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A TG-associated minor LPL haplotype supresses miR-29 binding on LPL 3'UTR
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International audience. Aim: Identification and functional study of Lipoprotein Lipase (LPL) 3'UTR polymorphisms associated with triglyceridemia and predicted to modulate miRNA binding.Methods: We sequenced LPL 3'UTR in 271 type 2 diabetic patients with documented lipid phenotypes. Haplotype analysis was performed using PHASE software. These data were validated by meta-analysis across four French general population samples. In silico analysis identified putative miRNA binding-sites created by LPL polymorphisms. Both LPL 3'UTR wild-type (WT) and variant (VAR) sequences were cloned in luciferase reporter gene plasmid. Transfection experiments in HEK293T and co-transfection with candidate miRNA-precursor was performed to study the regulation of the LPL constructs by specific miRNA.Results: A specific haplotype combining 8 LPL polymorphisms including the rs328 and 3'UTR minor alleles was found significantly associated with lower triglycerides concentration both in type 2 diabetes and in general populations. This haplotype was predicted to disrupt several miR-29 binding-sites. In vitro, the LPL-WT construct co-transfected with miR-29 showed a significant luciferase activity decrease (-25±6%, p< 0.001), whereas no difference was observed in the presence of the LPL-VAR construct. Moreover in HEK293T cells expressing endogenously miR-29, LPL-WT haplotype expression was lower compared to LPL-VAR haplotype (-32±7%, p<0.0001). Finally, the introduction of either the rs328 minor or major alleles did not modify the expression of LPL-WT and LPL-VAR haplotype.Conclusion: We identify a 'gain of function' LPL haplotype significantly associated with lower TG concentration which may increase LPL expression and activity by disrupting miR-29 binding-sites. The miRNA regulation of this haplotype is independent of rs328.
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