Conserved elements within first intron of aquaporin-5 (Aqp5) function as transcriptional enhancers

Flodby, Per; Zhou, Baosen; Ann, David K.; Kim, Kwang‐Jin; Minoo, Parviz; Crandall, Edward D.; Borok, Zea

doi:10.1016/j.bbrc.2007.02.076

Cited by 8 publications

(5 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In contrast, expression of the EGFP transgene was much greater in the SMG than in the lung at both mRNA and protein levels, consistent with our previous reports showing that the 4.3-kb Aqp5 promoter may differentially regulate Aqp5 transcription in the salivary gland and lung in vitro (3,10). In contrast, expression of the EGFP transgene was much greater in the SMG than in the lung at both mRNA and protein levels, consistent with our previous reports showing that the 4.3-kb Aqp5 promoter may differentially regulate Aqp5 transcription in the salivary gland and lung in vitro (3,10).…”

Section: Discussionsupporting

confidence: 90%

Rat aquaporin-5 4.3-kb 5′-flanking region differentially regulates expression in salivary gland and lung in vivo

Zhou¹,

Ann²,

Flodby³

et al. 2008

American Journal of Physiology-Cell Physiology

Self Cite

View full text Add to dashboard Cite

We previously cloned a 4.3-kb genomic fragment encompassing 5'-flanking regulatory elements of rat aquaporin-5 (Aqp5) that demonstrated preferential transcriptional activity in lung and salivary cells in vitro. To investigate the ability of Aqp5 regulatory elements to direct transgene expression in vivo, transgenic (TG) mice and rats were generated in which the 4.3-kb Aqp5 fragment directed the expression of enhanced green fluorescent protein (EGFP). RT-PCR revealed relative promoter specificity for the lung and salivary glands in TG mice. Immunofluorescence microscopy showed strong EGFP expression in salivary acinar cells but not in lung type I (AT1) cells, both known sites of endogenous AQP5 expression. Similar results were obtained in TG rats generated by lentiviral transgenesis. EGFP mRNA was detected in both salivary glands and lung. Robust EGFP fluorescence was observed in frozen sections of the rat salivary gland but not in the lung or other tested tissues. The percentage of EGFP-positive acinar cells was increased in parotid and submandibular glands of TG rats receiving a chronic injection of the beta-adrenergic receptor agonist isoproterenol. EGFP-positive cells in the lung that were also reactive with the AT1-cell specific monoclonal antibody VIIIB2 were identified by flow cytometry. These findings demonstrate that the 4.3-kb Aqp5 promoter/enhancer directs strong cell-specific transgene expression in salivary gland and low-level AT1 cell-specific expression in the lung. While these Aqp5 regulatory elements should be useful for functional studies in salivary glands, additional upstream or intronic cis-active elements are likely required for robust expression in the lung.

show abstract

Section: Discussionsupporting

confidence: 90%

Rat aquaporin-5 4.3-kb 5′-flanking region differentially regulates expression in salivary gland and lung in vivo

Zhou¹,

Ann²,

Flodby³

et al. 2008

American Journal of Physiology-Cell Physiology

Self Cite

View full text Add to dashboard Cite

show abstract

“…This new Cre line will enable detailed studies of gene function by gene targeting to modulate gene expression specifically in AT1 cells in the distal mouse lung, thereby advancing our understanding of the functional contribution of this important cell type to alveolar homeostasis at baseline and in disease. a lung epithelial cell line (MLE-15), whereas no significant enhancer activity was found in a cell line derived from salivary gland (Pa-4) (13). These transfection studies suggest that sequences in intron 1 of Aqp5 have important regulatory functions, although it is unknown exactly which regulatory sequences are required for high-level Aqp5 expression in the lung in vivo.…”

