APC1638T: a mouse model delineating critical domains of the adenomatous polyposis coli protein involved in tumorigenesis and development

This is the publisher's version, also available electronically from "http://genesdev.cshlp.org".The adenomatous polyposis coli (APC) gene is considered as the true gatekeeper of colonic epithelial proliferation: It is mutated in the majority of colorectal tumors, and mutations occur at early stages of tumor development in mouse and man. These mutant proteins lack most of the seven 20-amino-acid repeats and all SAMP motifs that have been associated with down-regulation of intracellular β-catenin levels. In addition, they lack the carboxy-terminal domains that bind to DLG, EB1, and microtubulin. APC also appears to be essential in development because homozygosity for mouse Apcmutations invariably results in early embryonic lethality. Here, we describe the generation of a mouse model carrying a targeted mutation at codon 1638 of the mouse Apc gene, Apc1638T, resulting in a truncated Apc protein encompassing three of the seven 20 amino acid repeats and one SAMP motif, but missing all of the carboxy-terminal domains thought to be associated with tumorigenesis. Surprisingly, homozygosity for the Apc1638T mutation is compatible with postnatal life. However, homozygous mutant animals are characterized by growth retardation, a reduced postnatal viability on the B6 genetic background, the absence of preputial glands, and the formation of nipple-associated cysts. Most importantly,Apc 1638T/1638T animals that survive to adulthood are tumor free. Although the full complement of Apc1638T is sufficient for proper β-catenin signaling, dosage reductions of the truncated protein result in increasingly severe defects in β-catenin regulation. The SAMP motif retained in Apc1638T also appears to be important for this function as shown by analysis of the Apc1572T protein in which its targeted deletion results in a further reduction in the ability of properly controlling β-catenin/Tcf signaling. These results indicate that the association with DLG, EB1, and microtubulin is less critical for the maintenance of homeostasis by APC than has been suggested previously, and that proper β-catenin regulation by APC appears to be required for normal embryonic development and tumor suppression

Characterization and comparison of the human and mouse dist1l α-globin complex reveals a tightly packed multiple gene cluster containing differentially expressed transcription units

In this paper, we describe the detailed analysis of about 75 kb of genomic DNA flanking the 5′ end of the mouse α-globin region and complete the transcription map of the human region. Previously, we established the homology of the human and mouse α-globin upstream flanking regions (αUFR) and characterized in detail the mouse α-globin major regulatory element (αMRE) and the mMPG DNA repair gene. Here, we extend our analysis with the construction of a detailed restriction map, the mapping and isolation of two nonglobin genes, named mDist1 and mProx1, the distribution of 18 DNase hypersensitive sites (HSSs) in erythroid and fibroblast cells, and the analysis of the mDist1, mMPG, and mProx1 expression levels in several adult tissues and during fetal development. In addition, the hDist1 gene is exactly localized 1.9 kb from the hMPG gene. The mapping results show that the Dist1, MPG, and Prox1 genes, together with the α-globin genes and the αMRE, form a tightly packed multiple gene cluster that is 50% more compact in mouse than in human. The expression results show that each of the genes present in this locus displays a characteristic expression pattern in adult tissues and during fetal development. The 18 DNase HSSs observed were scattered over this region. Interestingly, all the erythroid-sensitive HSSs were associated with the Prox1 transcription unit, whereas the only two pairs of fibroblast-sensitive HSSs present in this locus were located in the promoter regions of the mProx1 and mDist1/mMPG genes. The possible role of the erythroid- and fibroblastsensitive sites in the regulation of the mouse α-globin and nonglobin gene expression is discussed. The characterization of the mouse αUFR identifies most, if not all, of the structural elements possibly involved in the regulation of mα-globin gene expression and sheds light on the organization and evolution of the telomere-associated, GC-rich isochore family H3

Somatic Apc mutations are selected upon their capacity to inactivate the β-catenin downregulating activity

