Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium

Abstract There is evidence that the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel is highly expressed at the apical pole of ciliated cells in human bronchial epithelium (HBE), however recent studies have detected little CFTR mRNA in those cells. To understand this discrepa...

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Autores principales:	Yukiko Sato, Kamila R. Mustafina, Yishan Luo, Carolina Martini, David Y. Thomas, Paul W. Wiseman, John W. Hanrahan
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/1f668e4b5e9b420599dbb88a96c392a3
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spelling	oai:doaj.org-article:1f668e4b5e9b420599dbb88a96c392a32021-12-05T12:14:36ZNonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium10.1038/s41598-021-02420-x2045-2322https://doaj.org/article/1f668e4b5e9b420599dbb88a96c392a32021-12-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-02420-xhttps://doaj.org/toc/2045-2322Abstract There is evidence that the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel is highly expressed at the apical pole of ciliated cells in human bronchial epithelium (HBE), however recent studies have detected little CFTR mRNA in those cells. To understand this discrepancy we immunostained well differentiated primary HBE cells using CFTR antibodies. We confirmed apical immunofluorescence in ciliated cells and quantified the covariance of the fluorescence signals and that of an antibody against the ciliary marker centrin-2 using image cross-correlation spectroscopy (ICCS). Super-resolution stimulated emission depletion (STED) imaging localized the immunofluorescence in distinct clusters at the bases of the cilia. However, similar apical fluorescence was observed when the monoclonal CFTR antibodies 596, 528 and 769 were used to immunostain ciliated cells expressing F508del-CFTR, or cells lacking CFTR due to a Class I mutation. A BLAST search using the CFTR epitope identified a similar amino acid sequence in the ciliary protein rootletin X1. Its expression level correlated with the intensity of immunostaining by CFTR antibodies and it was detected by 596 antibody after transfection into CFBE cells. These results may explain the high apparent expression of CFTR in ciliated cells and reports of anomalous apical immunofluorescence in well differentiated cells that express F508del-CFTR.Yukiko SatoKamila R. MustafinaYishan LuoCarolina MartiniDavid Y. ThomasPaul W. WisemanJohn W. HanrahanNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-15 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Yukiko Sato Kamila R. Mustafina Yishan Luo Carolina Martini David Y. Thomas Paul W. Wiseman John W. Hanrahan Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
description	Abstract There is evidence that the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel is highly expressed at the apical pole of ciliated cells in human bronchial epithelium (HBE), however recent studies have detected little CFTR mRNA in those cells. To understand this discrepancy we immunostained well differentiated primary HBE cells using CFTR antibodies. We confirmed apical immunofluorescence in ciliated cells and quantified the covariance of the fluorescence signals and that of an antibody against the ciliary marker centrin-2 using image cross-correlation spectroscopy (ICCS). Super-resolution stimulated emission depletion (STED) imaging localized the immunofluorescence in distinct clusters at the bases of the cilia. However, similar apical fluorescence was observed when the monoclonal CFTR antibodies 596, 528 and 769 were used to immunostain ciliated cells expressing F508del-CFTR, or cells lacking CFTR due to a Class I mutation. A BLAST search using the CFTR epitope identified a similar amino acid sequence in the ciliary protein rootletin X1. Its expression level correlated with the intensity of immunostaining by CFTR antibodies and it was detected by 596 antibody after transfection into CFBE cells. These results may explain the high apparent expression of CFTR in ciliated cells and reports of anomalous apical immunofluorescence in well differentiated cells that express F508del-CFTR.
format	article
author	Yukiko Sato Kamila R. Mustafina Yishan Luo Carolina Martini David Y. Thomas Paul W. Wiseman John W. Hanrahan
author_facet	Yukiko Sato Kamila R. Mustafina Yishan Luo Carolina Martini David Y. Thomas Paul W. Wiseman John W. Hanrahan
author_sort	Yukiko Sato
title	Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
title_short	Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
title_full	Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
title_fullStr	Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
title_full_unstemmed	Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium
title_sort	nonspecific binding of common anti-cftr antibodies in ciliated cells of human airway epithelium
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/1f668e4b5e9b420599dbb88a96c392a3
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Nonspecific binding of common anti-CFTR antibodies in ciliated cells of human airway epithelium

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