45 research outputs found
Promoter Elements and Factors Involved In Hepatic Transcription of the Human Apoa-i Gene Positive and Negative Regulators Bind To Overlapping Sites
No full textDNase I footprinting analysis of the proximal apoA-I promoter sequences with rat liver nuclear extracts identified four protected regions: A, -22 to +17; B, -128 to -77; C, -175 to -148; and D, -220 to -190. Region D (-220 to -190) binds at least two distinct activities, designated AID1 and AID2, respectively, which can be separated by ion exchange chromatography. Region C (-175 to -148) forms five DNA protein complexes. Three of the complexes (2, 4, and 5) originate from the binding of more than one heat-stable nuclear factor, and two (1 and 3), from the binding of two heat-labile factors. The heat-stable factors bind in the -175 to -148 region and can be distinguished from C/EBP, which recognizes the same region, with DNA binding gel electrophoretic assays. Both factors 1 and 3 bind in the -168 to -148 apoA-I region. Despite the lack of a CCAAT motif in this region, the binding of factor 1 is competed out by oligonucleotides containing the binding sites of NFY and NFY*. Mutagenesis of the promoter region showed that mutations in the -171 to -166 and -158 to -153 regions diminished the binding of the heat-stable factors and reduced hepatic transcription to 14 and 8% of control, respectively. In contrast, a mutation in the -164 to -159 region abolished the binding of factor 1 and was associated with a 4.6-fold increase in hepatic transcription. These findings suggest that the heat-stable factors act as positive regulators, whereas factor 1 acts as a negative regulator in apoA-I gene transcription
Expression of multiple α2-adrenergic receptor messenger rna species in rat tissues
No full textWe used a human platelet α2-adrenergic receptor probe to study the tissue distribution and messenger RNA (mRNA) forms of the rat α2-adrenergic receptor. Under stringent conditions of hybridization and washing, we detected an mRNA species of 3.8 kb. The abundance of this form follows the order spleen, kidney, brain stem and cortex, and skeletal muscle and lung and is consistent with the reported abundance and tissue distribution of the receptor activity. A 3.0 kb mRNA form was also detected in cerebral cortex and brain stem and a 4.1 kb mRNA form was observed in kidney under less stringent hybridization conditions. The tissue distribution of the 3.0 kb form is different from that of α1,-and β-adrenergic receptors and the D2 dopaminergic receptor. The mRNA analysis combined with Southern blot analysis of rat and human genomic DNA indicate that: 1) in addition to a 3.8 kb rat os-adrenergic receptor transcript, there are other mRNA forms in the rat that do not correspond to previously described adrenergic receptor mRNA species and 2) more tban one α2-adrenergic receptor gene in the rat Is expressed in a tissue-specific manner. © 1990 American Heart Association, Inc
Transcriptional Regulation of the Human ApoA-I Gene in Cell Culture and in ApoA-I Transgenic Mice Regulation of ApoA-I Gene Expression and Prospects to Increase Plasma ApoA-I and HDL Levels
No full textAbbreviations there is a linkage and common regulatory mechanism of the apoA-I/apoCIII/apoA-IV gene cluster The receptor specificity of the HREs of the apoA-I promoter and the apoCIII enhancer were determined by DNA binding gel electrophoresis assays. These analyses established that both HREs present in the proximal apoA-I promoter bind HNF-4, other orphan receptors, and a variety of liganddependent nuclear receptors with different affinities Transcriptional regulation of the human apoA-I gene in transgenic mice. To validate the conclusions drawn by the in vitro experiments, we generated a variety of transgenic mouse lines which express the WT A-I/CIII cluster or the same cluster with the mutations in the HREs and the binding sites of SP1 and other transcription factors. In these constructs, the apoCIII gene was replaced by the CAT gen
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No full textNom de l'autor obtingut del principi del textEl v. del darrer f. en blanc, signatures: a-t6El comentari envolta el text i registre després del colofóCaplletres xilogràfiques de dues mide
Role of Esrrg in the Fibrate-Mediated Regulation of Lipid Metabolism Genes in Human ApoA-I Transgenic Mice
No full textWe have used a new ApoA-I transgenic mouse model to identify by global gene expression profiling, candidate genes that affect lipid and lipoprotein metabolism in response to fenofibrate treatment. Multilevel bioinformatical analysis and stringent selection criteria (2-fold change, 0% false discovery rate) identified 267 significantly changed genes involved in several molecular pathways. The fenofibrate-treated group did not have significantly altered levels of hepatic human APOA-I mRNA and plasma ApoA-I compared with the control group. However, the treatment increased cholesterol levels to 1.95-fold mainly due to the increase in high-density lipoprotein (HDL) cholesterol. The observed changes in HDL are associated with the upregulation of genes involved in phospholipid biosynthesis and lipid hydrolysis, as well as phospholipid transfer protein. Significant upregulation was observed in genes involved in fatty acid transport and β-oxidation, but not in those of fatty acid and cholesterol biosynthesis, Krebs cycle and gluconeogenesis. Fenofibrate changed significantly the expression of seven transcription factors. The estrogen receptor-related gamma gene was upregulated 2.36-fold and had a significant positive correlation with genes of lipid and lipoprotein metabolism and mitochondrial functions, indicating an important role of this orphan receptor in mediating the fenofibrate-induced activation of a specific subset of its target genes.National Institutes of Health (HL48739 and HL68216); European Union (LSHM-CT-2006-0376331, LSHG-CT-2006-037277); the Biomedical Research Foundation of the Academy of Athens; the Hellenic Cardiological Society; the John F Kostopoulos Foundatio
