Abstract
Cells alter their gene expression in response to exposure to various environmental changes. Epigenetic mechanisms such as DNA methylation are believed to regulate the alterations in gene expression patterns. In vitro and in vivo studies have documented changes in cellular proliferation, cytoskeletal remodeling, signal transduction, bone mineralization and immune deficiency under the influence of microgravity conditions experienced in space. However microgravity induced changes in the epigenome have not been well characterized. In this study we have used Next-generation Sequencing (NGS) to profile ground-based “simulated” microgravity induced changes on DNA methylation (5-methylcytosine or 5mC), hydroxymethylation (5-hydroxymethylcytosine or 5hmC), and simultaneous gene expression in cultured human lymphoblastoid cells. Our results indicate that simulated microgravity induced alterations in the methylome (~60% of the differentially methylated regions or DMRs are hypomethylated and ~92% of the differentially hydroxymethylated regions or DHMRs are hyperhydroxymethylated). Simulated microgravity also induced differential expression in 370 transcripts that were associated with crucial biological processes such as oxidative stress response, carbohydrate metabolism and regulation of transcription. While we were not able to obtain any global trend correlating the changes of methylation/ hydroxylation with gene expression, we have been able to profile the simulated microgravity induced changes of 5mC over some of the differentially expressed genes that includes five genes undergoing differential methylation over their promoters and twenty five genes undergoing differential methylation over their gene-bodies. To the best of our knowledge, this is the first NGS-based study to profile epigenomic patterns induced by short time exposure of simulated microgravity and we believe that our findings can be a valuable resource for future explorations.
Highlights
During space flight, astronauts are exposed to powerful environmental assaults such as microgravity, cosmic radiation and magnetic fields that have the potential to impinge upon cellular ontogeny through epigenetic modifications [1]
We applied MeDIP coupled with high-throughput sequencing to identify the differences in the genome-wide patterns of 5mC upon simulated microgravity on TK6 cells. 2.8x108 and 1.8x108 reads were obtained during MeDIP-seq from TK6 cells subjected to static and simulated microgravity respectively and more than 90% of these reads aligned to the human genome GRCh37/ hg19, 2009 Assembly (S1 Table and S2 Fig)
The similarity between the replicates was evident as hierarchical dendrogram displayed distinct clustering of biological replicates in two groups (S3 Fig) and sequence coverage analyses displayed that MeDIP-seq reads generated from the samples covered similar number of bases of the reference genome (S4 Fig)
Summary
Astronauts are exposed to powerful environmental assaults such as microgravity, cosmic radiation and magnetic fields that have the potential to impinge upon cellular ontogeny through epigenetic modifications [1]. Studies of human lymphoblast and lymphoblast cell cultures following periods of simulated microgravity have demonstrated alterations in metabolic processes and DNA repair pathways which could in turn signify an increased susceptibility to malignancy [12, 13]. These studies indicate exposure to microgravity during space flight alters gene expression patterns and subsequently cellular physiology
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