Browsing by Subject "Cystic Fibrosis Transmembrane Conductance Regulator"

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Open Access
A C-terminal motif found in the beta2-adrenergic receptor, P2Y1 receptor and cystic fibrosis transmembrane conductance regulator determines binding to the Na+/H+ exchanger regulatory factor family of PDZ proteins.
(Proc Natl Acad Sci U S A, 1998-07-21) Hall, RA; Ostedgaard, LS; Premont, RT; Blitzer, JT; Rahman, N; Welsh, MJ; Lefkowitz, RJ
The Na+/H+ exchanger regulatory factor (NHERF) binds to the tail of the beta2-adrenergic receptor and plays a role in adrenergic regulation of Na+/H+ exchange. NHERF contains two PDZ domains, the first of which is required for its interaction with the beta2 receptor. Mutagenesis studies of the beta2 receptor tail revealed that the optimal C-terminal motif for binding to the first PDZ domain of NHERF is D-S/T-x-L, a motif distinct from those recognized by other PDZ domains. The first PDZ domain of NHERF-2, a protein that is 52% identical to NHERF and also known as E3KARP, SIP-1, and TKA-1, exhibits binding preferences very similar to those of the first PDZ domain of NHERF. The delineation of the preferred binding motif for the first PDZ domain of the NHERF family of proteins allows for predictions for other proteins that may interact with NHERF or NHERF-2. For example, as would be predicted from the beta2 receptor tail mutagenesis studies, NHERF binds to the tail of the purinergic P2Y1 receptor, a seven-transmembrane receptor with an intracellular C-terminal tail ending in D-T-S-L. NHERF also binds to the tail of the cystic fibrosis transmembrane conductance regulator, which ends in D-T-R-L. Because the preferred binding motif of the first PDZ domain of the NHERF family of proteins is found at the C termini of a variety of intracellular proteins, NHERF and NHERF-2 may be multifunctional adaptor proteins involved in many previously unsuspected aspects of intracellular signaling.
Open Access
A complex intronic enhancer regulates expression of the CFTR gene by direct interaction with the promoter.
(J Cell Mol Med, 2009-04) Ott, Christopher J; Suszko, Magdalena; Blackledge, Neil P; Wright, Jane E; Crawford, Gregory E; Harris, Ann
Genes can maintain spatiotemporal expression patterns by long-range interactions between cis-acting elements. The cystic fibrosis transmembrane conductance regulator gene (CFTR) is expressed primarily in epithelial cells. An element located within a DNase I-hypersensitive site (DHS) 10 kb into the first intron was previously shown to augment CFTR promoter activity in a tissue-specific manner. Here, we reveal the mechanism by which this element influences CFTR transcription. We employed a high-resolution method of mapping DHS using tiled microarrays to accurately locate the intron 1 DHS. Transfection of promoter-reporter constructs demonstrated that the element displays classical tissue-specific enhancer properties and can independently recruit factors necessary for transcription initiation. In vitro DNase I footprinting analysis identified a protected region that corresponds to a conserved, predicted binding site for hepatocyte nuclear factor 1 (HNF1). We demonstrate by electromobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP) that HNF1 binds to this element both in vitro and in vivo. Moreover, using chromosome conformation capture (3C) analysis, we show that this element interacts with the CFTR promoter in CFTR-expressing cells. These data provide the first insight into the three- dimensional (3D) structure of the CFTR locus and confirm the contribution of intronic cis-acting elements to the regulation of CFTR gene expression.
Open Access
Regionally distinct progenitor cells in the lower airway give rise to neuroendocrine and multiciliated cells in the developing human lung.
(Proceedings of the National Academy of Sciences of the United States of America, 2023-06) Conchola, Ansley S; Frum, Tristan; Xiao, Zhiwei; Hsu, Peggy P; Kaur, Kamika; Downey, Michael S; Hein, Renee FC; Miller, Alyssa J; Tsai, Yu-Hwai; Wu, Angeline; Holloway, Emily M; Anand, Abhinav; Murthy, Preetish Kadur Lakshminarasimha; Glass, Ian; Tata, Purushothama R; Spence, Jason R
Using scRNA-seq and microscopy, we describe a cell that is enriched in the lower airways of the developing human lung and identified by the unique coexpression of SCGB3A2/SFTPB/CFTR. To functionally interrogate these cells, we apply a single-cell barcode-based lineage tracing method, called CellTagging, to track the fate of SCGB3A2/SFTPB/CFTR cells during airway organoid differentiation in vitro. Lineage tracing reveals that these cells have a distinct differentiation potential from basal cells, giving rise predominantly to pulmonary neuroendocrine cells and a subset of multiciliated cells distinguished by high C6 and low MUC16 expression. Lineage tracing results are supported by studies using organoids and isolated cells from the lower noncartilaginous airway. We conclude that SCGB3A2/SFTPB/CFTR cells are enriched in the lower airways of the developing human lung and contribute to the epithelial diversity and heterogeneity in this region.
Open Access
Understanding Cystic Fibrosis Transmembrane Conductance Regulator expression in Heart Failure
(2019-04-22) Ramesh, Arjun
Cardiovascular disease, specifically congestive heart failure, is a leading cause of death in the United States. Cystic Fibrosis (CF) is caused by the mutations to the cystic fibrosis transmembrane conductance regulator (CFTR) gene. These mutations result in a defective or absent CFTR protein in the lung epithelial cells. Not a well-known concept is that the CFTR protein is present in more than just the lungs - its presence in the cardiac tissue may be critical for heart function. Preliminary research from the Bowles lab demonstrates that CFTR expression and function are reduced in diseased human cardiac tissue. Also, DNA sequencing suggests a potential cause for this diminishment: a genetic enhancer in the CFTR gene is different in heart failure patients compared to healthy controls. This difference has been seen in a small study of 48 patients. In this independent study, I examined the CFTR gene through PCR and gel electrophoresis analysis, as well as consolidating previous work from the Bowles lab and others in the CFTR field, to provide an in depth look at this region of the gene. The challenges of this PCR study have held back results of amplification data due to complications with the protocol and electrophoresis. Near the end of the study, successful PCR and gel electrophoresis was completed, showing the technique was achievable after trouble-shooting. This technique will be applied to a larger sample set of genomic DNA to be amplified and sequenced. This study may set the stage for using the CFTR enhancer region as a biomarker of heart failure. In addition, it may provide preliminary data to the Bowles lab for the use of CFTR modulation in the treatment of heart failure.