The APC gene, originally identified as the gene for familial adenomatous polyposis (FAP), is now considered as the true 'gatekeeper' of colonic epithelial proliferation. Its main tumor suppressing activity seems to reside in the capacity to properly regulate intracellular β-catenin signaling. Most somatic APC mutations are detected between codons 1286 and 1513, the mutation cluster region (MCR). This clustering can be explained either by the presence of mutation-prone sequences within the MCR, or by the selective advantage provided by the resulting truncated polypeptides. Here, a Msh2-deficient mouse model (Msh2(Δ7N)) was generated and bred with Apc(1638N) and Apc(Min) that allowed the comparison of the somatic mutation spectra along the Apc gene in the different allelic combinations. Mutations identified in Msh2(Δ7N/Δ7N) tumors are predominantly dinucleotide deletions at simple sequence repeats leading to truncated Apc polypeptides that partially retain the 20 a.a. β-catenin downregulating motifs. In contrast, the somatic mutations identified in the wild type Apc allele of Msh2(Δ7N/Δ7N/Apc(+/1638N) and Msh2(Δ7N/Δ7N)/Apc(+Min) tumors are clustered more to the 5' end, thereby completely inactivating the β-catenin downregulating activity of APC. These results indicate that somatic Apc mutations are selected during intestinal tumorigenesis and that inactivation of the β-catenin downregulating function of APC is likely to represent the main selective factor. (C) 2000 Wiley-Liss, Inc

Apc modulates embryonic stem-cell differentiation by controlling the dosage of β-catenin signaling

The Wnt signal-transduction pathway induces the nuclear translocation of membrane-bound β-catenin (Catnb) and has a key role in cell-fate determination. Tight somatic regulation of this signal is essential, as uncontrolled nuclear accumulation of β-catenin can cause developmental defects and tumorigenesis in the adult organism. The adenomatous polyposis coli gene (APC) is a major controller of the Wnt pathway and is essential to prevent tumorigenesis in a variety of tissues and organs. Here, we have investigated the effect of different mutations in Apc on the differentiation potential of mouse embryonic stem (ES) cells. We provide genetic and molecular evidence that the ability and sensitivity of ES cells to differentiate into the three germ layers is inhibited by increased doses of β-catenin by specific Apc mutations. These range from a severe differentiation blockade in Apc alleles completely deficient in β-catenin regulation to more specific neuroectodermal, dorsal mesodermal and endodermal defects in more hypomorphic alleles. Accordingly, a targeted oncogenic mutation in Catnb also affects the differentiation potential of ES cells. Expression profiling of wildtype and Apc-mutated teratomas supports the differentiation defects at the molecular level and pinpoints a large number of downstream structural and regulating genes. Chimeric experiments showed that this effect is cell-autonomous. Our results imply that constitutive activation of the Apc/β-catenin signaling pathway results in differentiation defects in tissue homeostasis, and possibly underlies tumorigenesis in the colon and other self-renewing tissues

Apc1638T: a mouse model delineating critical domains of the adenomatous polyposis coli protein involved in tumorigenesis and development

The adenomatous polyposis coli (APC) gene is considered as the true gatekeeper of colonic epithelial proliferation: It is mutated in the majority of colorectal tumors, and mutations occur at early stages of tumor development in mouse and man. These mutant proteins lack most of the seven 20-amino-acid repeats and all SAMP motifs that have been associated with down-regulation of intracellular β-catenin levels. In addition, they lack the carboxy-terminal domains that bind to DLG, EB1, and microtubulin. APC also appears to be essential in development because homozygosity for mouse Apc mutations invariably results in early embryonic lethality. Here, we describe the generation of a mouse model carrying a targeted mutation at codon 1638 of the mouse Apc gene, Apc1638T, resulting in a truncated Apc protein encompassing three of the seven 20 amino acid repeats and one SAMP motif, but missing all of the carboxy-terminal domains thought to be associated with tumorigenesis. Surprisingly, homozygosity for the Apc1638T mutation is compatible with postnatal life. However, homozygous mutant animals are characterized by growth retardation, a reduced postnatal viability on the B6 genetic background, the absence of preputial glands, and the formation of nipple-associated cysts. Most importantly, Apc(1638T/1638T) animals that survive to adulthood are tumor free. Although the full complement of Apc1638T is sufficient for proper β-catenin signaling, dosage reductions of the truncated protein result in increasingly severe defects in β-catenin regulation. The SAMP motif retained in Apc1638T also appears to be important for this function as shown by analysis of the Apc1572T protein in which its targeted deletion results in a further reduction in the ability of properly controlling β-catenin/Tcf signaling. These results indicate that the association with DLG, EB1, and microtubulin is less critical for the maintenance of homeostasis by APC than has been suggested previously, and that proper β-catenin regulation by APC appears to be required for normal embryonic development and tumor suppression