Tractable Cre-lox system for stochastic alteration of genes in mice

Abstract

We developed a cell division-activated Cre-lox system for stochastic recombination of loxP-flanked loci in mice. Cre activation by frameshift reversion is modulated by DNA mismatch-repair status and occurs in individual cells surrounded by normal tissue, mimicking spontaneous cancer-causing mutations. This system should be particularly useful for delineating pathways of neoplasia, and determining the developmental and aging consequences of specific gene alterations.

Figure 1. Generation and characterization of Pms2^cre mice

(a) A schematic of the Pms2^cre allele activation by a −1-bp frameshift in the 12A run. SA, splice acceptor; IRES, internal ribosome entry site. Boxes 1 and 3–5 represent exons. (b–f) β-galactosidase staining in whole-mount mouse intestinal sections. Different magnification images of Pms2^cre/+ (b,c) and Pms2^cre/cre (d–f) small intestine. Scale bars, 1 mm (d), 0.5 mm (b), 0.25 mm (c,e) and 0.1 mm (f). Arrow in b indicates the single spot in the field. (g–i) Hematoxylin and eosin–stained 5-μm paraffin sections of Pms2^cre/cre small intestine crypt. Scale bars, 50 μm (g) and 25 μm (h,i). (j) Distribution of average number of blue spots in the gastrointestinal tract of age-matched Pms2^cre/cre (n = 10) and Pms2^cre/+ (n = 4) mice. Error bars, s.d.

Figure 2. Activation of K-ras results in larger regions of β-galactosidase staining

(a) Distribution of average spots per region in Pms2^cre/cre, LSL-K-ras^G12D mice (n = 5). Error bars, s.d. The size spot per area increases compared to Pms2^cre/cre, Rosa26r mice. (b,c) β-galactosidase staining in whole-mount small intestine from Pms2^cre/cre, LSL-K-ras^G12D mice shows expanded (large) regions of staining. Scale bars, 1 mm (b) and 0.5 mm (c).