title: Genomic DNA recombination with cell-penetrating peptide-tagged cre protein in mouse skeletal and cardiac muscle.
abstract: The Cre-loxP recombination system has been used to promote DNA recombination both in vitro and in vivo. For in vivo delivery, Cre expression is commonly achieved through the use of tissue/cell type-specific promoters, viral infection, or drug inducible transcription and protein translocation to promote targeted DNA excision. The development of cell permeable (or penetrating) peptide tagged proteins has facilitated the delivery of Cre recombinase protein into cells in culture, organotypic slide culture, or in living animals. In this report, we generated bacterially expressed, his-tagged Cre protein with either a cardiac targeting peptide or an antennapedia peptide at the C-terminus and demonstrated efficient uptake and recombination in both cell culture and mice. To facilitate delivery to cardiac and skeletal muscle, we mixed proteins with pluronic F-127 hydrogel and delivered Cre protein into reporter Rosa26mTmG mouse skeletal muscle or Rosa26LacZ cardiac muscle via ultrasound guided injection. Activation of reporter gene expression indicated that these Cre proteins were enzymatically active. Recombination events were detected only in the vicinity of injection areas. In conclusion, we have developed a method to deliver enzymatically active Cre protein locally to skeletal muscle and cardiac muscle that may be adapted for use with other proteins.
authors.affiliation: Division of Cardiology, Department of Medicine, University of Washington, Seattle, Washington.
authors.forename: Wei-Ming
authors.initials: WM
authors.lastname: Chien
authors.forename: Yonggang
authors.initials: Y
authors.lastname: Liu
authors.forename: Michael T
authors.initials: MT
authors.lastname: Chin
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D057846:Cell-Penetrating Peptides / Q000378:metabolism*; D018014:Gene Transfer Techniques; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018482:Muscle, Skeletal / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D011995:Recombination, Genetic
chemical_list: D057846:Cell-Penetrating Peptides; C045073:Cre recombinase; D019426:Integrases
keywords: Cre recombinase protein; antennapedia peptide; cardiac and skeletal muscle; cardiac targeting peptide; pluronic F-127