Insulin-like growth factor 1 (IGF1) exerts important endocrine and paracrine functions in the cardiovascular system. We identified the common variant -1411C>T in the IGF1 upstream promoter P1, located within several overlapping transcription factor binding sites. Using transient transfection assays, we identified this site as a functional enhancer. The T allele-carrying enhancer, compared with the C allelic portion, exerts significantly reduced or even abrogated activity, respectively, in SaOs-2 and HepG2 (all P<0.0001) as well as in differentiated THP-1 macrophages. Electrophoretic mobility shift assay and subsequent supershift experiments in HepG2 identified c-Jun as the binding partner exclusively to the T allele, whereas CCAAT/enhancer-binding protein delta and interferon consensus site-binding protein/interferon-regulating factor 8 interacted only with the C allelic promoter portion. Furthermore, genotyping in a case-control study for essential hypertension (n=745 hypertensive patients; n=769 normotensive control subjects) for this variant revealed an odds ratio for hypertension of 0.73 (95% confidence interval 0.58-0.91, P=0.006) associated with the T allele, and normotensive subjects carrying the protective T allele displayed a significant decrease in diastolic (P=0.036) and systolic (P=0.024) blood pressure levels. We here report detection of a functional enhancer module in the upstream IGF1 promoter region, which might play a key role in local IGF1 bioavailability. Whether -1411C>T is also associated with other IGF1-related disease phenotypes should be evaluated further in population studies.
Abstract-The G protein-coupled receptor kinase 4 is involved in renal sodium handling and blood pressure regulation.Missense variants have already been tested functionally and are associated with hypertension, but no data on promoter analyses are yet available. We scanned 94 hypertensive white subjects for genetic variation and performed promoter reporter gene analyses in HEK293T, COS7, and SaOs-2 cells. Transient transfections with various full lengths and wild-type deletion constructs revealed that 1851 bp of the flanking region and 275 bp of the 5Ј-untranslated region were sufficient for transcriptional activities and composed a powerful cis-active element in the distal 293 bp. The Ϫ1702T and ϩ2T alleles resulted in drastic general reductions of promoter function, whereas an activity increasing effect of ϩ268C was cell type specific. Electrophoretic mobility-shift assay, supershift, and cotransfection analyses of transcription factor binding sites predicted in silico (Alibaba2.1/Transfac7) resulted in allele-specific binding patterns of nuclear proteins and identified the participation of CCAAT/enhancer-binding protein transcription factor family members. The G protein-coupled receptor kinase 4 core promoter resides in the first 1851 bp upstream of its transcription start site. The 4 identified genetic variants within this region exert allele-specific impact on both cell typeand stimulation-dependent transcription and may affect the expression balance of renal G protein-coupled receptor kinase 4. (Hypertension. 2008;52:737-746.)
Human thrombopoietin (TPO) is involved in cardiovascular disease as it regulates megakaryocyte development and enhances platelet adhesion/aggregation. The THPO promoter structure is still controversial. By reverse transcription-PCR, we confirm that THPO transcription is cell line-dependently initiated at two alternative promoters, which we newly designated P1a and P1. We subsequently electrophoretically scanned and resequenced these portions in 95 and 57 patients with cardiovascular disease, respectively, and identified seven variants (
Objective— Osteoprotegerin (OPG) has been reported to be involved in the development of atherosclerotic disease, and OPG gene variation has been associated with plasma OPG levels and different cardiovascular disease phenotypes. However, the genetic architecture of the OPG promoter and its transcriptional regulation are poorly characterized. Methods and Results— We identified 1008 bp of the OPG 5′-flanking region to be sufficiently transcriptionally active in osteosarcoma cell lines and generated serial promoter deletion constructs. Individual subcloning revealed the existence of 3 molecular haplotypes (MolHaps): [T −960 -A −946 -G −900 -T −864 ; MolHap1, wild type], [T −960 -G −946 -G −900 -T −864 ; MolHap2], [C −960 -G −946 -A −900 -G −864 ; MolHap4]. Compared to MolHap1, transcriptional activities of MolHaps 2 and 4 were significantly reduced ( P =0.0018). Whereas introduction of the −159C allele reduced transcriptional activities of the full-length constructs ( P =0.0014), it significantly increased activities of the deletion constructs ( P =0.0005). Electrophoretic mobility shift, competition, and chromatin immunoprecipitation assays revealed specific DNA:protein interactions for the MolHaps with Sp1 and NF-1, and identified Egr1 interacting exclusively with the −159T allele. Conclusions— We propose new structural and transcriptional features within the OPG promoter region and identified MolHaps being differentially transcriptionally active and allele-dependently interacting with a proximal polymorphic site.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.