An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology
Basal cells are multipotent stem cells responsible for the repair and regeneration of all the epithelial cell types present in the proximal lung. In mice, the elusive origins of basal cells and their contribution to lung development were recently revealed by high-resolution, lineage tracing studies....
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sg-ntu-dr.10356-1473052023-03-05T16:49:25Z An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology Goh, Kim Jee Tan, Ee Kim Lu, Hao Roy, Sudipto Dunn, Norris Ray Lee Kong Chian School of Medicine (LKCMedicine) Institute of Medical Biology, A*STAR Science::Medicine Cell Biology Developmental Biology Basal cells are multipotent stem cells responsible for the repair and regeneration of all the epithelial cell types present in the proximal lung. In mice, the elusive origins of basal cells and their contribution to lung development were recently revealed by high-resolution, lineage tracing studies. It however remains unclear if human basal cells originate and participate in lung development in a similar fashion, particularly with mounting evidence for significant species-specific differences in this process. To address this outstanding question, in the last several years differentiation protocols incorporating human pluripotent stem cells (hPSC) have been developed to produce human basal cells in vitro with varying efficiencies. To facilitate this endeavour, we introduced tdTomato into the human TP63 gene, whose expression specifically labels basal cells, in the background of a previously described hPSC line harbouring an NKX2-1GFP reporter allele. The functionality and specificity of the NKX2-1GFP;TP63tdTomato hPSC line was validated by directed differentiation into lung progenitors as well as more specialised lung epithelial subtypes using an organoid platform. This dual fluorescent reporter hPSC line will be useful for tracking, isolating and expanding basal cells from heterogenous differentiation cultures for further study. Agency for Science, Technology and Research (A*STAR) Nanyang Technological University Published version We are grateful to Professor Darrell Kotton, the National Center for Advancing Translational Sciences (NCATS; grant number: U01TR001810) and the Center for Regenerative Medicine for the BU3 NKX2-1GFP hiPSC line. We would also like to acknowledge the support of Seri Mustafah from the Agency for Science, Technology and Research Singapore (A*STAR) Flow Cytometry Core Facility for technical assistance with FACS sorting experiments. K.J.G, L.H. and S.R. are supported by A*STAR. E.K.T. and N.R.D. are supported by start-up funds provided by the Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore. 2021-03-26T08:20:44Z 2021-03-26T08:20:44Z 2021 Journal Article Goh, K. J., Tan, E. K., Lu, H., Roy, S. & Dunn, N. R. (2021). An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology. Scientific Reports, 11(1). https://dx.doi.org/10.1038/s41598-021-83825-6 2045-2322 https://hdl.handle.net/10356/147305 10.1038/s41598-021-83825-6 33633173 2-s2.0-85101746095 1 11 en Scientific Reports © 2021 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. application/pdf |
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Science::Medicine Cell Biology Developmental Biology Goh, Kim Jee Tan, Ee Kim Lu, Hao Roy, Sudipto Dunn, Norris Ray An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
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Basal cells are multipotent stem cells responsible for the repair and regeneration of all the epithelial cell types present in the proximal lung. In mice, the elusive origins of basal cells and their contribution to lung development were recently revealed by high-resolution, lineage tracing studies. It however remains unclear if human basal cells originate and participate in lung development in a similar fashion, particularly with mounting evidence for significant species-specific differences in this process. To address this outstanding question, in the last several years differentiation protocols incorporating human pluripotent stem cells (hPSC) have been developed to produce human basal cells in vitro with varying efficiencies. To facilitate this endeavour, we introduced tdTomato into the human TP63 gene, whose expression specifically labels basal cells, in the background of a previously described hPSC line harbouring an NKX2-1GFP reporter allele. The functionality and specificity of the NKX2-1GFP;TP63tdTomato hPSC line was validated by directed differentiation into lung progenitors as well as more specialised lung epithelial subtypes using an organoid platform. This dual fluorescent reporter hPSC line will be useful for tracking, isolating and expanding basal cells from heterogenous differentiation cultures for further study. |
author2 |
Lee Kong Chian School of Medicine (LKCMedicine) |
author_facet |
Lee Kong Chian School of Medicine (LKCMedicine) Goh, Kim Jee Tan, Ee Kim Lu, Hao Roy, Sudipto Dunn, Norris Ray |
format |
Article |
author |
Goh, Kim Jee Tan, Ee Kim Lu, Hao Roy, Sudipto Dunn, Norris Ray |
author_sort |
Goh, Kim Jee |
title |
An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
title_short |
An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
title_full |
An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
title_fullStr |
An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
title_full_unstemmed |
An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology |
title_sort |
nkx2-1gfp and tp63tdtomato dual fluorescent reporter for the investigation of human lung basal cell biology |
publishDate |
2021 |
url |
https://hdl.handle.net/10356/147305 |
_version_ |
1759858315274223616 |