Identification of genes that are down-regulated in human colon tumors and are dependent on APC

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The tumor suppressor gene adenomatous polyposis coli (APC) is mutated in a majority of colorectal tumors. Numerous studies have shown that APC binds to and promotes degradation of the wnt-signaling component β-catenin. Mutations in APC lead to increased transcriptional activity of β-catenin/Tcf/Lef complexes and are thought to cause neoplasms by promoting proliferation and preventing proper differentiation of colonocytes. In order to better understand the mechanisms of colonocyte differentiation, we performed microarray analyses on colon adenomas and carcinomas compared to normal colon. We found that colon adenomas and carcinomas have elevated β-catenin target gene expression, as expected. Our analyses also revealed a preponderance of genes involved in normal colonocyte differentiation that were down-regulated in tumors compared to normal. To investigate the ability of APC to positively regulate gene expression, we used a model system in which wild type APC was reconstituted into the APC-deficient cell line HT29, and performed microarray analyses. Induction of APC over a 48-hour timecourse generated a list of 65 genes with a correlation coefficient of 0.98 that showed parallel induction to APC. Among those were a colon-specific retinol dehydrogenase (RDHL), epithelial-specific transcription factor ESE-1/ESX/ELF-3, carcinoembryonic antigen-related cell adhesion molecule CEACAM1, caspase-7, and transcription factor GATA-binding protein 2. We confirmed induction of these genes by northern blot or quantitative rt-pcr. We then examined human colon tumors and found that these gene transcripts were down-regulated in at least 50% of the samples assayed. Probing into the mechanisms of regulation by APC, we found that induction of certain genes, like RDHL, were independent of β-catenin in that introduction of dominant negative Lef did not reactivate the gene. These findings support a role for APC in regulating genes involved in normal differentiation of the colon by a pathway distinct from β-catenin/Tcf/Lef. Loss of APC may result in decreased expression of these genes, thereby contributing to tumor progression.