Identification of up-regulated and down-regulated cis-natural antisense transcripts in the human B
Mitsuru Chiba1, Tomisato Miura, Kosuke Kasai
1Department of Biomedical Sciences, Division of Medical Life Sciences, Hirosaki University Graduate School of Health Sciences, Hirosaki, Aomori 036-8564, Japan. mchiba32@cc.hirosaki-u.ac.jp
Abstract:
Ionizing radiation (IR) causes DNA injury and induces multiple signal mechanisms, including the regulation of DNA repair, the cell cycle and gene expression through the activation of p53-related pathways. Cis-natural antisense transcripts (cis-NATs), which are transcribed from the DNA strand opposite to that for mRNA of the gene, are recognized as important regulators of gene expression in eukaryotic cells, but the effects on cis-NAT expression by IR are unknown to date. Therefore, we investigated the effects of X-ray irradiation on cis-NAT expression together with mRNA expression using a human B lymphoblast cell line (IM-9), a custom-microarray and strand-specific RT-qPCR. Eighteen, 33 and 106 mRNAs were demonstrated to be differentially expressed in IM-9 cells after 1, 2 and 4 Gy irradiation, respectively, as compared to 0 Gy by microarray analysis (fold change, FC >2.0). On the other hand, 10, 22 and 43 NATs were demonstrated to be differentially expressed in IM-9 cells after 1, 2 and 4 Gy irradiation, respectively, as compared to 0 Gy by microarray analysis (FC >2.0). Among these mRNAs/NATs, the IR dose-dependent up-regulation of mRNAs and cis-NATs of MDM2 and CDKN1A were confirmed by strand-specific RT-qPCR. Additionally, the cis-NATs of MDM2 were indicated to be localized in the cytoplasm, while cis-NATs of CDKN1A were located in the nucleus and cytoplasm. In conclusion, the radiation-responsive cis-NATs in conjunction with mRNAs were identified for the first time in the present study. It is possible that these cis-NATs regulate the gene expression in a post-transcriptional fashion. The IR dose-dependent up- and down-regulation of these mRNAs/cis-NATs may be a marker for ionizing radiation.
Insights
Ionizing radiation (IR) affects gene expression by altering natural antisense transcripts (NATs). Researchers identified radiation-responsive cis-NATs and mRNAs, suggesting their potential as biomarkers for IR exposure.
Area of Science:
- Molecular Biology
- Radiation Biology
- Genomics
Background:
- Ionizing radiation (IR) triggers cellular responses, including DNA repair and cell cycle regulation via p53 pathways.
- Cis-natural antisense transcripts (cis-NATs) are key gene expression regulators, but their response to IR remains uncharacterized.
Purpose of the Study:
- To investigate the impact of X-ray irradiation on cis-NAT expression alongside mRNA expression.
- To identify novel radiation-responsive cis-NATs and their potential role in cellular response to IR.
Main Methods:
- Utilized a human B lymphoblast cell line (IM-9) exposed to varying doses of X-ray irradiation (1, 2, and 4 Gy).
- Employed custom-microarray analysis to assess differential mRNA and cis-NAT expression.
- Validated key findings using strand-specific quantitative reverse transcription PCR (RT-qPCR).
Main Results:
- Microarray analysis revealed dose-dependent differential expression of numerous mRNAs and cis-NATs following IR.
- Confirmed IR dose-dependent up-regulation of MDM2 and CDKN1A mRNAs and their corresponding cis-NATs.
- Determined subcellular localization of cis-NATs for MDM2 (cytoplasm) and CDKN1A (nucleus and cytoplasm).
Conclusions:
- Identified radiation-responsive cis-NATs and mRNAs for the first time, offering new insights into IR effects.
- Hypothesized that these cis-NATs may regulate gene expression post-transcriptionally.
- Proposed that the IR dose-dependent expression of these mRNAs/cis-NATs could serve as biomarkers for ionizing radiation exposure.
