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Targeting Gene Expression to the Lung
Published in Kenneth L. Brigham, Gene Therapy for Diseases of the Lung, 2020
Jeffrey A. Whitsett, Stephan W. Glasser
AP-1 family members bind to a cis-acting element located near a TTF-1 binding site in the first intron of the mouse gene. Its occupation and function are critical to SP-B gene expression but do not suffice to confer lung cell specificity per se. Together the SP-B (F1) and SP-B (F2) serve to confer lung cellspecific gene expression in the context of the basal SP-B promoter. The element does not activate gene expression from other basal promoter elements (20). In contrast, an upstream SP-B enhancer, located approximately -350 from the start of transcription of the human SP-B gene, contains TTF-1 sites (two or three in the human or mouse gene, respectively) that markedly enhance gene expression and serve as an enhancer of viral promoters. TTF-1 is critical to the activity of the distal element; mutations blocking TTF-1 binding to this site markedly inhibited enhancer activity (24). The distal SP-B enhancer acts in an orientation-independent fashion and can activate a minimal promoter from the herpes simplex thymidine kinase gene, producing lung cellselective control of gene expression after transfection of respiratory epithelial cells in vitro. Such elements can therefore be introduced into gene transfer vectors to activate gene expression in a respiratory epithelial cell selective manner.
Expression of Major Histocompatibility Antigens on Fetal and Placental Cells
Published in Gérard Chaouat, The Immunology of the Fetus, 2020
The transient expression of CAT (chloramphenicol acetyl transferase)-gene constructs with 1.4 kb of the promoter sequence of H-2Ld was analyzed in F9 cells. The native upstream region of 1.4 kb gave very low activity as expected. Deletions analysis of this construct in differentiated and undifferentiated cells suggested that there is a negative regulation of the expression of H-2Ld in EC cells due to the presence of a cis-acting element situated between position -195bp (base pair upstream to the start site) and -l61bp.94 The H-2Kd gene does not appear to be subjected to a negative regulation in undifferentiated EC cells.101 This region was first described in the H-2Kd gene promoter as the Class-I regulatory element (CRE) (see Figure 8). It is a highly conserved sequence among MHC Class-I genes, also present in the 5’ upstream region of the B-2m gene, which is able to increase the CAT-activity in NIH3T3 cells that express MHC Class-I genes constitutively, that is, it induces an enhancing effect (Figure 8). This enhancer is about five times less active in nondiffer-entiated PCOAza RI EC cells, compared to the differentiated cell line derived from it.95
The Role of Plasminogen Activator Inhibitor Type 1 (PAI-1) in the Clinical Setting, Including Deep Vein Thrombosis
Published in Pia Glas-Greenwalt, Fibrinolysis in Disease Molecular and Hemovascular Aspects of Fibrinolysis, 2019
More recently a third polymorphism was found in the promoter region of the PAI-1 gene.48,50 The single base pair insertion/deletion polymorphism (GACACGTG4 or 5AGT) is located 675 base pairs upstream from the transcription initiation site of the PAI-1 gene. Individuals homozygous for the del allele (GGGG) exhibited higher plasma PAI-1 activity than those with at least one ins allele (5 Gs). In the patients, the difference was significant (4G/4G: 21.5 ng/ml versus at least one 5G: 16.1 mg/ml; p = 0.03). Gelshift assays, footprinting studies, and transfection experiments with the promoter region linked to a reporter gene suggested that the 5G polymorphic site functions as a part of a cis-acting element for the binding of a protein that acts as a transcription repressor: in particular, this element appears to be involved in the blunting of the cells’ response to the stimulating effect of interleukin-1 on PAI-1 expression.
