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Identification of ATM-dependent long non-coding RNAs induced in response to DNA damage
Marta Podralska1, Marcin Piotr Sajek2, Antonina Bielicka1
1Institute of Human Genetics, Polish Academy of Sciences, Poznan, Poland.
Abstract:
DNA damage response (DDR) is a complex process, essential for cell survival. Especially deleterious type of DNA damage are DNA double-strand breaks (DSB), which can lead to genomic instability and malignant transformation if not repaired correctly. The central player in DSB detection and repair is the ATM kinase which orchestrates the action of several downstream factors. Recent studies have suggested that long non-coding RNAs (lncRNAs) are involved in DDR. Here, we aimed to identify lncRNAs induced upon DNA damage in an ATM-dependent manner. DNA damage was induced by ionizing radiation (IR) in immortalized lymphoblastoid cell lines derived from 4 patients with ataxia-telangiectasia (AT) and 4 healthy donors. RNA-seq revealed 10 lncRNAs significantly induced 1 h after IR in healthy donors, whereas none in AT patients. 149 lncRNAs were induced 8 h after IR in the control group, while only three in AT patients. Among IR-induced mRNAs, we found several genes with well-known functions in DDR. Gene Set Enrichment Analysis and Gene Ontology revealed delayed induction of key DDR pathways in AT patients compared to controls. The induction and dynamics of selected 9 lncRNAs were confirmed by RT-qPCR. Moreover, using a specific ATM inhibitor we proved that the induction of those lncRNAs is dependent on ATM. Some of the detected lncRNA genes are localized next to protein-coding genes involved in DDR. We observed that induction of lncRNAs after IR preceded changes in expression of adjacent genes. This indicates that IR-induced lncRNAs may regulate the transcription of nearby genes. Subcellular fractionation into chromatin, nuclear, and cytoplasmic fractions revealed that the majority of studied lncRNAs are localized in chromatin. In summary, our study revealed several lncRNAs induced by IR in an ATM-dependent manner. Their genomic co-localization and co-expression with genes involved in DDR suggest that those lncRNAs may be important players in cellular response to DNA damage.
Insights
This study identifies long non-coding RNAs (lncRNAs) induced by DNA damage in an ATM-dependent manner. These lncRNAs may play a role in regulating genes involved in the DNA damage response.
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- DNA double-strand breaks (DSBs) are critical DNA damage events.
- The ATM kinase is central to DNA damage response (DDR).
- Long non-coding RNAs (lncRNAs) are increasingly recognized for their roles in cellular processes.
Purpose of the Study:
- To identify lncRNAs induced by DNA damage in an ATM-dependent manner.
- To investigate the role of lncRNAs in the cellular response to DNA damage.
Main Methods:
- Ionizing radiation (IR) was used to induce DNA damage in lymphoblastoid cell lines from healthy donors and ataxia-telangiectasia (AT) patients.
- RNA sequencing (RNA-seq) was performed to analyze lncRNA and mRNA expression.
- Quantitative real-time PCR (RT-qPCR) validated lncRNA induction.
- ATM inhibition and subcellular fractionation were used to further characterize lncRNA function and localization.
Main Results:
- 10 and 149 lncRNAs were significantly induced by IR in healthy donors at 1h and 8h, respectively, with minimal induction in AT patients.
- ATM kinase activity is crucial for the induction of these IR-responsive lncRNAs.
- Gene expression analysis revealed delayed DDR pathway activation in AT patients.
- Many identified lncRNAs are located near DDR-related genes and their induction precedes changes in adjacent gene expression.
- The majority of the studied lncRNAs were found to be chromatin-localized.
Conclusions:
- Several lncRNAs are induced by ionizing radiation in an ATM-dependent manner.
- These lncRNAs may function as regulators of nearby genes involved in DNA damage response.
- The findings highlight a novel role for lncRNAs in the complex DNA damage response network.
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