Pancreas-specific Cre driver lines and considerations for their prudent use

Abstract

Cre/LoxP has broad utility for studying the function, development, and oncogenic transformation of pancreatic cells in mice. Here we provide an overview of the Cre driver lines that are available for such studies. We discuss how variegated expression, transgene silencing, and recombination in undesired cell types have conspired to limit the performance of these lines, sometimes leading to serious experimental concerns. We also discuss preferred strategies for achieving high-fidelity driver lines and remind investigators of the continuing need for caution when interpreting results obtained from any Cre/LoxP-based experiment performed in mice.

Figure 1. Cre-mediated recombination in mouse tissues

Cre/LoxP can be used to conditionally eliminate or activate expression of genes. A. Conditional gene knockout. A so-called “floxed” allele is generated by using gene targeting to flank a coding exon of a gene of interest with two tandemly-oriented LoxP sites. Ideally, the codon that is floxed should be of a length that is not divisible by three since this will cause a frame-shift in the protein being encoded. Intercrossing of a Cre driver mouse with a floxed allele mouse will lead to the excision of flanked exon, and loss of a functional protein. B. Conditional gene activation. A “Lox-Stop-Lox” (LSL) allele is also generated by gene targeting. In this case a gene of interest (GOI) is engineered to lie downstream of an LSL cassette containing tandem LoxP sites flanking a selectable marker, usually neomycin, and multiple polyA signal sequences. Interbreeding of the LSL-GOI allele to a Cre driver mouse leads to activation of the GOI in a tissue-specific manner. This strategy is frequently used to derive reporter lines whose expression is activated by Cre.

Figure 2. Highly simplified scheme of pancreas development

The pancreas is an endodermally-derived organ that arises in a stepwise, progressive manner that involves the specification and differentiation of definitive endoderm (DE) into specific cell types. First, DE is specified into pre-pancreatic multipotent progenitor cells (MPCs). Second, the pancreatic MPCs are specified into either an acinar, ductal or endocrine cell lineage. Third, the endocrine cell lineage is further specified into five different endocrine cell types. On account of this progressive cellular differentiation, Cre driver lines that are expressed early in development will result in recombination across multiple cell lineages. In addition, genes that have been used to drive Cre expression in specific lineages and/or cell types are indicated in the boxes. For example, the expression of Ptf1a-Cre, since it is expressed in pancreatic MPCs during development, will result in a recombined allele in all three main pancreatic cell types in adult animals. Similarly, the expression of Ngn3-Cre in pre-endocrine progenitor cells will result in recombined allele being present in all five endocrine cell types.