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.

Cardio-facio-cutaneous (CFC) syndrome is a sporadic developmental disorder involving characteristic craniofacial features, cardiac defects, ectodermal abnormalities, and developmental delay. We demonstrate that heterogeneous de novo missense mutations in three genes within the mitogen-activated protein kinase (MAPK) pathway cause CFC syndrome. The majority of cases (18 out of 23) are caused by mutations in BRAF, a gene frequently mutated in cancer. Of the 11 mutations identified, two result in amino acid substitutions that occur in tumors, but most are unique and suggest previously unknown mechanisms of B-Raf activation. Furthermore, three of five individuals without BRAF mutations had missense mutations in either MEK1 or MEK2, downstream effectors of B-Raf. Our findings highlight the involvement of the MAPK pathway in human development and will provide a molecular diagnosis of CFC syndrome.

We have investigated the contribution of CDK4 and CDK2 inhibition to G1 arrest in colon cancers following inhibition of the MEK/MAP kinase pathway. CDK4 inhibition is sufficient to cause arrest, but inhibition of CDK2 by p27 Kip1 redistribution or ectopic expression has no effect on proliferation. Likewise, inhibition of CDK2 through expression of dominant-negative (DN) CDK2 or antisense oligonucleotides did not prevent cell proliferation in these cells. We therefore tested whether CDK2 activity is dispensable in other cells. Surprisingly, osteosarcomas and Rb-negative cervical cancers continued to proliferate after depletion of CDK2 through antisense oligonucleotides or small interfering (si) RNA. Here we report of sustained cell proliferation in the absence of CDK2, and we suggest that CDK2 is not a suitable target for cancer therapy.