Mutations in the adenomatous polyposis coli (APC) tumour-suppressor gene occur in most human colon cancers. Loss of functional APC protein results in the accumulation of beta-catenin. Mutant forms of beta-catenin have been discovered in colon cancers that retain wild-type APC genes, and also in melanomas, medulloblastomas, prostate cancer and gastric and hepatocellular carcinomas. The accumulation of beta-catenin activates genes that are responsive to transcription factors of the TCF/LEF family, with which beta-catenin interacts. Here we show that beta-catenin activates transcription from the cyclin D1 promoter, and that sequences within the promoter that are related to consensus TCF/LEF-binding sites are necessary for activation. The oncoprotein p21ras further activates transcription of the cyclin D1 gene, through sites within the promoter that bind the transcriptional regulators Ets or CREB. Cells expressing mutant beta-catenin produce high levels of cyclin D1 messenger RNA and protein constitutively. Furthermore, expression of a dominant-negative form of TCF in colon-cancer cells strongly inhibits expression of cyclin D1 without affecting expression of cyclin D2, cyclin E, or cyclin-dependent kinases 2, 4 or 6. This dominant-negative TCF causes cells to arrest in the G1 phase of the cell cycle; this phenotype can be rescued by expression of cyclin D1 under the cytomegalovirus promoter. Abnormal levels of beta-catenin may therefore contribute to neoplastic transformation by causing accumulation of cyclin D1.

Beta-catenin regulates expression of cyclin D1 in colon carcinoma cells.

The adenomatous polyposis coli (APC) tumor suppressor protein binds to beta-catenin, a protein recently shown to interact with Tcf and Lef transcription factors. The gene encoding hTcf-4, a Tcf family member that is expressed in colonic epithelium, was cloned and characterized. hTcf-4 transactivates transcription only when associated with beta-catenin. Nuclei of APC-/- colon carcinoma cells were found to contain a stable beta-catenin-hTcf-4 complex that was constitutively active, as measured by transcription of a Tcf reporter gene. Reintroduction of APC removed beta-catenin from hTcf-4 and abrogated the transcriptional transactivation. Constitutive transcription of Tcf target genes, caused by loss of APC function, may be a crucial event in the early transformation of colonic epithelium.

/pdf/constitutive-transcriptional-activation-by-a-b-catenin-tcf-27dkig5evt.pdf

Constitutive Transcriptional Activation by a β-Catenin-Tcf Complex in APC−/− Colon Carcinoma

The regulation of cell growth and survival can be subverted by a variety of genetic defects that alter transcriptional programs normally responsible for controlling cell number. High throughput analysis of these gene expression patterns should ultimately lead to the identification of minimal expression profiles that will serve as common denominators in assigning a cancer to a given category. In the course of defining the common denominators, though, we should not be too surprised to find that cancers within a single category may nevertheless exhibit seemingly disparate genetic defects. The wnt pathway has already provided an outstanding example of this. We now know of three regulatory genes in this pathway that are mutated in primary human cancers and several others that promote experimental cancers in rodents (Fig. 1). In all of these cases the common denominator is the activation of gene transcription by -catenin. The resulting gene expression profile should provide us with a signature common to those cancers carrying defects in the wnt pathway. In this review, the wnt pathway will be covered from the perspective of cancer, with emphasis placed on molecular defects known to promote neoplastic transformation in humans and in animal models.

/pdf/wnt-signaling-and-cancer-2fdsemer2h.pdf

Wnt signaling and cancer

The p53 tumour suppressor gene is the most widely mutated gene in human tumorigenesis. p53 encodes a transcriptional activator whose targets may include genes that regulate genomic stability, the cellular response to DNA damage, and cell-cycle progression. Introduction of wild-type p53 into cell lines that have lost endogenous p53 function can cause growth arrest or induce a process of cell death known as apoptosis. During normal development, self-reactive thymocytes undergo negative selection by apoptosis, which can also be induced in immature thymocytes by other stimuli, including exposure to glucocorticoids and ionizing radiation. Although normal negative selection involves signalling through the T-cell receptor, the induction of apoptosis by other stimuli is poorly understood. We have investigated the requirement for p53 during apoptosis in mouse thymocytes. We report here that immature thymocytes lacking p53 die normally when exposed to compounds that may mimic T-cell receptor engagement and to glucocorticoids but are resistant to the lethal effects of ionizing radiation. These results demonstrate that p53 is required for radiation-induced cell death in the thymus but is not necessary for all forms of apoptosis.

