Brown adipose tissue (BAT) is positively associated with cardiometabolic health in humans. Strategies focusing on activating BAT in order to burn excess energy may have significant value in the context of obesity (Becher et al., 2021). BAT is highly abundant in newborns to maintain body temperature but deteriorates at differing rates as we age (Zoico et al., 2019), limiting the therapeutic approach of tissue activation. The individual variability in BAT depots in adulthood remains an open question in the field, with exposure to environmental factors potentially playing a role. For example, parental cold exposure has recently been shown to have transgenerational effects on offspring thermogenesis (Oelkrug et al., 2020; Sun et al., 2018). Our laboratory has bred mice over several generations at thermoneutrality, creating thermo-naïve animals, overcoming the cold-induced transgenerational programming of BAT thermogenesis. Preliminary data show that cold exposure during infancy protects against diet-induced obesity in adulthood. These data suggest that cold-exposure during puberty imprints a thermogenic memory in adipocytes which can mitigate the effects of diet-induced obesity in later life. As humans experience an involution of their BAT after infancy, cold-induced programming of enhanced BAT capacity and thermogenic function bares translational significance to combat cardiometabolic diseases in adulthood. The aim of the proposed research is to identify the underlying molecular mechanisms which maintain BAT thermogenesis beyond infancy and uncover the cold-induced molecular memory imprinted during puberty which primes thermogenesis in adulthood. To this end, single-nuclear and ATAC-sequencing has been performed on these animals to determine epigenetic and cell-specific alterations underlying the observed phenotype. Additionally, single-nuclear and ATAC-seq will additionally be performed on Marsupial animals who have been subjected to similar environmental stimuli during development to assess species differences. Furthermore, we will investigate other tissues (skeletal muscle, WAT) to further our understanding of this priming effect. Hence, the transcriptomic and epigenomic analysis are vital to the success of this project.