Transcriptional regulation of CYP3A4 by nuclear receptors in human hepatocytes under hypoxia
Published in Drug Metabolism Reviews, 2020
Xuechun Yuan, Hui Lu, Anpeng Zhao, Yidan Ding, Qiong Min, Rong Wang
Moreover, another response element located on the upstream of CYP3A4 gene (from −11.4 to −10.5 kb relative to the transcription start site) has been identified as the constitutive liver enhancer module of CYP3A4 gene (CLEM4), which contains an array of cis-acting elements encompassing ∼900 bp, activated by the collaboration of several transcription factors including HNF-1, HNF-4, USF1 and AP-1. Among them, HNF-4 and its binding sites may be more crucial for the expression of hepatic CYP3A4 gene (Matsumura et al. 2004). Furthermore, a specific cis-acting element has been confirmed to exist in CYP3A4 gene enhancer, which allows the binding of HNF4α and thereby permits PXR- and CAR-mediated gene activation (Tirona et al. 2003). As a result, HNF-4 should not be overlooked in mediating the transcription of CYP3A4 gene. A brief overview of the 5′-flanking region of CYP3A4 gene regulation by these mentioned NRs in the liver is showed in Figure 1.
Transcriptional control of the MUC16 promoter facilitates follicle-stimulating hormone peptide-conjugated shRNA nanoparticle-mediated inhibition of ovarian carcinoma in vivo
Published in Drug Delivery, 2018
Ming-Xing Zhang, Shan-Shan Hong, Qing-Qing Cai, Meng Zhang, Jun Chen, Xiao-Yan Zhang, Cong-Jian Xu
Here, we predicted the promoter sequences in the regions that were 4 kbp upstream of MUC16 and measured transcriptional activity in ovarian cancer cells with dual-luciferase reporter assay. As shown in our study, although the MUC16.1 and MUC16.2 promoter constructs resulted in higher luciferase gene expression in HEY cells, the nanoparticle complexes containing the MUC16.2 promoter-driven plasmid did not significantly reduce the gro-α levels. It seemed that the promoter sequence with TAAA repeats, the MUC16.1 promoter, showed high transcriptional activity, and this sequence might contain an atypical TATA box and be a transcriptional recognition site. The plasmid containing gro-α shRNA driven by this MUC16 promoter sequence decreased gro-α protein secretion in ovarian cancer cells. However, the MUC16.2 promoter sequence had no possible transcriptional recognition sites such as TATA box. The MUC16.3 promoter sequence with CT repeats that made up a potential cis-acting element did not exhibit transcriptional activity. However, the transcriptional activity of the MUC16 promoter that we screened was lower than that of the positive control, namely, the CMV promoter. Some DNA elements, including enhancers, might need to be integrated into vectors to increase transcriptional activity in future studies (Rama et al., 2015).
Sickle cell disease in the era of precision medicine: looking to the future
Published in Expert Review of Precision Medicine and Drug Development, 2019
Martin H Steinberg, Sara Kumar, George J. Murphy, Kim Vanuytsel
Five common haplotypes of the HbS gene, defined by polymorphisms or SNPs cis to HBB, are associated with different HbF levels and disease phenotypes [10–12]. Patients with the Arab-Indian (AI) and Senegal haplotype have the mildest disease; Bantu or Central African Republic (CAR) haplotype patients have the most severe disease; Benin and Cameroon haplotype patients are intermediate. The association of a haplotype with disease severity correlates with the HbF level typical of each haplotype. In adult haplotype homozygotes, levels of HbF average about 20% in the AI, 10% in the Senegal and 7% to 4% in the Cameroon, Benin and Bantu haplotypes. The mechanism underlying differential HbF expression among haplotypes is unclear and is most likely to be a cis-acting effect. In children with the AI haplotype HbF levels average about 30%. These children can have a mild disease but as they age and HbF levels fall to 15% to 20% the disease becomes more severe [13,14]. The AI haplotype has unique cis and trans-acting variants but whether or not they are responsible for the high HbF is unknown [15–19]. A C-T polymorphism (rs7482144) 158 bp 5ʹ to HBG2 (–158) in the proximal promoter of this gene creates a cleavage site for the restriction endonuclease Xmn1. This SNP is present only in the AI and Senegal haplotypes. It is associated with increased expression of only HBG2 with a corresponding increase only in Gγ globin. There is little evidence that rs7482144 is functional. Recent studies of BCL11A binding in the HbF gene promoters do not support a mechanistic role for rs7482144 suggesting that it is in linkage disequilibrium with the functional cis-acting element of these haplotypes [20]. Ascertaining HbS-associated haplotypes has been useful epidemiologically and anthropologically but their prognostic relevance in individuals is minimal.