Characterization of bacterial artificial chromosome transgenic mice expressing mCherry fluorescent protein substituted for the murine smooth muscle alpha-actin gene.
Smooth muscle alpha actin (SMA) is a cytoskeletal protein expressed by mesenchymal and smooth muscle cell types, including mural cells (vascular smooth muscle cells and pericytes). Using Bacterial Artificial Chromosome (BAC) recombineering technology, we generated transgenic reporter mice that express a membrane localized cherry red fluorescent protein (mCherry), driven by the full-length SMA promoter and intronic sequences. We determined that the founders and F1 progeny of five independent lines contain 1-3 copies of the mCherry-substituted BAC vector. Furthermore, we characterized the expression of SMA-mCherry in relation to endogenous SMA in the embryo and in adult tissues, and found that the transgenic reporter in each line recapitulated endogenous SMA expression at all time points. We were also able to isolate SMA expressing cells from embryonic tissues using fluorescence-activated cell sorting (FACS). We demonstrated that this marker can be combined with other vital fluorescent reporters and it can be used for live imaging of embryonic cardiodynamics. Therefore, these transgenic mice will be useful for isolating live SMA-expressing cells via FACS and for studying the emergence, behavior, and regulation of SMA-expressing cells, including vascular smooth muscle cells and pericytes throughout embryonic and postnatal development.
Armstrong
John J
JJ
Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030, USA.
Larina
Irina V
IV
Dickinson
Mary E
ME
Zimmer
Warren E
WE
Hirschi
Karen K
KK
D000199:Actins / Q000235:genetics*; D000367:Age Factors; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D004622:Embryo, Mammalian / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins; D006321:Heart / Q000196:embryology / Q000502:physiology*; D008164:Luminescent Proteins; D051379:Mice; D008822:Mice, Transgenic; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D020286:Pericytes / Q000166:cytology; D011401:Promoter Regions, Genetic / Q000235:genetics; D013050:Spectrometry, Fluorescence; D000097573:Red Fluorescent Protein
D000199:Actins; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein