Use of Cre-adenovirus and CAR transgenic mice for efficient deletion of genes in post-thymic T cells.
Conditional gene deletion using lineage-specific transgenic expression of Cre has been useful for defining the role of specific gene products in mice in vivo. However, this technology has had limitations for studies of peripheral T cell biology, since the T-lineage promoters commonly used are active early in thymic development. As such, T cell development can be altered by the resulting genetic alterations, thus limiting the interpretation of the data in post-thymic T cell studies. Thus, new strategies are needed to delete targeted genes directly in peripheral T lymphocytes. The availability of transgenic mice expressing the CAR in the T cell compartment enabled testing of Cre-mediated recombination using an adenoviral vector in naïve peripheral T cells in vitro, even without cellular activation. Using Rosa26R reporterxCAR transgenic mice, we describe conditions by which Cre-mediated deletion of targeted genes can be achieved with primary T cells in vitro. These cells can also be adoptively transferred into defined recipient mice for study in vivo. We use conditional PTEN-deficient mice as proof of concept to confirm the value of this technique for deleting a negative regulator of T cell activation. This technology should be broadly applicable for studies of T cell-specific gene deletion to gain understanding of function in the post-thymic T cell compartment.
Zha
Yuanyuan
Y
Department of Pathology and Department of Medicine, University of Chicago, Chicago, IL 60637, USA.
Shah
Ramila
R
Locke
Frederick
F
Wong
Austin
A
Gajewski
Thomas F
TF
D000256:Adenoviridae; D000818:Animals; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D017353:Gene Deletion; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors; D051379:Mice; D008822:Mice, Transgenic; D051059:PTEN Phosphohydrolase / Q000172:deficiency / Q000235:genetics*; D011506:Proteins / Q000235:genetics*; D022661:RNA, Untranslated; D011991:Receptors, Virus / Q000235:genetics; D011995:Recombination, Genetic; D013601:T-Lymphocytes / Q000502:physiology*; D014161:Transduction, Genetic
C568772:CLMP protein, mouse; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; C110853:Gt(ROSA)26Sor non-coding RNA, mouse; D011506:Proteins; D022661:RNA, Untranslated; D011991:Receptors, Virus; D051059:PTEN Phosphohydrolase; C487757:Pten protein, mouse