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Fig. 1. Inducible lung epithelial-specific ablation of Rb. (A) Male mice heterozygous for both CC10-rtTA and tetCre transgenes, and homozygous for the floxed Rb gene allele (Rb^LoxP) were bred to female mice bearing a floxed Rb allele and a mutated null Rb allele (Rb^-). Dams were treated with doxycycline (ovals), which activates the rtTA (arches) expressed specifically in lung epithelium under control of the CC10 promoter. Activated rtTA induces Cre expression leading to excision and functional loss of Rb. (B) PCR analysis on lung or tail DNA obtained from day 1 pups taken from doxycycline-treated dams, or control dams not treated with doxycycline. Representative results are shown. Recombination at the floxed Rb allele (Rb^Rec) is dependent upon the presence of both CC10-rtTA and tetCre transgenes, and is detected in lung but not tail DNA. No recombination is detected in lung DNA obtained from control pups not treated with doxycycline. (C-F) Enzymatic staining for ß-gal (blue) in lungs from ROSA26 reporter mice harboring the CC10-rtTA and tetCre transgenes, and treated with doxycycline in utero. (C) Whole-mount staining of lungs from double transgenic day 1 pups (+) and control littermates (C) lacking one or both transgenes. (D,F) Staining of adult lung sections. Note that majority of ciliated and non-ciliated epithelial cells show staining. Inset in D highlights the scattered punctuate staining observed in the alveolar region, consistent with the localization of Type II cells. (E) Enzymatic staining for ß-gal (blue) followed by immunohistochemistry for CGRP (brown/black) in day 1 lung. Arrow marks a cell positive for both ß-gal activity and CGRP. br, bronchi/bronchioles; bv, blood vessel; a, alveoli.

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