| 1. |
- Heverin, Maura, et al.
(författare)
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On the regulatory importance of 27-hydroxycholesterol in mouse liver
- 2017
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Ingår i: Journal of Steroid Biochemistry and Molecular Biology. - : Elsevier. - 0960-0760 .- 1879-1220. ; 169, s. 10-21
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Forskningsöversikt (refereegranskat)abstract
- 27-Hydroxycholesterol (27OH) is a strong suppressor of cholesterol synthesis and a weak activator of LXR in vitro. The regulatory importance of 27OH in vivo is controversial. Here we utilized male mice with increased levels of 27OH either due to increased production (CYP27A1 transgenic mice) or reduced metabolism (Cyp7b1-/- mice). We also used mice lacking 27OH due to a knockout of Cyp27a1. The latter mice were treated with cholic acid to compensate for reduced bile acid synthesis. The effects of the different levels of 27OH on Srebp- and other LXR-regulated genes in the liver were investigated. In the liver of CYP27tg mice we found a modest increase of the mRNA levels corresponding to the LXR target genes Cyp7b1 and Abca1. A number of other LXR-regulated genes were not affected. The effect on Abca1 mRNA was not seen in the liver of Cyp7b1-/- mice. There were little or no effects on cholesterol synthesis. In the liver of the Cyp27-/- mice treated with 0.025% cholic acid there was no significant effect of the knockout on the LXR target genes. In a previous work triple-knockout mice deficient in the biosynthesis of 24S-hydroxycholesterol, 25-hydroxycholesterol and 27OH were shown to have impaired response to dietary cholesterol, suggesting side-chain oxidized oxysterols to be mediators in cholesterol-induced effects on LXR target genes at a transcriptional level (Chen W. et al., Cell Metab. 5 (2007) 73-79). The hydroxylated oxysterol responsible for the effect was not defined. We show here that treatment of wildtype mice with dietary cholesterol under the same conditions as in the above study induced the LXR target genes Lpl, Abcg8 and Srebp1c in wild type mice but failed to activate the same genes in mice lacking 27-hydroxycholesterol due to a knockout of Cyp27. We failed to demonstrate the above effects at the protein level (Abcg8) or at the activity level (Lpl). The results suggest that 27OH is not an important regulator of Srebp- or LXR regulated genes under basal conditions in mouse liver. On the other hand 27OH appears to mediate cholesterol-induced effects on some LXR target genes at a transcriptional level under some in vivo conditions.
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| 2. |
- Reilly, Sarah-Jayne, et al.
(författare)
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The nudix hydrolase 7 is an acyl-CoA diphosphatase involved in regulating peroxisomal coenzyme A homeostasis.
- 2008
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Ingår i: Journal of Biochemistry. - : Oxford University Press (OUP). ; 144:5, s. 655-663
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Tidskriftsartikel (refereegranskat)abstract
- Coenzyme A (CoASH) is an obligate cofactor for lipids undergoing beta-oxidation in peroxisomes. Although the peroxisomal membrane appears to be impermeable to CoASH, peroxisomes contain their own pool of CoASH. It is believed that CoASH enters peroxisomes as acyl-CoAs, but it is not known how this pool is regulated. The mouse nudix hydrolase 7 (NUDT7alpha) was previously identified in peroxisomes as a CoA-diphosphatase, and therefore suggested to be involved in regulation of peroxisomal CoASH levels. Here we show that mouse NUDT7alpha mainly acts as an acyl-CoA diphosphatase, with highest activity towards medium chain acyl-CoAs, and much lower activity with CoASH. Nudt7alpha mRNA is highly expressed in liver, brown adipose tissue and heart, similar to enzymes involved in peroxisomal lipid degradation. Nudt7alpha mRNA is downregulated by Wy-14,643, a peroxisome proliferator-activated receptor alpha (PPARalpha) ligand, in a PPARalpha dependent manner in mouse liver. In highly purified peroxisomes, nudix hydrolase activity is highest with C(6)-CoA and is decreased by fibrate treatment. Under certain conditions, such as treatment with peroxisome proliferators or fasting, an increase in peroxisomal CoASH levels has been reported, which is in line with a decreased expression/activity of NUDT7alpha. Taken together these data suggest that NUDT7alpha function is tightly linked to peroxisomal CoASH/acyl-CoA homeostasis.
