The online version of this article (doi:10.1186/1476-4598-11-18) contains supplementary material, which is available to authorized users.
The authors declare that they have no competing interests.
SS, XW, SM and HD conceived and designed the experiments. SS, XW and HD generated the gene-targeting vector and produced the knockout mice and characterized the tumor phenotypes. SS and SM performed cytogenetic analysis. ZN, MR and SK provided reagents and technical supports for this study. SS and HD wrote the manuscript with subsequent contributions from all authors. All authors read and approved the final manuscript.
HLTF (Helicase-like Transcription Factor) is a DNA helicase protein homologous to the SWI/SNF family involved in the maintenance of genomic stability and the regulation of gene expression. HLTF has also been found to be frequently inactivated by promoter hypermethylation in human colon cancers. Whether this epigenetic event is required for intestinal carcinogenesis is unknown.
To address the role of loss of HLTF function in the development of intestinal cancer, we generated Hltf deficient mice. These mutant mice showed normal development, and did not develop intestinal tumors, indicating that loss of Hltf function by itself is insufficient to induce the formation of intestinal cancer. On the Apc min/+ mutant background, Hltf - deficiency was found to significantly increase the formation of intestinal adenocarcinoma and colon cancers. Cytogenetic analysis of colon tumor cells from Hltf -/-/Apc min/+ mice revealed a high incidence of gross chromosomal instabilities, including Robertsonian fusions, chromosomal fragments and aneuploidy. None of these genetic alterations were observed in the colon tumor cells derived from Apc min/+ mice. Increased tumor growth and genomic instability was also demonstrated in HCT116 human colon cancer cells in which HLTF expression was significantly decreased.
Taken together, our results demonstrate that loss of HLTF function promotes the malignant transformation of intestinal or colonic adenomas to carcinomas by inducing genomic instability. Our findings highly suggest that epigenetic inactivation of HLTF, as found in most human colon cancers, could play an important role in the progression of colon tumors to malignant cancer.
Additional file 1: Loss of Hltf function does not affect the cellular proliferation in small intestine and colon. The small intestines and colons collected from 2-month old Hltf +/+ and Hltf -/- mice were stained with Haematoxylin-eosin and anti-Brdu antibody. Both Hltf +/+ and Hltf -/- intestine and colon displayed normal morphology and a similar number of BrdU positive cells within the crypt (2 h after BrdU injection) and in crypt-villus axis (24 h after BrdU injection). (PDF 11 MB)12943_2012_1000_MOESM1_ESM.PDF
Additional file 2: Loss of Hltf function does not affect the differentiation of epithelial cells in small intestine and colon. The small intestines and colons from 2-month old Hltf +/+ and Hltf -/- mice were analyzed using several intestinal cell-lineage markers. The Goblet cells were determined by staining with Alcian blue and periodic acid-Schiff (PAS). The enteroendocrine cells were analyzed by immuno-staining with anti-chromogranin A, and the Paneth cells in small intestine were detected by anti-lysozyme antibody. Both Hltf +/+ and Hltf -/- intestines or colons showed very similar staining patterns for these markers. (PDF 12 MB)12943_2012_1000_MOESM2_ESM.PDF
Additional file 3: Hltf -/- /Apc min/+ mice frequently developed invasive intestinal adenocarcinomas. Several additional Haematoxylin-eosin stained images demonstrate the formation of invasive intestinal adenocarcinomas (indicated by arrows) in Hltf -/-/Apc min/+ mice. (PDF 5 MB)12943_2012_1000_MOESM3_ESM.PDF
Additional file 4: List of genes that showed expression changes between Hltf+/+ and Hltf-/- mouse ES cells and in HCT116 cells with and without HLTF knockdown. The microarray expression assays were carried out by the Center for Applied Genomics at the Toronto Hospital for Sick Children. Three independent cell lines from each group were analyzed. The changes with the statistical significance (p < 0.05) are highlighted by yellow. Red indicates the change of gene expression cannot be validated by real-time PCR. (PDF 498 KB)12943_2012_1000_MOESM4_ESM.PDF
Additional file 5: List of PCR primers that were applied in this study. (PDF 61 KB)12943_2012_1000_MOESM5_ESM.PDF
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