Studies in animal models linked gestational exposures to endocrine disrupting chemicals (EDCs) with the onset of disease in exposed and unexposed descendants. Many groups found such transgenerational effects of chemical exposures, which were proposed to be examples of epigenetic inheritance. Transgenerational effects of environmental exposures have substantial support in the literature. Yet the concept that responses to environmental exposures can be transmitted to subsequent generations through the germline without DNA mutations remains controversial because the underlying mechanisms have not been explained satisfactorily. Understanding how effects of environmental exposures are transmitted to unexposed generations without DNA mutations is a fundamental, unanswered question in biology. We developed a highly reproducible animal model for transgenerational inheritance of obesity. When pregnant F0 mouse dams were treated with environmentally-relevant (nM) doses of TBT via their drinking water throughout gestation, increased fat accumulation was detected in F1-F4 generation male descendants. Affected TBT-group males developed a transgenerational “thrifty phenotype”: they were resistant to fat loss during fasting, rapidly gained weight when dietary fat was increased and retained this fat even after being returned to a normal, low-fat diet. Our published and preliminary results led us to propose a new model for transgenerational inheritance - that prenatal TBT exposure altered higher-order chromatin structure (HOCS), changing secondary epigenetic modifiers that inhibited expression of insulin degrading enzyme (Ide) causing diet-induced hyperinsulinemia and obesity. Here we propose a comprehensive series of experiments designed to determine exactly how exposure of pregnant F0 dams to TBT alters HOCS in F1-F3 primordial germ cells (PGCs), why these changes are inherited, rather than reversed to the normal state, how these changes affect lower-level epigenetic regulators controlling expression of Ide and why does the phenotype only occur in males. Aim 1 will identify mechanisms that drive changes HOCS near the Ide gene and how these interact with lower-level epigenetic regulators to modulate Ide expression in the adult liver. Aim 2 tests whether epigenetic interventions, such as dissolving the HOCS alterations or releasing Ide expression from repression in PGCs or adults can prevent or reverse the transgenerational predisposition to male-specific metabolic phenotypes. Deciphering the underlying mechanisms will have profound implications for how the field views transgenerational inheritance and how future experiments are planned and conducted. This new understanding will be critical to explaining the etiology of non-communicable diseases such as obesity and type 2 diabetes, targeting their causes and ameliorating their effects. Our results will have broad implications for understanding epigenetic transmission of the effects of environmental stressors and ...