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| 3. |
- Tillander, Veronika
(författare)
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Peroxisomal and mitochondrial enzymes involved in lipid metabolism : studies on function and regulation
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Fatty acids constitute a major part of the energy that we obtain from the diet and are also the principal source for mammals to store energy. To use the incoming or stored fatty acids as energy, the fatty acids needs to be metabolized of which the majority of fatty acids will be degraded by the mitochondrial β-oxidation system that in the end generates energy to the cell in the form of ATP. However, this organelle is not able to handle all kinds of fatty acids of which very long chain fatty acids, long chain methyl-branched fatty acids and dicarboxylic acids are such cumbersome fatty acids. Therefore a second organelle, the peroxisome, is required for metabolism of these particular fatty acids. Also peroxisomes contain a β-oxidation system and similar to the mitochondrial system is the initial substrate a CoA-esterified fatty acid, so-called acyl-CoA. This thesis will focus on some enzymes that are active on these acyl-CoA esters, but that are not directly involved in the β-oxidation per se. Instead they contribute to the regulation of both acyl-CoA and free coenzyme A levels in different cellular compartments. This thesis will also include how these fatty acid degrading systems can be regulated at gene level by affecting different transcription factors by dietary ligands and by fasting. The peroxisomal Nudix hydrolase 7α (NUD7α), previously believed to be a CoASH degrading enzyme, was demonstrated to be a medium chain diphosphatase, most active on medium chain acyl-CoA esters, to produce 3’,5’-ADP and the corresponding 4’-acylphosphopantetheine thereof. NUDT7α expression and activity was down regulated by PPARα activation, which would prevent CoASH degradation and support a high rate of the β-oxidation in peroxisomes during these conditions. Peroxisomes are not only needed for the degradation of complex lipids, but are also essential for many other metabolic pathways such as bile acid and etherphospholipid synthesis and the degradation of D-amino acids and glyoxylate. The expression of gene transcripts that code for the proteins involved in these peroxisomal pathways was investigated almost throughout the whole mouse body with the aim to map the tissue expression of these pathways. The peroxisomal β-oxidation system is present in all examined tissues, however with differences in magnitude. More specifically expressed pathways are e.g. glyoxylate and D-amino acid degradation pathways. Auxiliary enzymes to the peroxisomal β-oxidation showed tissue specific expression, suggesting a high degree of tissue specific metabolite patterns, also being dependent on the metabolic state. The study also shows that PPARα is of major importance for the regulation in liver of the peroxisomal “transcriptome” during fasting. Mitochondria degrade both fatty acids and amino acids and the mitochondrial acyl-CoA thioesterase 9 (ACOT9) was shown to hydrolyze both long chain acyl-CoAs as well as short chain acyl-CoA intermediates and products of branched-chain amino acid metabolism. Kinetic characterization of the enzyme suggests a thigh regulation of the activity during different metabolic conditions in the mitochondria. Dietary ω-3 PUFAs from fish oil (FO) and krill oil (KO) cause different changes in lipid profiles and gene regulation when supplemented to mice. FO lowered most plasma lipids whereas KO only significantly lowered non-esterified fatty acids in plasma. FO showed a classical PPARα activation response by up regulating genes for fatty acid utilization and oxidation whereas KO down regulates genes for cholesterol and fatty acid synthesis.
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| 4. |
- Tillander, Veronika
(författare)
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The mammalian peroxisome : ‛its function and distribution’
- 2010
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Peroxisomes are essential organelles for normal cell functions in all organisms from yeast to human. Their important contribution in different metabolic pathways is clearly visible by the severe phenotypes seen in the majority of peroxisomal diseases, where the symptoms often leads to an early death. Peroxisomes are involved in the synthesis of etherphospholipids and bile acids, in the metabolism of certain amino acids, purines and glyoxylate, and do also harbour an advanced system for degradation of various types of fatty acids and complex lipids. Peroxisomes are dynamic organelles that respond to different physiological and pharmacological changes by changing their number and contents of certain proteins. We carried out a tissue expression and regulation study on the majority of all known peroxisomal proteins (here called the ‘Pexiome’) in mouse at mRNA level to investigate if and how the different pathways may differ in their expression through out the mouse body. We studied how the mRNA expression varies in liver, kidney and intestinal epithelial in response to 12 hrs fasting, and also the effect of peroxisome proliferator activating receptor α (PPARα) agonist administration on gene expression in liver. The results show that indeed the mRNA expression of different genes varies markedly among tissues, while a number of genes seem to have a very wide tissue expression, which is in line with the content of peroxisomes in all cell types. Interestingly, fasting has a profound effect on the expression of the ‘Pexiome’ and also affects the peroxisomal gene expression in a strongly tissue specific manner. By examination of mouse livers from fasted and PPARα agonist treated animals on PPARα (+/+) and PPARα (-/-) backgrounds, it was evident that the regulation of most of the peroxisomal genes by fasting is far more complex than just involving PPARα activation. We also carried out an in depth study on the mouse peroxisomal Nudix hydrolase 7α (NUDT7α), which had previously been shown to act as a CoASH diphosphatase. Our data show that NUDT7α preferably cleaves off 3’,5’-ADP from the CoA-moiety of medium chain acyl-CoA’s. The expression of the enzyme at mRNA level was down regulated during PPARα activation in liver, and we also found that the total Nudix hydrolase activity was decreased in rat liver peroxisomes isolated from clofibrate treated mice. These findings suggest that NUDT7α may be an important regulator of the peroxisomal CoASH pool, and likely also regulates the β-oxidation of fatty acids in the peroxisome at substrate level
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| 5. |
- Waluk, Dominik P., et al.
(författare)
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Molecular characterization of two members of the glycine N-acyltransferase gene family in human: glycine N-acyl transferase-like 1 (GLYATL1) and glycine N-acyltransferase-like 3 (GLYATL3).
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- N-acyl amino acids are a group of endogenous lipid mediators that regulate a variety of cellular physiological functions. The discovery of N-acyl amino acids in many biological systems has allowed research to focus on their functions as well as pathways for production of these signalling lipids. We have previously identified that human glycine N-acyltransferase-like 2 (hGLYATL2) is involved in the enzymatic formation of N-acyl glycines. hGLYATL2 is localized in a gene cluster with other glycine N-acyltransferase genes. Here, we have characterized human glycine N-acyltransferase-like 1 (hGLYATL1) and human glycine N-acyltransferase-like 3 (hGLYATL3), which are members of this gene family. Our results show that hGLYATL1 is localized to the endoplasmic reticulum (ER) but the intracellular localization of hGLYATL3 remains to be determined. The hGLYATL1 mRNA shows highest expression in liver and kidney, whereas mRNA of hGLYATL3 is expressed in pancreas and liver. Using bioinformatics we determined the overall three-dimensional (3D) structures of hGLYATL1 and hGLYATL3 enzymes, with predicted binding site residues. In summary, we have characterized novel members of glycine N-acyltransferases that may be involved in the production of lipid signalling molecules, in particular N-acyl glycines.
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