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Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter

Mouse models that combine specific loxP-flanked gene sequences with Cre recombinase expressed from cell-regulated promoters have become important tools to investigate gene function. Critically however, expression of Cre recombinase may not always be restricted to the target cell or tissue of interes...

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Autores principales: Bittner-Eddy, Peter D., Fischer, Lori A., Costalonga, Massimo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763954/
https://www.ncbi.nlm.nih.gov/pubmed/31616418
http://dx.doi.org/10.3389/fimmu.2019.02228
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author Bittner-Eddy, Peter D.
Fischer, Lori A.
Costalonga, Massimo
author_facet Bittner-Eddy, Peter D.
Fischer, Lori A.
Costalonga, Massimo
author_sort Bittner-Eddy, Peter D.
collection PubMed
description Mouse models that combine specific loxP-flanked gene sequences with Cre recombinase expressed from cell-regulated promoters have become important tools to investigate gene function. Critically however, expression of Cre recombinase may not always be restricted to the target cell or tissue of interest due to promiscuous activity of the driving promoter. Expression of Cre recombinase and, by extension, excision of the loxP-flanked gene may occur in non-target cells and may not be readily apparent. Here we report on the fidelity of Cre recombinase expressed from the il17a or Foxp3 promoters by combining them with a constitutively expressed floxed-stopped tdTomato reporter gene. Foxp3-driven Cre recombinase in F(1) mice induced tdTomato red fluorescent protein in Treg cells but also in a range of other immune cells. Frequency of tdTomato expression was variable but positively correlated (p < 0.0001) amongst lymphoid (B cells and CD8 T cells) and blood-resident myeloid cells (dendritic cells, monocytes, neutrophils) suggesting stochastic activity of the Foxp3 promoter rather than developmental regulation in common ancestral progenitors. Interestingly, frequency of tdTomato(+) dendritic cells, monocytes and neutrophils did not correlate with the tdTomato(+) fraction in eosinophils, indicating that activity of the Foxp3 promoter in eosinophils occurred after the split from a common multipotent progenitor. When these F(1) mice were crossed to achieve homozygosity of the promoter and reporter gene, a novel visually red phenotype was observed segregating amongst littermates. The red coloration was widespread and prevalent in non-immune tissues. Thymocytes examined from these red mice showed that all four subsets of immature thymocytes (CD4(−) CD8(−)) based on differential expression of CD25 and CD44 were expressing tdTomato. Finally, we show evidence of Foxp3 Cre recombinase independent tdTomato expression, suggesting germ line transmission of an activated tdTomato reporter gene. Our data highlights potential issues with conclusions drawn from using specifically the B6.129(Cg)-Foxp3(tm4(YFP/Cre)Ayr)/J mice.
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spelling pubmed-67639542019-10-15 Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter Bittner-Eddy, Peter D. Fischer, Lori A. Costalonga, Massimo Front Immunol Immunology Mouse models that combine specific loxP-flanked gene sequences with Cre recombinase expressed from cell-regulated promoters have become important tools to investigate gene function. Critically however, expression of Cre recombinase may not always be restricted to the target cell or tissue of interest due to promiscuous activity of the driving promoter. Expression of Cre recombinase and, by extension, excision of the loxP-flanked gene may occur in non-target cells and may not be readily apparent. Here we report on the fidelity of Cre recombinase expressed from the il17a or Foxp3 promoters by combining them with a constitutively expressed floxed-stopped tdTomato reporter gene. Foxp3-driven Cre recombinase in F(1) mice induced tdTomato red fluorescent protein in Treg cells but also in a range of other immune cells. Frequency of tdTomato expression was variable but positively correlated (p < 0.0001) amongst lymphoid (B cells and CD8 T cells) and blood-resident myeloid cells (dendritic cells, monocytes, neutrophils) suggesting stochastic activity of the Foxp3 promoter rather than developmental regulation in common ancestral progenitors. Interestingly, frequency of tdTomato(+) dendritic cells, monocytes and neutrophils did not correlate with the tdTomato(+) fraction in eosinophils, indicating that activity of the Foxp3 promoter in eosinophils occurred after the split from a common multipotent progenitor. When these F(1) mice were crossed to achieve homozygosity of the promoter and reporter gene, a novel visually red phenotype was observed segregating amongst littermates. The red coloration was widespread and prevalent in non-immune tissues. Thymocytes examined from these red mice showed that all four subsets of immature thymocytes (CD4(−) CD8(−)) based on differential expression of CD25 and CD44 were expressing tdTomato. Finally, we show evidence of Foxp3 Cre recombinase independent tdTomato expression, suggesting germ line transmission of an activated tdTomato reporter gene. Our data highlights potential issues with conclusions drawn from using specifically the B6.129(Cg)-Foxp3(tm4(YFP/Cre)Ayr)/J mice. Frontiers Media S.A. 2019-09-20 /pmc/articles/PMC6763954/ /pubmed/31616418 http://dx.doi.org/10.3389/fimmu.2019.02228 Text en Copyright © 2019 Bittner-Eddy, Fischer and Costalonga. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Immunology
Bittner-Eddy, Peter D.
Fischer, Lori A.
Costalonga, Massimo
Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title_full Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title_fullStr Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title_full_unstemmed Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title_short Cre-loxP Reporter Mouse Reveals Stochastic Activity of the Foxp3 Promoter
title_sort cre-loxp reporter mouse reveals stochastic activity of the foxp3 promoter
topic Immunology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763954/
https://www.ncbi.nlm.nih.gov/pubmed/31616418
http://dx.doi.org/10.3389/fimmu.2019.02228
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