p53 is required for radiation-induced apoptosis in mouse thymocytes

RAG-1-deficient mice have no mature B and T lymphocytes

Nuclear localization of beta-catenin by interaction with transcription factor LEF-1.

This study demonstrates that blastocyst-derived embryonic stem cells (ES cells) can be used as a vehicle for transgenesis. The method is nearly as efficient as other methods, and the introduced neomycin phosphotransferase (neo) gene is stably transmitted through several generations with no apparent loss in G418 resistance. An important factor contributing to the efficiency of this process is the rigorous selection, before blastocyst injection, of genetically transformed cells for in vitro developmental pluripotency. One of the advantages of the ES cell route to transgenesis is that it provides investigators with the opportunity to screen for the desired genetic alterations before reintroducing the ES cells into the animal.

https://www.pnas.org/content/pnas/83/23/9065.full.pdf

Transgenesis by means of blastocyst-derived embryonic stem cell lines

Enhancer trap integrations in mouse embryonic stem cells give rise to staining patterns in chimaeric embryos with a high frequency and detect endogenous genes

Danforth's short tail (Sd) is a semidominant mutation on mouse chromosome 2 that acts cell autonomously in the notochord and leads to its disintegration, and thus causes severe defects in somite patterning and vertebral column development. The molecular nature of the Sd gene and mutation is unknown, and it is unclear whether Sd is a loss-of-function mutation and the semidominant inheritance of the Sd phenotype is due to haploinsufficiency, or whether Sd represents a gain-of-function mutation in a gene essential for notochord development and maintenance. Here, we report on the genetic interaction between Sd and an insertional mutation called Etl4lacZ, which provides genetic evidence that Sd is a gain-of-function mutation. Etl4lacZ is an enhancer trap insertion, which gives rise to lacZ expression in distinct cell types, including the notochord. In homozygosity, the lacZ insertion leads to abnormal vertebrae in the caudal part of the vertebral column. Etl4lacZ maps approximately 0.75 cM distal to Sd, and in double heterozygotes modifies the Sdphenotype contrarily, depending on the chromosomal configuration of the Sd and Etl4lacZ mutations: when Etl4lacZ is present on the chromosome wild type for Sd (Sd +/+ Etl4lacZ; trans configuration), the Sd phenotype is enhanced, i.e., vertebral malformations extend to more anterior positions and the vertebral body of the axis is further reduced. Conversely, when Etl4lacZ is present on the same chromosome as Sd (SdEtl4lacZ /+ +; cis configuration), the Sd phenotype is attenuated, i.e., vertebral malformations are confined to more posterior levels, and the dens axis, which is severely reduced or absent in Sd heterozygotes, is restored. The different effect of the Etl4lacZ insertion on Sd, depending on its presence in trans or cis, suggests a direct interaction of the transgene insertion with the Sd gene. Additionally, the attenuation of the Sd phenotype by Etl4lacZ in cis suggests that Sd is a gain-of-function mutation and lends support to the idea that Etl4lacZ is a new allele of Sd. Dev. Genet. 23:86–96, 1998. © 1998 Wiley-Liss, Inc.

Reinhard Korn

Papers

Nuclear localization of beta-catenin by interaction with transcription factor LEF-1.

Transgenesis by means of blastocyst-derived embryonic stem cell lines

Enhancer trap integrations in mouse embryonic stem cells give rise to staining patterns in chimaeric embryos with a high frequency and detect endogenous genes

Genetic interactions suggest that Danforth's short tail (Sd) is a gain-of-function mutation.