The CCAAT/enhancer binding proteins (C/EBPs) are a family of basic leucine zipper (B-ZIP) transcription factors that play important roles in cellular differentiation, proliferation, survival, and apoptosis, and metabolism, inflammation, and transformation [1–3]. Six C/EBP family members have been identified that share the N-terminal basic amino acid-containing region necessary for DNA binding and the highly conservative C-terminal leucine zipper (B-ZIP) dimerization motif [4, 5]. Through the B-ZIP domain, they can homo- and/or hetero-dimerize with each other to bind specific DNA sequences [4, 5].
Among the family members, the most studied are C/EBPα and C/EBPβ. C/EBPα is responsible for blocking proliferation, promoting differentiation and suppressing tumorigenesis, thus being considered as a tumor suppressor. Significant down-regulation of C/EBPα expression is found in cancers of various tissues such as mammary gland , lung , and the epidermis . Further, specific somatic mutations are found in ~10% of acute myeloid leukemia (AML) patients [9, 10]. The reintroduction of C/EBPα blocked the in vivo tumorigenicity of AML and skin carcinogenesis [11, 12]. In contrast, C/EBPβ is more complex, but has been implicated in playing a role in tumorigenesis, depending on the cellular context and C/EBPβ forms present, out of three translational isoforms . In particular, one of the C/EBPβ isoforms, the C/EBP-liver-enriched inhibitory protein acts as dominant-negative, and the increased expression inhibits the transcriptional activation of genes involved in differentiation, while its over-expression is found in breast cancers [13, 14].
In the lung, three C/EBPs, C/EBPα, C/EBPβ, and C/EBPδ, are highly expressed with various degrees of expression depending on protein subtypes, cell types, and/or developmental stages [15–18]. Similar to tumors of other organs, C/EBPα is described as a tumor suppressor in lung cancer [7, 19], while C/EBPβ regulates the expression of Matrix Metalloproteinase (MMP) 1 that mediates extracellular matrix remodeling and promotes tumor invasion . The role of C/EBPδ in lung carcinogenesis is not clear. Because of multiple forms of C/EBPs expressed in the lung, how C/EBPs as a whole contribute to lung carcinogenesis is not known.
A-C/EBP is a dominant negative form of C/EBP that contains leucine zipper dimerization domain and an acidic region, replacing the basic region of C/EBPs. A-C/EBP heterodimerizes with all C/EBP family members through the leucine zipper region of the B-ZIP, and the acidic region forms coiled coil structure to stabilize the heterodimer, thus blocking the DNA binding of C/EBPs . A transgenic mouse line (TetO-A-C/EBP) was established that expresses the A-C/EBP dominant negative gene  under the regulated control of the tetracycline operon (TetO) promoter . The TetO-A-C/EBP mice were further crossed with transgenic mice CCSP-rtTA that express the rtTA under the promoter of lung epithelial Clara cell-specific CCSP (SCGB1A1) gene . The progeny of this cross, named CCSP-rtTA;TetO-A-C/EBP mice were previously shown to highly express A-C/EBP in the presence of doxycycline and suppress the transcription of C/EBP-responsive genes such as secretoglobin 3a2 in lung [18, 22].
In this study, CCSP-rtTA;TetO-A-C/EBP mice were subjected to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung carcinogenesis bioassay  in the presence and absence of doxycycline, and the effect of A-C/EBP expression on the development and/or progression of lung cancer was examined.