Abstract
Backgroundβ-oxidation of long and very long chain fatty acyl-CoA derivatives occurs in peroxisomes, which are ubiquitous subcellular organelles of eukaryotic cells. This pathway releases acetyl-CoA as precursor for several key molecules such as cholesterol. Numerous enzymes participating to cholesterol and fatty acids biosynthesis pathways are co-localized in peroxisomes and some of their encoding genes are known as targets of the NFY transcriptional regulator. However, until now no interaction between NFY transcription factor and genes encoding peroxisomal β-oxidation has been reported.ResultsThis work studied the interactions between NFY factor with the rat gene promoters of two enzymes of the fatty acid β-oxidation, MFP-1 (multifunctional protein type 1) and ThB (thiolase B) and their involvement in the cholesterol dependent-gene regulation. Binding of this nuclear factor to the ATTGG motif of the MFP-1 and of the ThB promoters was demonstrated by EMSA (Electrophoretic Mobility Shift Assay) and super shift assay. In contrast, in spite of the presence of putative Sp1 binding sites in these promoters, competitive EMSA did not reveal any binding. The promoter-dependent luciferase gene expression was downregulated by cholesterol in MFP-1 and ThB promoters harbouring constructs.ConclusionsThis work describes for the first time a NFY interaction with promoter sequences of the peroxisomal β-oxidation encoding genes. It suggests that cholesterol would negatively regulate the expression of genes involved in β-oxidation, which generates the initial precursor for its own biosynthesis, via at least the NFY transcription factor.
Highlights
Background major pathways providing cellular energyBeta-oxidationFatty acid β-oxidation is with glycolysis one of the two of long and very long chain fatty acyl-CoA derivativesLipids in Health and Disease 2004, 3 http://www.lipidworld.com/content/3/1/4FNiugculeroet1ide sequences of the 5'-flanking regions of the multifunctional protein-1 (MFP-1) and thiolase-B (ThB) genes respectively Nucleotide sequences of the 5'-flanking regions of the multifunctional protein-1 (MFP-1) and thiolase-B (ThB) genes respectively
It suggests that cholesterol would negatively regulate the expression of genes involved in β-oxidation, which generates the initial precursor for it own biosynthesis, via at least the NFY transcription factor
Identification of trans-acting factors recognizing ATTGG motifs of the MFP-1 and ThB promoters In silico sequences analysis of rat peroxisomal genes encoding MFP-1 and ThB reveals the presence of inverted CCAAT boxes in both promoters, which are positioned from the transcription start sites at: -68/-64 for the MFP-1 gene and at -56/-60 and -105/-109 for the ThB gene (Figure 1)
Summary
Background major pathways providing cellular energyBeta-oxidationFatty acid β-oxidation is with glycolysis one of the two of long and very long chain fatty acyl-CoA derivatives (page number not for citation purposes)Lipids in Health and Disease 2004, 3 http://www.lipidworld.com/content/3/1/4FNiugculeroet1ide sequences of the 5'-flanking regions of the multifunctional protein-1 (MFP-1) and thiolase-B (ThB) genes respectively Nucleotide sequences of the 5'-flanking regions of the multifunctional protein-1 (MFP-1) and thiolase-B (ThB) genes respectively. The inverted CCAAT motif is http://www.lipidworld.com/content/3/1/4 a functional cis-acting element present in promoters and enhancer regions of many eukaryotic genes including those encoding for two type I collagens, albumin, hsp, major histocompatibility complex (MHC) class II and numerous cell-cycle-related proteins such as thymidine kinase and cyclins [5,6,7,8,9]. Portions of the amino acid sequences of mammalian NFY-A, NFY-B, and NFY-C are evolutionarily conserved, with a high degree of sequence identity with segments of yeast HAP3, HAP2, and HAP5, respectively [16,18,19] These polypeptides are components of a Saccharomyces cerevisiae CCAAT binding transcription factor which controls several nuclear genes involved in the mitochondrial function [20]. The role of NFY has been demonstrated in the transcriptional regulation of genes encoding enzymes implicated in two major synthetic pathways: 3hydroxy-3-methylglutaryl-CoA reductase and farnesyl diphosphate synthase for cholesterol biosynthesis, and acetyl-CoA carboxylase and fatty acid synthase for fatty acids biosynthesis [21,22,23,24,25]
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