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Tunable Protein Stabilization In Vivo Mediated by Shield-1 in Transgenic Medaka.

Froschauer A, Kube L, Kegler A, Rieger C, Gutzeit HO - PLoS ONE (2015)

Bottom Line: Techniques for conditional gene or protein expression are important tools in developmental biology and in the analysis of physiology and disease.We present the application of the DD-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life.Shield-1 administration to the water leads to concentration-dependent induction of a YFP reporter gene in various organs and in spermatogonia at the cellular level.

View Article: PubMed Central - PubMed

Affiliation: Institute of Zoology, Technische Universität Dresden, Dresden, Germany.

ABSTRACT
Techniques for conditional gene or protein expression are important tools in developmental biology and in the analysis of physiology and disease. On the protein level, the tunable and reversible expression of proteins can be achieved by the fusion of the protein of interest to a destabilizing domain (DD). In the absence of its specific ligand (Shield-1), the protein is degraded by the proteasome. The DD-Shield system has proven to be an excellent tool to regulate the expression of proteins of interests in mammalian systems but has not been applied in teleosts like the medaka. We present the application of the DD-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life. Shield-1 administration to the water leads to concentration-dependent induction of a YFP reporter gene in various organs and in spermatogonia at the cellular level.

No MeSH data available.


Related in: MedlinePlus

Induction of YFP in adult medaka.The fluorescence of non-transgenic adult fish and of transgenic individuals of Ola-Tg(actin-DD-YFP)13 was photographed after the indicated treatment with vehicle only (0.1% ethanol) or 1 μM Shield-1, respectively. (A) Adult females are shown as combinations of 2 overlapping photographs in toto; exposure time: 13 sec, scale bar: 5 mm. (B) Brains of the same individuals as in A; exposure time: 21 sec, scale bar: 1 mm. (C) Testes of adult males show the strongest fluorescence at the outer area at the end of the tubules (arrows); exposure time: 13 sec, scale bar 1 mm.
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pone.0131252.g003: Induction of YFP in adult medaka.The fluorescence of non-transgenic adult fish and of transgenic individuals of Ola-Tg(actin-DD-YFP)13 was photographed after the indicated treatment with vehicle only (0.1% ethanol) or 1 μM Shield-1, respectively. (A) Adult females are shown as combinations of 2 overlapping photographs in toto; exposure time: 13 sec, scale bar: 5 mm. (B) Brains of the same individuals as in A; exposure time: 21 sec, scale bar: 1 mm. (C) Testes of adult males show the strongest fluorescence at the outer area at the end of the tubules (arrows); exposure time: 13 sec, scale bar 1 mm.

Mentions: Adult male and female individuals were treated with different concentrations of Shield-1 and analyzed after 24 hours. The induced fluorescence was visible in toto, in brain and in gonads (Fig 3 and S3 Fig). The induction of YFP in brain (Fig 3B) shows that Shield-1 effectively passes the blood brain barrier. Other tissues like gills, gut, liver, kidney, spleen and heart also responded to the treatment (not shown). We did not compare the fluorescence levels between different organs due to their differences in cellular composition, thickness and autofluorescence, but the induction was dose dependent in all organs.


Tunable Protein Stabilization In Vivo Mediated by Shield-1 in Transgenic Medaka.

Froschauer A, Kube L, Kegler A, Rieger C, Gutzeit HO - PLoS ONE (2015)

Induction of YFP in adult medaka.The fluorescence of non-transgenic adult fish and of transgenic individuals of Ola-Tg(actin-DD-YFP)13 was photographed after the indicated treatment with vehicle only (0.1% ethanol) or 1 μM Shield-1, respectively. (A) Adult females are shown as combinations of 2 overlapping photographs in toto; exposure time: 13 sec, scale bar: 5 mm. (B) Brains of the same individuals as in A; exposure time: 21 sec, scale bar: 1 mm. (C) Testes of adult males show the strongest fluorescence at the outer area at the end of the tubules (arrows); exposure time: 13 sec, scale bar 1 mm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4493054&req=5

pone.0131252.g003: Induction of YFP in adult medaka.The fluorescence of non-transgenic adult fish and of transgenic individuals of Ola-Tg(actin-DD-YFP)13 was photographed after the indicated treatment with vehicle only (0.1% ethanol) or 1 μM Shield-1, respectively. (A) Adult females are shown as combinations of 2 overlapping photographs in toto; exposure time: 13 sec, scale bar: 5 mm. (B) Brains of the same individuals as in A; exposure time: 21 sec, scale bar: 1 mm. (C) Testes of adult males show the strongest fluorescence at the outer area at the end of the tubules (arrows); exposure time: 13 sec, scale bar 1 mm.
Mentions: Adult male and female individuals were treated with different concentrations of Shield-1 and analyzed after 24 hours. The induced fluorescence was visible in toto, in brain and in gonads (Fig 3 and S3 Fig). The induction of YFP in brain (Fig 3B) shows that Shield-1 effectively passes the blood brain barrier. Other tissues like gills, gut, liver, kidney, spleen and heart also responded to the treatment (not shown). We did not compare the fluorescence levels between different organs due to their differences in cellular composition, thickness and autofluorescence, but the induction was dose dependent in all organs.

Bottom Line: Techniques for conditional gene or protein expression are important tools in developmental biology and in the analysis of physiology and disease.We present the application of the DD-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life.Shield-1 administration to the water leads to concentration-dependent induction of a YFP reporter gene in various organs and in spermatogonia at the cellular level.

View Article: PubMed Central - PubMed

Affiliation: Institute of Zoology, Technische Universität Dresden, Dresden, Germany.

ABSTRACT
Techniques for conditional gene or protein expression are important tools in developmental biology and in the analysis of physiology and disease. On the protein level, the tunable and reversible expression of proteins can be achieved by the fusion of the protein of interest to a destabilizing domain (DD). In the absence of its specific ligand (Shield-1), the protein is degraded by the proteasome. The DD-Shield system has proven to be an excellent tool to regulate the expression of proteins of interests in mammalian systems but has not been applied in teleosts like the medaka. We present the application of the DD-Shield technique in transgenic medaka and show the ubiquitous conditional expression throughout life. Shield-1 administration to the water leads to concentration-dependent induction of a YFP reporter gene in various organs and in spermatogonia at the cellular level.

No MeSH data available.


Related in: MedlinePlus