Transformation of normal NIH-3T3 cells into highly oncogenic and invasive cancer cells by MBD2 overexpression
Bibliographic record
Abstract
3925 Introduction: Cancer cells are hallmarked by global DNA hypomethylation and regional hypermethylation of tumor suppressor genes. The latter has been extensively studied while not much attention has been given to global DNA hypomethylation. MBD2 is the only protein that has been shown to directly remove the methyl group from CG di-nucleotides; though, it has also been reported to be a transcriptional repressor. Inhibition of MBD2 attenuates tumorigenesis, metastasis, and reverses the hypomethylation of metastatic genes in cell culture and in vivo. Additionally, germ line deletion of the mbd2 gene protects mice from intestinal tumors. Therefore, we tested the hypothesis that MBD2 causes oncogenic and metastatic transformation of cells by triggering global hypomethylation. Results: MBD2 and a mutant MBD2 lacking the MBD domain were stably and transiently expressed in mouse fibroblast NIH-3T3 cells. We determined that MBD2 induces global hypomethylation and histone H3 lysine 9 acetylation in an MBD domain dependent manner. Several assays revealed that the MBD2 cell lines, but not the mutant nor the control cell lines, were transformed and highly invasive in cell culture. Only the MBD2 cell lines were highly tumorigenic and invasive when injected into Nude mice while also showing a remarkable ability to invade and degrade bone tissue when injected into the tibia of SCID mice. We also determined that MBD2 is required for maintaining a transformed state triggered by activated HA-RAS in NIH-3T3 cells. MBD2 knockdown in these cells attenuated transformation and increased DNA methylation supporting our hypothesis. To delineate the effects of MBD2 overexpression on specific gene expression, a genomic approach using affymetrix expression arrays was employed. Four hundred and forty four genes were identified to have their mRNA expression altered by MBD2 expression. Genes involved in tumorgenesis, bone degradation, or with no known function that were induced in both MBD2 and HA-Ras expressing cell lines were chosen for further study. Knocking down selected candidate genes9 mRNA in MBD2 overexpressing NIH-3T3 cells resulted in attenuated tumorigenesis while knocking down MBD2 in HA-RAS cells significantly reduced these genes9 expression levels. CHIP assay of the promoter and bisulfite sequencing of the same regions for the candidate genes revealed that MBD2 binds only to regions associated with demethylated DNA and that these regions are associated with H3 lysine 9 acetylation. Summary: Our results support the hypothesis that MBD2, through activation of global cancer pathways, transforms normal fibroblast cells into highly oncogenic, invasive, and metastatic cancer cells; therefore, MBD2 is a novel cancer therapeutic candidate. Supported by a grant from the National Cancer Institute of Canada to MS.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.001 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".