Project Summary/Abstract The serotonergic (5HTergic) system is implicated in a wide range of neurodevelopmental and neuropsychiatric phenomena, including the regulation of mood and stress reactivity. While 5HT actions have been assumed to be mediated exclusively through 5HT receptors and their synaptic effects, recent studies have demonstrated the presence of nuclear pools of 5HT in dorsal raphe serotonergic neurons and forebrain target neurons. Our laboratory has established that 5HT forms covalent bonds with histone H3—resulting in H3 glutamine 5 serotonin (H3Q5ser)–a process known as H3 serotonylation. We have further shown that H3 serotonylation plays key regulatory roles in establishing normal embryonic and adult patterns of brain transcriptional plasticity. However, functional roles for H3 serotonylation during early post-natal brain development, time points encompassing critical periods of neural plasticity, have largely been unexplored, and the impact of environmental stimuli (aberrant or otherwise) on this modification during early life remains unknown. I hypothesize that H3 serotonylation trajectories vary – in a region-specific manner – across the post-natal brain to control critical aspects of neurodevelopmental gene expression and early life stress disrupts these patterns directly influencing vulnerability to stress-related behavioral abnormalities. Under the mentorship of Drs. Ian Maze and Eric Nestler at the Icahn School of Medicine at Mount Sinai, I will address this hypothesis with three distinct aims using a variety of approaches. In Aim 1, I will functionally assess H3 serotonylation dynamics during post-natal development, examine how early life stress disrupts these patterns, and elucidate the causal relationship of this novel histone modification on gene expression regulation using strategic integration of epigenomic and transcriptomic approaches (ie. RNA sequencing and CUT&RUN). In Aim 2, I will characterize how disruptions in post-natal serotonylation-associated gene expression directly result in alterations in brain circuitry leading to increased vulnerability to stress-related behavioral abnormalities using advanced gene-editing techniques and behavioral analyses. In Aim 3, I will characterize brain-wide patterns of H3 serotonylation across post-natal brain development and in response to early like stress using whole brain clearing and imaging techniques (ie. iDISCO+). Overall, this project will provide novel insight into the ways that h3 serotonylation controls brain development and the mechanisms by which disruptions to this posttranslational mark cause aberrant pathophysiological states.