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Temporal control of the Cre recombinase in transgenic mice by a tetracycline responsive promoter
L St-Onge1, P A Furth, P Gruss
1Department of Molecular Cell Biology, Max-Planck-Institute for Biophysical Chemistry, Göttingen, Germany.
Abstract:
Gene-targeted mice derived from embryonic stem cells are a useful tool to study gene function during development. However, if the mutation is embryonic lethal and the gene is deleted from the onset of development, later functions in adult animals cannot be studied. Recently, the bacterial Cre-loxP site-specific recombination system has successfully been used in transgenic animals to produce tissue-specific and temporal deletions [Gu et al. (1993) Cell, 73, 1155"""""" Gu et al. (1994) Science, 265,103--106; Kuhn et al. (1995) Science, 269, 1427-1429]. We have evaluated the tetracycline responsive binary system [Gossen and Bujard (1992) Proc. Natl. Acad. Sci. USA, 89, 5547-5551] for its ability to transiently express the Cre recombinase in transgenic mice. In this system, a transactivator fusion protein composed of the tetracycline repressor (tetR) and the acidic domain of the herpes simplex viral protein 16 (VP16) can regulate the expression of the Cre gene from a promoter containing tet-operator (tetO) sequences. In the absence of tetracycline, the Cre gene is expressed and will induce site-specific recombination between two loxP sites. In the presence of tetracycline, the Cre gene will not be expressed and recombination will not occur.
Insights
Researchers developed a novel method using the tetracycline-inducible system to control Cre recombinase expression in transgenic mice. This allows for temporal control of gene deletion, enabling the study of gene functions in adult animals.
Area of Science:
- Genetics
- Molecular Biology
- Developmental Biology
Background:
- Gene-targeted mice are crucial for studying gene function during development.
- Embryonic lethal mutations prevent the study of later gene functions in adult animals.
- The Cre-loxP system allows for tissue-specific and temporal gene deletions in transgenic animals.
Purpose of the Study:
- To evaluate the tetracycline-responsive binary system for transient Cre recombinase expression in transgenic mice.
- To enable temporal control over gene deletions, overcoming limitations of embryonic lethal mutations.
- To facilitate the study of gene functions in adult animals.
Main Methods:
- Utilized a tetracycline-responsive binary system to regulate Cre recombinase expression.
- Employed a transactivator fusion protein (tetracycline repressor-VP16) to control Cre gene expression from a tet-operator-containing promoter.
- Manipulated tetracycline levels to control Cre gene expression and subsequent loxP site-specific recombination.
Main Results:
- Demonstrated that the tetracycline-responsive system can transiently express Cre recombinase in transgenic mice.
- Showed that Cre gene expression and recombination are regulated by the presence or absence of tetracycline.
- Established a method for temporal control of gene deletion using the Cre-loxP system.
Conclusions:
- The tetracycline-inducible system provides a powerful tool for temporal control of Cre recombinase activity in transgenic mice.
- This approach enables the study of gene functions that are essential for embryonic development but also play roles in adult animals.
- Facilitates research into gene function by allowing conditional gene inactivation at specific times.