PROJECT SUMMARY The overarching goal of the research presented in this application is to understand how environmental chemicals trigger a deregulation of the epigenome that can resist epigenetic reprogramming in germ cells and therefore can become heritable. Inorganic arsenic (iAs) is a model epigenetic toxicant owing to its well described impact on global DNA hypomethylation coinciding with a reduction in the levels of the universal methyl donor SAM, used towards DNA and histone methylation. iAs is also a chemical with well-established transgenerational epigenetic inheritance effects, producing heritable reproductive and metabolic dysfunctions and neurobehavioral outcomes for multiple generations. However, iAs shows remarkable complexity in its epigenetic impact since even in the context of global DNA hypomethylation, some loci show hypermethylation and the effect on histone methylation are non- uniform with many methylated histone marks showing increases while others show a decrease. It is unclear how iAs causes such varied epigenetic effect and how these effects are maintained during reprogramming. Here, we propose to leverage a unique model of germ cell reprogramming based on the step-wise differentiation of mouse Embryonic Stem Cells (ESCs) into Primordial Germ Cell-Like Cells (PGCLCs). PGCLCs undergo epigenetic reprogramming in vitro and are transcriptionally and functionally similar to in vivo PGCs since they generate viable offspring upon transplantation in vivo. We will build on compelling preliminary data showing that in mESCs, at levels where sodium arsenite does not cause a significant increase in ROS levels, a pronounced metabolic impact on the methionine cycle and on the transsulfuration pathways are observed concomitant with its epigenetic impact. We also show that two enzymes crucial for the one-carbon and iAs metabolism, MAT2A and AS3MT, respectively, are strongly upregulated in response to iAs and translocate to the nucleus where they associate