The armadillo repeat domain of Apc suppresses intestinal tumorigenesis.

The adenomatous polyposis coli (APC) gene is known to act as a tumor suppressor gene in both sporadic and hereditary colorectal cancer by negatively regulating WNT signaling. Familial adenomatous polyposis (FAP) patients develop intestinal polyps due to the presence of a single germline mutation in APC. The severity of the FAP phenotype is a function of the position of the APC mutation, indicating a complex role for APC that extends beyond the canonical WNT pathway. APC encodes a large protein with multiple functional domains, including an armadillo repeat domain that has been linked to protein-protein interactions. To determine the effect of the armadillo repeat domain on intestinal tumorigenesis, we generated a congenic mouse line (Apc ( Δ242 )) carrying a gene trap cassette between exons 7 and 8 of the murine Apc gene. Apc ( Δ242/+) mice express a truncated Apc product lacking the armadillo repeat domain as part of a fusion protein with ß-geo. Expression of the fusion product was confirmed by X-gal staining, ensuring that Apc ( Δ242 ) is not a null allele. In contrast, Apc ( Min/+) mice produce a truncated Apc product that contains an intact armadillo repeat domain. On the C57BL/6J background, Apc ( Δ242/+) mice develop more polyps than do Apc ( Min/+) mice along the entire length of the small intestine; however, polyps were significantly smaller in Apc ( Δ242/+) mice. In addition, polyp multiplicity in Apc ( Δ242/+) mice is affected by polymorphisms between inbred strains. These data suggest that the armadillo repeat domain of the Apc protein suppresses tumor initiation in the murine intestine while also promoting tumor growth.

Guanylyl cyclase C suppresses intestinal tumorigenesis by restricting proliferation and maintaining genomic integrity.

BACKGROUND AND AIMS: The most commonly lost gene products in colorectal carcinogenesis include guanylin and uroguanylin, endogenous ligands for guanylyl cyclase C (GCC). Beyond intestinal fluid balance, GCC mediates diarrhea induced by bacterial enterotoxins, and an inverse relationship exists between enterotoxigenic Escherichia coli infections producing the exogenous GCC ligand ST and colorectal cancer worldwide. However, the role of GCC in neoplasia remains obscure. METHODS: Intestinal tumorigenesis was examined in wild-type (Gcc(+/+)) and GCC-deficient (Gcc(-/-)) mice carrying mutations in Apc (Apc(Min/+)) or exposed to the carcinogen azoxymethane. Markers of DNA damage, loss of Apc heterozygosity, and beta-catenin mutations were used to assess genomic integrity. Hyperproliferation was explored using Ki67 and cell cycle markers. Apoptosis was quantified by transferase biotin-dUTP nick end labeling analysis. RESULTS: In colons of Apc(Min/+) mice, deletion of Gcc increased tumor incidence and multiplicity, reflecting uncoupling of loss of genomic integrity and compensatory apoptosis. Conversely, in the small intestine, elimination of Gcc increased tumorigenesis by enhancing proliferation without altering genomic integrity. Moreover, these distinct but mutually reinforcing mechanisms collaborate in azoxymethane-exposed mice, and deletion of Gcc increased tumor initiation and growth associated with hypermutation and hyperproliferation, respectively, in conjunction with attenuated apoptosis. CONCLUSIONS: GCC suppresses tumor initiation and growth by maintaining genomic integrity and restricting proliferation. This previously unrecognized role of GCC in inhibiting tumorigenesis, together with the invariant disruption in guanylin and uroguanylin expression early in carcinogenesis, and the uniform over-expression of GCC by tumors, underscores the potential of oral administration of GCC ligands for targeted prevention and therapy of colorectal cancer.

The modifier of Min 2 (Mom2) locus: embryonic lethality of a mutation in the Atp5a1 gene suggests a novel mechanism of polyp suppression.

Inactivation of the APC gene is considered the initiating event in human colorectal cancer. Modifier genes that influence the penetrance of mutations in tumor-suppressor genes hold great potential for preventing the development of cancer. The mechanism by which modifier genes alter adenoma incidence can be readily studied in mice that inherit mutations in the Apc gene. We identified a new modifier locus of ApcMin-induced intestinal tumorigenesis called Modifier of Min 2 (Mom2). The polyp-resistant Mom2R phenotype resulted from a spontaneous mutation and linkage analysis localized Mom2 to distal chromosome 18. To obtain recombinant chromosomes for use in refining the Mom2 interval, we generated congenic DBA.B6 ApcMin/+, Mom2R/+ mice. An intercross revealed that Mom2R encodes a recessive embryonic lethal mutation. We devised an exclusion strategy for mapping the Mom2 locus using embryonic lethality as a method of selection. Expression and sequence analyses of candidate genes identified a duplication of four nucleotides within exon 3 of the alpha subunit of the ATP synthase (Atp5a1) gene. Tumor analyses revealed a novel mechanism of polyp suppression by Mom2R in Min mice. Furthermore, we show that more adenomas progress to carcinomas in Min mice that carry the Mom2R mutation. The absence of loss of heterozygosity (LOH) at the Apc locus, combined with the tendency of adenomas to progress to carcinomas, indicates that the sequence of events leading to tumors in ApcMin/+ Mom2R/+ mice is consistent with the features of human tumor initiation and progression.