Mouse Cre-LoxP system: general principles to determine tissue-specific roles of target genes

Abstract

Genetically engineered mouse models are commonly preferred for studying the human disease due to genetic and pathophysiological similarities between mice and humans. In particular, Cre-loxP system is widely used as an integral experimental tool for generating the conditional. This system has enabled researchers to investigate genes of interest in a tissue/cell (spatial control) and/or time (temporal control) specific manner. A various tissue-specific Cre-driver mouse lines have been generated to date, and new Cre lines are still being developed. This review provides a brief overview of Cre-loxP system and a few commonly used promoters for expression of tissue-specific Cre recombinase. Also, we finally introduce some available links to the Web sites that provides detailed information about Cre mouse lines including their characterization.

Keywords: Cre portal sites; Cre-loxP system; tissue specific promoter.

Figure 1. Mechanism of Cre-loxP system. (A) An overview of Cre-loxP system. 38 kDa Cre recombinase recognizes the loxP sites of specific 34 bp DNA sequences. (B) General breeding strategy for conditional mutation using loxP and Cre driving mouse line. In principle, one mouse must have a tissue-specific driven cre gene and another mouse have loxP flanked (floxed) alleles of interest gene Y. Expression of Cre recombinase excises floxed loci and inactivates the gene Y.

Figure 2. Principles of Inducible Cre-loxP mutation system. (A) Tamoxifen (Tam)-inducible System of estrogen receptor fused to Cre (CreER). In the absence of tamoxifen, expressed fusion protein, CreER, interacts with heat shock protein 90 (HSP90) and exists in cytoplasm (1). Administration of Tam disrupts the interaction of HSP90 with CreER (2). Interaction of ER with Tam induces nuclear translocation of Cre (3). In the nucleus, the CreER recognizes the loxP sites (4) and inactivates the gene Y in tissue X (5). (B and C) Doxycycline (Dox)-induced Tetracyclin (Tet)-on and -off systems. (B) In Tet-on system, ubiquitous or tissue-specific promoter driven rtTA is expressed. In the absence of Dox, inactivated rtTA is unable to bind tetO₇ (7 repeats of a 19 nuclotide tetO minimal promoter, also referred to as TRE) sequence of cre gene. Cre is not expressed. Following Dox administration, Dox interacts with the rtTA and allows to activate. Activated rtTA binds to tetO₇ promoter of cre and induces the Cre expression. (C) In Tet-off system, in the absence of Dox, activated tTA is able to bind tetO₇ (TRE) sequence of cre and induces the Cre expression. Upon Dox administration, tTA interacted with Dox is inactivated. Inactivated rTA is not able to bind to tetO₇ promoter and therefore Cre expression is inhibited.