Carboxyl terminus of apolipoprotein A-I (ApoA-I) is necessary for the transport of lipid-free ApoA-I but not prelipidated ApoA-I particles through aortic endothelial cells
No full textHigh density lipoproteins (HDL) and apolipoprotein A-I (apoA-I) must leave the circulation and pass the endothelium to exert their atheroprotective actions in the arterial wall. We previously demonstrated that the transendothelial transport of apoA-I involves ATP-binding cassette transporter (ABC) A1 and re-secretion of lipidated particles. Transendothelial transport of HDL is modulated by ABCG1 and the scavenger receptor BI (SR-BI). We hypothesize that apoA-I transport is started by the ABCA1-mediated generation of a lipidated particle which is then transported by ABCA1-independent pathways. To test this hypothesis we analyzed the endothelial binding and transport properties of initially lipid-free as well as prelipidated apoA-I mutants. Lipid-free apoA-I mutants with a defective carboxyl-terminal domain showed an 80% decreased specific binding and 90% decreased specific transport by aortic endothelial cells. After prior cell-free lipidation of the mutants, the resulting HDL-like particles were transported through endothelial cells by an ABCG1- and SR-BI-dependent process. ApoA-I mutants with deletions of either the amino terminus or both the amino and carboxyl termini showed dramatic increases in nonspecific binding but no specific binding or transport. Prior cell-free lipidation did not rescue these anomalies. Our findings of stringent structure-function relationships underline the specificity of transendothelial apoA-I transport and suggest that lipidation of initially lipid-free apoA-I is necessary but not sufficient for specific transendothelial transport. Our data also support the model of a two-step process for the transendothelial transport of apoA-I in which apoA-I is initially lipidated by ABCA1 and then further processed by ABCA1-independent mechanisms
Parameters calculated from DLS experiments for apoE3 variants (Figure 8).
No full text<p>Calculated particle diameters are given in nm. Errors indicate peak width. Peak 2 corresponds to ∼1% of the total mass of the protein sample and indicates higher order aggregates. Units are in nm. Values are means ± SD from three experiments.</p><p>*non-detected.</p><p>**I = diameter calculated based on the distribution of intensities of scattering, V = diameter calculated after normalization of the scattering distribution based on particle volume occupancy.</p
Intracellular and extracellular processing of human apolipoprotein A-I: secreted apolipoprotein A-I isoprotein 2 is a propeptide.
No full textHDL-apoA-I induces the expression of angiopoietin like 4 (ANGPTL4) in endothelial cells via a PI3K/AKT/FOXO1 signaling pathway
No full textBackground: High Density Lipoprotein (HDL) and its main protein component, apolipoprotein A-I (apoA-I), have numerous atheroprotective functions on various tissues including the endothelium. Therapies based on reconstituted HDL containing apoA-I (rHDL-apoA-I) have been used successfully in patients with acute coronary syndrome, peripheral vascular disease or diabetes but very little is known about the genomic effects of rHDL-apoA-I and how they could contribute to atheroprotection. Objective: The present study aimed to understand the endothelial signaling pathways and the genes that may contribute to rHDL-apoA-I-mediated atheroprotection. Methods: Human aortic endothelial cells (HAECs) were treated with rHDL-apoA-I and their total RNA was analyzed with whole genome microarrays. Validation of microarray data was performed using multiplex RT-qPCR. The expression of ANGPTL4 in EA.hy926 endothelial cells was determined by RT-qPCR and Western blotting. The contribution of signaling kinases and transcription factors in ANGPTL4 gene regulation by HDL-apoA-I was assessed by RT-qPCR, Western blotting and immunofluorescence using chemical inhibitors or siRNA-mediated gene silencing. Results: It was found that 410 transcripts were significantly changed in the presence of rHDL-apoA-I and that angiopoietin like 4 (ANGPTL4) was one of the most upregulated and biologically relevant molecules. In validation experiments rHDL-apoA-I, as well as natural HDL from human healthy donors or from transgenic mice overexpressing human apoA-I (TgHDL-apoA-I), increased ANGPTL4 mRNA and protein levels. ANGPTL4 gene induction by HDL was direct and was blocked in the presence of inhibitors for the AKT or the p38 MAP kinases. TgHDL-apoA-I caused phosphorylation of the transcription factor forkhead box O1 (FOXO1) and its translocation from the nucleus to the cytoplasm. Importantly, a FOXO1 inhibitor or a FOXO1-specific siRNA enhanced ANGPTL4 expression, whereas administration of TgHDL-apoA-I in the presence of the FOXO1 inhibitor or the FOXO1-specific siRNA did not induce further ANGPTL4 expression. These data suggest that FOXO1 functions as an inhibitor of ANGPTL4, while HDL-apoA-I blocks FOXO1 activity and induces ANGPTL4 through the activation of AKT. Conclusion: Our data provide novel insights into the global molecular effects of HDL-apoA-I on endothelial cells and identify ANGPTL4 as a putative mediator of the atheroprotective functions of HDL-apoA-I on the artery wall, with notable therapeutic potential. © 2018 Elsevier Inc
EFFECTS OF APOE VARIANTS ON PLTP INTERACTION AND ACTIVATION OF LOW AND HIGH ACTIVITY FORMS OF PLTP
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