PROJECT SUMMARY Genomic imprinting is an epigenetic phenomenon in which somatic genes are mono-allelically expressed depending on their parental origin. Imprinted genes play critical roles in prenatal and postnatal development, including growth, metabolism, and neurodevelopment. Thus, genomic imprinting is essential for organismal viability, and disrupted imprinting frequently results in human disease. Imprinted genes are abundantly expressed in the brain; however, investigation into their regulatory mechanisms and physiological significance in the brain has been limited. Emerging evidence suggests that chromatin structure at imprinted domains may differ between the two parental alleles, implying a mechanistical model where differential chromatin structures may confer allele-specific regulatory roles. Nevertheless, the exploration of the impact of allele-specific chromatin structure on imprinted gene regulation is still in its early stages. The Mest-Copg2 imprinted domain, located on mouse chromosome 6 and human chromosome 7, presents a neuron-specific imprinting pattern and is associated with developmental and postnatal growth defects and atypical maternal behavior. In preliminary data, striking differences in parental chromatin structures at the Mest-Copg2 domain were observed. Yet, the extent to which chromatin structures play a role in imprinted expression of Mest-Copg2 remains entirely uncharted. Thus, this proposed research delves into the regulatory mechanism and functional role of the Mest-Copg2 domain. The focus of Aim 1 will be elucidating how allelic chromatin structures are established in the Mest- Copg2 domain. Epigenetic perturbation tools will be used to establish what are the basis of allelic chromatin structures in this domain. Aim 2 will identify cis-regulatory elements modulating Mest-Copg2 imprinted expression. CRISPRi experiments will be conducted in hybrid primary neurons to validate the activities of cis- regulatory elements in mediating imprinted Mest-Copg2 expression. In Aim 3, the physiological implication of neuron specific Copg2 imprinting will be investigated to provide insights into the role of genomic imprinting in neurons. These results will provide a novel regulatory mechanism mediated by chromatin structure, as well as functional implication of genomic imprinting in the Mest-Copg2 domain. The Harvard University will provide a vibrant research atmosphere with ample opportunities for inter- and intradepartmental collaborations. This environment, coupled with the guidance of Dr. Whipple and Dr. Dulac, will provide valuable training opportunities to develop scientific and professional skills necessary for becoming an independent researcher.

PROJECT NARRATIVE This project aims to unravel the intricacies of imprinted gene regulation through the investigation of chromatin organizations. By focusing on neuron-specific imprinted genes, the results will elucidate the molecular mechanisms and physiological functions of imprinted genes in the brain. The insights gained from this research will enhance our understanding of human imprinting disorders and related neuropathological conditions.