Section: Clinical Relevancementioning

confidence: 85%

Directed Expression of Cre in Alveolar Epithelial Type 1 Cells

Flodby

Borok²,

Banfalvi³

et al. 2010

Am J Respir Cell Mol Biol

Self Cite

View full text Add to dashboard Cite

Pulmonary alveolar epithelium is comprised of two morphologically and functionally distinct cell types, alveolar epithelial type (AT) I and AT2 cells. Genetically modified mice with cell-specific Cre/loxP-mediated knockouts of relevant genes in each respective cell type would be useful to help elucidate the relative contributions of AT1 versus AT2 cells to alveolar homeostasis. Cre has previously been efficiently expressed in AT2 cells in mouse lung with the surfactant protein (SP)-C promoter; however, no transgenic mouse expressing Cre in AT1 cells has so far been available. To develop an AT1 cell-specific transgenic Cre mouse, we generated a knockin of a Cre-IRES-DsRed cassette into exon 1 of the endogenous aquaporin 5 (Aqp5) gene, a gene expressed specifically in AT1 cells in the distal lung epithelium, resulting in the mouse line, Aqp5-Cre-IRES-DsRed (ACID). Endogenous Aqp5 and transgenic Cre in ACID mice showed a very similar pattern of tissue distribution by RT-PCR. To analyze Cre activity, ACID was crossed to two Cre reporter strains, R26LacZ and mT/mG. Double-transgenic offspring demonstrated reporter gene expression in a very high fraction of AT1 cells in the distal lung, whereas AT2 cells were negative. As expected, variable reporter expression was detected in several other tissues where endogenous Aqp5 is expressed (e.g., submandibular salivary gland and stomach). ACID mice should be of major utility in analyzing the functional contribution of AT1 cells to alveolar epithelial properties in vivo with Cre/loxP-mediated gene deletion technology.

show abstract

“…Multiple examples were found, including enhancers in the first introns of peptidylargi-nine deiminase type I 27 and aquaporin 5 28 and 5′ to the keratin 5 gene. 29 The location of peaks of open chromatin in HTE cells coinciding with these mapped enhancers are shown in online supplementary figure 3.…”

Section: Resultsmentioning

confidence: 99%

A genome-wide analysis of open chromatin in human tracheal epithelial cells reveals novel candidate regulatory elements for lung function

Bischof

Ott

Leir

et al. 2011

Thorax

View full text Add to dashboard Cite

Background Distal cell-type-specific regulatory elements may be located at very large distances from the genes that they control and are often hidden within intergenic regions or in introns of other genes. The development of methods that enable mapping of regions of open chromatin genome wide has greatly advanced the identification and characterisation of these elements. Methods Here we use DNase I hypersensitivity mapping followed by deep sequencing (DNase-seq) to generate a map of open chromatin in primary human tracheal epithelial (HTE) cells and use bioinformatic approaches to characterise the distribution of these sites within the genome and with respect to gene promoters, intronic and intergenic regions. Results Genes with HTE-selective open chromatin at their promoters were associated with multiple pathways of epithelial function and differentiation. The data predict novel cell-type-specific regulatory elements for genes involved in HTE cell function, such as structural proteins and ion channels, and the transcription factors that may interact with them to control gene expression. Moreover, the map of open chromatin can identify the location of potentially critical regulatory elements in genome-wide association studies (GWAS) in which the strongest association is with single nucleotide polymorphisms in non-coding regions of the genome. We demonstrate its relevance to a recent GWAS that identifies modifiers of cystic fibrosis lung disease severity. Conclusion Since HTE cells have many functional similarities with bronchial epithelial cells and other differentiated cells in the respiratory epithelium, these data are of direct relevance to elucidating the molecular basis of normal lung function and lung disease.

show abstract

Conserved elements within first intron of aquaporin-5 (Aqp5) function as transcriptional enhancers

Cited by 8 publications

References 27 publications

Rat aquaporin-5 4.3-kb 5′-flanking region differentially regulates expression in salivary gland and lung in vivo

Rat aquaporin-5 4.3-kb 5′-flanking region differentially regulates expression in salivary gland and lung in vivo

Directed Expression of Cre in Alveolar Epithelial Type 1 Cells

A genome-wide analysis of open chromatin in human tracheal epithelial cells reveals novel candidate regulatory elements for lung function

Contact Info

Product

Resources

About