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Published on: March 4, 2020
Novel radiation response genes identified in gene-trapped MCF10A mammary epithelial cells
Jennifer Malone1, Robert Ullrich
1Department of Pathology, University of Colorado Health Sciences Center, Aurora, Colorado 80045-0508, USA. Jennifer.Malone@uchsc.edu
Abstract:
We have used a gene-trapping strategy to screen human mammary epithelial cells for radiation response genes. Relative mRNA expression levels of five candidate genes in MCF10A cells were analyzed, both with and without exposure to radiation. In all five cases, the trapped genes were significantly down-regulated after radiation treatment. Sequence analysis of the fusion transcripts identified the trapped genes: (1) the human androgen receptor, (2) the uncharacterized DREV1 gene, which has known homology to DNA methyltransferases, (3) the human creatine kinase gene, (4) the human eukaryotic translation elongation factor 1 beta 2, and (5) the human ribosomal protein L27. All five genes were down-regulated significantly after treatment with varying doses of ionizing radiation (0.10 to 4.0 Gy) and at varying times (2-30 h after treatment). The genes were also analyzed in human fibroblast and lymphoblastoid cell lines to determine whether the radiation response being observed was cell-type specific. The results verified that the observed radiation response was not a cell-type-specific phenomenon, suggesting that the genes play essential roles in the radiation damage control pathways. This study demonstrates the potential of the gene-trap approach for the identification and functional analysis of novel radiation response genes.
Insights
This study identified five key genes, including the androgen receptor and ribosomal protein L27, that are down-regulated after radiation exposure in human cells. These findings highlight their crucial role in DNA damage control pathways.
Area of Science:
- Molecular Biology
- Genetics
- Radiation Biology
Background:
- Identifying genes involved in cellular radiation response is crucial for understanding DNA damage control.
- Gene-trapping offers a powerful strategy for discovering novel genes with specific cellular functions.
Purpose of the Study:
- To screen human mammary epithelial cells for radiation response genes using a gene-trapping approach.
- To identify and characterize genes whose expression is altered following ionizing radiation exposure.
Main Methods:
- Utilized a gene-trapping strategy in MCF10A human mammary epithelial cells.
- Analyzed mRNA expression levels of candidate genes before and after radiation treatment.
- Performed sequence analysis of fusion transcripts to identify trapped genes.
- Validated findings in human fibroblast and lymphoblastoid cell lines.
Main Results:
- Five candidate genes, including human androgen receptor, DREV1, creatine kinase, eukaryotic translation elongation factor 1 beta 2, and ribosomal protein L27, were identified.
- All five genes showed significant down-regulation post-ionizing radiation exposure across various doses (0.10–4.0 Gy) and time points (2–30 h).
- The observed radiation response was consistent across different human cell types, indicating a non-cell-type-specific phenomenon.
Conclusions:
- The identified genes play essential roles in cellular radiation damage control pathways.
- The gene-trap approach is effective for discovering novel radiation response genes.
- This research provides insights into the molecular mechanisms underlying cellular responses to ionizing radiation.
