bp and 1092 bp, respectively, and encode proteins of 361 and 363 residues with a calculated molecular mass of 40.3 and 40.4 kDa, respectively. The substrate preference of the two isoforms was similar yet they differed from SMT1; kinetically SMT2-1 and SMT2-2 generated k cat values for the optimal substrate 24(28)methylene lophenol of 0.8 min −1 and 1.34 min −1, respectively, compared to the activity of SMT1 that generated a k cat for the optimal substrate cycloartenol of 0.6 min −1. SMT2-2 was purified to homogeneity and the subunit organization shown to be tetrameric in similar fashion to other cloned SMTs. Analysis of the accumulated products catalyzed by the recombinant enzymes demonstrated that soybean SMT2-1 and SMT2-2 operate transalkylation activities analogous to the soybean plant SMT1. Metabolite analyses correlated with transcript profiling of the three SMT isoforms during soybean maturation clearly demonstrated that SMT isoform expression determines specific C24-methyl to C24-ethyl ratios to flowering whereas with seed development there is a disconnection such that the SMT transcript levels decrease against an increase in sterol content; generally SMT2-2 is expressed more than SMT2-1 or SMT1. These observations suggest that the genes that encode SMT1 and SMT2 in sitosterol biosynthesis may have undergone divergent evolution. In support of this proposition, the genomic organization for SMT1 of fungi and protozoa align very closely with one another and to those of the plant SMT2; both sets of SMTs lack introns. Unexpectedly, the SMT1 from Glycine max and other embryophytes of diverse origin possess disparate intron–exon characteristics that can be shown relates back to the algae. Our results suggest that the order of SMT1 appearing before SMT2 in phytosterol synthesis arose recently in plant evolution in response to duplication of a more primitive SMT gene likely to have been bifunctional and catalytically promiscuous.

Dietary cholesterol utilization by the housefly, Musca domestica

The amount of cholesterol and the distribution of its metabolites were studied in various larval tissues of the housefly ( Musca domestica), reared on a high cholesterol diet containing the radioactive cholesterol. The sterol content was also examined in adults derived from larvae reared on increasing dietary cholesterol concentration in separate batches. The results suggest that when the dietary sterol concentration was raised from 0.002 to 0.02% wet weight (an optimal concentration required for a maximal growth and development of insect), the sterol content of six-day old larvae was increased approx. 4-fold. However, a further 10-fold increase of dietary cholesterol (to 0.22% of wet weight) was associated with only a 1.5-fold increase in larval sterol content, and no increase in the sterol content of adult insects derived from such larvae was observed. This increase of sterol content of whole larvae was found to be confined to the larval cuticle and composite gut fractions and may be attributed to unabsorbed cholesterol in the gut, and to a solubilizing effect of the cuticle. These findings suggest that when the cholesterol requirement for the maximal growth of the insect has been reached, larvae are able to regulate the intake of ingested cholesterol, and no more is taken up through the gut even when a high gut concentration of cholesterol is present. The adults contained a higher percentage of esterified sterols than the larvae, approx. 25% of the sterol in females and 14% of sterol in males were esterified.

Sterols in hardening winter wheat

Abstract The main sterols in winter wheat crowns and roots were sitosterol and campesterol, with significant amounts of stigmasterol and traces of cholesterol. The main groups of sterol-containing lipids were free sterols, steryl glucosides, steryl esters and esterified steryl glucosides. Sterol analysis within each group showed little difference between them. Steryl esters were relatively rich in cholesterol and poor in stigmasterol. Free sterols were rich in stigmasterol. Low temperature caused an increase in sterol content but had little effect on sterol composition and sterol to lipid P ratio. There was some increase in steryl esters and some decrease in free sterols. Cholesterol and stigmasterol decreased in the steryl ester and free sterol fractions, respectively. There was little evidence for involvement of sterols in winter wheat frost hardening.

The Effects of Catabolite Derepression on the Accumulation of Steryl Esters and the Activity of -Hydroxymethylglutaryl-CoA Reductase in Saccharomyces cerevisiae

Summary: The transition from the fermentative mode of growth on glucose to the oxidative mode of growth on ethanol by Saccharomyces cerevisiae is accompanied by many changes collectively known as catabolite derepression. Catabolite derepression caused a progressive increase in the total content of cellular sterols, amounting to a fourfold increase after 8 h; the additional sterols existed almost entirely as esters of monoenoic fatty acids. The increase in sterol content could be explained by the large increase in the activity of β-hydroxymethylglutaryl-CoA reductase (EC 1.1.1.34), but a secondary factor in sterol accumulation may have been the slower rate of cell growth during ethanol oxidation. Subsequent anaerobic growth of derepressed cells resulted in the hydrolysis of steryl esters to free sterols, and a progressive depletion of cellular sterol content.