Since the discovery of the CFTR gene over a decade ago, Cystic Fibrosis (CF) has
been regarded as amenable to intervention by gene therapy. The ultimate aim of gene
therapy must be the correction, within cells capable of repopulating the tissue, of the
genetic defect in its chromosomal context. Towards that end, a mouse model
designed to evaluate the efficiency of gene correction was created, and a transgenic
approach was taken to the investigation of a putative progenitor cell population in the
adult murine respiratory tract.
Before gene correction systems can be considered as valid therapeutic agents, their
utility in the cells and tissues of living animals must be demonstrated. Thus, an in
vivo system permitting the simple quantification of correction frequency in a wide
range of tissues would be a valuable resource for the gene correction community.
The generation and analysis of a transgenic mouse carrying an inactivated, but
potentially correctable, reporter transgene is described.
The full potential of a gene correction strategy to provide a single-dose, permanent
solution to a genetically-diseased tissue will only be realised once the therapy is able
to target resident stem cells. For CF lung disease, this will require the prior
identification of stem cells in the respiratory epithelium. Previous work has indicated
that potential stem cells are spatially coincident with small groups of cells expressing
high levels of keratin 5 (K5) protein in the proximal murine trachea. In order to
investigate lineage arising from this putative stem cell niche, transgenic mice have
been generated which express an inducible form of Cre recombinase from the K5
promoter. Preliminary experiments demonstrate recombination of a conditional
reporter gene after induction of Cre activity in K5-expressing tissue. Comparison of
the inducible system with a constitutive K5 promoter-driven Cre line validated the
choice of the former, as the clarity of data obtained from the conventional system
was undermined as a result of K5 promoter activity causing reporter gene activation
prior to the onset ofthe experiment.
In the course of these studies it became evident that the conventional, constitutive
Cre line gave rise to segregating patterns of reporter gene activation. While some
mice displayed the expected K5-derived expression profile, other animals
demonstrated ubiquitous expression. Universal activation of the conditional reporter
was detected only in animals derived from females carrying the Cre transgene, and
was found to be the result of unanticipated production of Cre protein in the maternal
germline. This transgenic line is unusual and valuable in offering a choice of tissuespecific and generalised recombination of floxed alleles.