Simulated Microgravity-Induced Epigenetic Changes in Human Lymphocytes Kamaleshwar P. Singh, 1 * Ragini Kumari, 2 and James W. DuMond 2 1 Department of Environmental Toxicology, The Institute of Environmental and Human Health (TIEHH), Texas Tech University, Lubbock, Texas 79409 2 Department of Biology, Texas Southern University, Houston, Texas 77004 ABSTRACT Real space flight and modeled microgravity conditions result in changes in the expression of genes that control important cellular functions. However, the mechanisms for microgravity-induced gene expression changes are not clear. The epigenetic changes of DNA methylation and chromatin histones modifications are known to regulate gene expression. The objectives of this study were to investigate whether simulated microgravity alters (a) the DNA methylation and histone acetylation, and (b) the expression of DNMT1, DNMT3a, DNMT3b, and HDAC1 genes that regulate epigenetic events. To achieve these objectives, human T-lymphocyte cells were grown in a rotary cell culture system (RCCS) that simulates microgravity, and in parallel under normal gravitational conditions as control. The microgravity-induced DNA methylation changes were detected by methylation sensitive-random amplified polymorphic DNA (MS-RAPD) analysis of genomic DNA. The gene expression was measured by Quantitative Real-time PCR. The expression of DNMT1, DNMT3a, and DNMT3b was found to be increased at 72 h, and decreased at 7 days in microgravity exposed cells. The MS-RAPD analysis revealed that simulated microgravity exposure results in DNA hypomethylation and mutational changes. Gene expression analysis revealed microgravity exposure time-dependent decreased expression of HDAC1. Decreased expression of HDAC1 should result in increased level of acetylated histone H3, however a decreased level of acetylated H3 was observed in microgravity condition, indicating thereby that other HDACs may be involved in regulation of H3 deacetylation. The findings of this study suggest that epigenetic events could be one of the mechanistic bases for microgravity-induced gene expression changes and associated adverse health effects. J. Cell. Biochem. 111: 123–129, 2010. ß 2010 Wiley-Liss, Inc. KEY WORDS: DNA METHYLATION; SIMULATED MICROGRAVITY; EPIGENETICS; RAPD; DNMT1; HDAC1; HISTONE ACETYLATION S everal studies suggest that space flight conditions result in profound changes in various physiological systems causing adverse health effects including cardiovascular changes [Baevsky et al., 2007], bone loss [Vico et al., 2000], muscular atrophy [Hayes et al., 1992], and decline in cellular immune function [Leach et al., 1990]. The mechanism of these microgravity and space flight- associated physiological changes are not well understood. Recent studies have shown that microgravity conditions affect the expression of genes including some of the genes associated with cellular functions. For example, altered expression of cell proliferation and growth factor cascades-associated genes was found in male rats exposed to spaceflight condition (NASA-STS-90 neurolab) for 17 days [Taylor et al., 2002]. A gene chip microarray analysis revealed microgravity-induced changes in the expression of genes belonging to various functional categories including those directly related to immune response, cell proliferation and differentiation, protein folding, transport and degradation, as well as apoptosis [Ward et al., 2006]. Our recent study revealed that simulated microgravity results in decreased expression of genes for DNA repair and cell cycles [Kumari et al., 2009]. While the role of genetic events of DNA mutations in gene expression changes is well established, accumulating evidences suggest that epigenetic changes of DNA methylation and histone modifications play important role in regulation of gene expression [Kang et al., 2001; Roh et al., 2006]. In normal cells, DNA methylation occurs predominantly in repetitive genomic regions, and CpG islands of the promoters generally remain unmethylated [Robertson, 2005]. The maintenance of the DNA methylation patterns is essential for mammalian development and for the normal functioning of the adult organism. Journal of Cellular Biochemistry ARTICLE Journal of Cellular Biochemistry 111:123–129 (2010) 123 Authors have no conflict of interest. *Correspondence to: Dr. Kamaleshwar P. Singh, PhD, Department of Environmental Toxicology, Institute of Environmental and Human Health, Texas Tech University, Lubbock, TX 79409. E-mail: kamaleshwar.singh@tiehh.ttu.edu Received 9 January 2010; Accepted 16 April 2010  DOI 10.1002/jcb.22674  ß 2010 Wiley-Liss, Inc. Published online 12 May 2010 in Wiley Online Library (wileyonlinelibrary.com).