Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites

Abstract Inhibitory G proteins (Gi proteins) are highly homologous but play distinct biological roles. However, their isoform-specific detection remains challenging. To facilitate the analysis of Gαi3 expression, we generated a Gnai3- iresGFP reporter mouse line. An internal ribosomal entry site...

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Autores principales:	Veronika Leiss, Ellen Reisinger, Annika Speidel, Sandra Beer-Hammer, Bernd Nürnberg
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/109c206beb054c498b8de76fdeab599d
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spelling	oai:doaj.org-article:109c206beb054c498b8de76fdeab599d2021-12-02T16:14:17ZAnalyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites10.1038/s41598-021-93591-02045-2322https://doaj.org/article/109c206beb054c498b8de76fdeab599d2021-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-93591-0https://doaj.org/toc/2045-2322Abstract Inhibitory G proteins (Gi proteins) are highly homologous but play distinct biological roles. However, their isoform-specific detection remains challenging. To facilitate the analysis of Gαi3 expression, we generated a Gnai3- iresGFP reporter mouse line. An internal ribosomal entry site (IRES) was inserted behind the stop-codon of the Gnai3 gene to initiate simultaneous translation of the GFP cDNA together with Gαi3. The expression of GFP was confirmed in spleen and thymus tissue by immunoblot analysis. Importantly, the GFP knock-in (ki) did not alter Gαi3 expression levels in all organs tested including spleen and thymus compared to wild-type littermates. Flow cytometry of thymocytes, splenic and blood cell suspensions revealed significantly higher GFP fluorescence intensities in homozygous ki/ki animals compared to heterozygous mice (+/ki). Using cell-type specific surface markers GFP fluorescence was assigned to B cells, T cells, macrophages and granulocytes from both splenic and blood cells and additionally blood-derived platelets. Moreover, immunofluorescent staining of the inner ear from knock-in mice unraveled GFP expression in sensory and non-sensory cell types, with highest levels in Deiter’s cells and in the first row of Hensen’s cells in the organ of Corti, indicating a novel site for Gαi3 expression. In summary, the Gnai3- iresGFP reporter mouse represents an ideal tool for precise analyses of Gαi3 expression patterns and sites.Veronika LeissEllen ReisingerAnnika SpeidelSandra Beer-HammerBernd NürnbergNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Veronika Leiss Ellen Reisinger Annika Speidel Sandra Beer-Hammer Bernd Nürnberg Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
description	Abstract Inhibitory G proteins (Gi proteins) are highly homologous but play distinct biological roles. However, their isoform-specific detection remains challenging. To facilitate the analysis of Gαi3 expression, we generated a Gnai3- iresGFP reporter mouse line. An internal ribosomal entry site (IRES) was inserted behind the stop-codon of the Gnai3 gene to initiate simultaneous translation of the GFP cDNA together with Gαi3. The expression of GFP was confirmed in spleen and thymus tissue by immunoblot analysis. Importantly, the GFP knock-in (ki) did not alter Gαi3 expression levels in all organs tested including spleen and thymus compared to wild-type littermates. Flow cytometry of thymocytes, splenic and blood cell suspensions revealed significantly higher GFP fluorescence intensities in homozygous ki/ki animals compared to heterozygous mice (+/ki). Using cell-type specific surface markers GFP fluorescence was assigned to B cells, T cells, macrophages and granulocytes from both splenic and blood cells and additionally blood-derived platelets. Moreover, immunofluorescent staining of the inner ear from knock-in mice unraveled GFP expression in sensory and non-sensory cell types, with highest levels in Deiter’s cells and in the first row of Hensen’s cells in the organ of Corti, indicating a novel site for Gαi3 expression. In summary, the Gnai3- iresGFP reporter mouse represents an ideal tool for precise analyses of Gαi3 expression patterns and sites.
format	article
author	Veronika Leiss Ellen Reisinger Annika Speidel Sandra Beer-Hammer Bernd Nürnberg
author_facet	Veronika Leiss Ellen Reisinger Annika Speidel Sandra Beer-Hammer Bernd Nürnberg
author_sort	Veronika Leiss
title	Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
title_short	Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
title_full	Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
title_fullStr	Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
title_full_unstemmed	Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites
title_sort	analyses of gnai3-iresgfp reporter mice reveal unknown gαi3 expression sites
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/109c206beb054c498b8de76fdeab599d
work_keys_str_mv	AT veronikaleiss analysesofgnai3iresgfpreportermicerevealunknowngai3expressionsites AT ellenreisinger analysesofgnai3iresgfpreportermicerevealunknowngai3expressionsites AT annikaspeidel analysesofgnai3iresgfpreportermicerevealunknowngai3expressionsites AT sandrabeerhammer analysesofgnai3iresgfpreportermicerevealunknowngai3expressionsites AT berndnurnberg analysesofgnai3iresgfpreportermicerevealunknowngai3expressionsites
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Analyses of Gnai3-iresGFP reporter mice reveal unknown Gαi3 expression sites

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