Decoding the dynamics and regulation of parental allelic expression at single-cell resolution

Project Summary The inheritance of both the maternal and paternal genes is required for normal development to proceed. Yet, we are not equal products of both parents’ genes. With exclusive expression of specific genes from only one parent, allelic effects have been suggested to influence genetic architecture and disease risk, but little is known about their contributions to development. Fundamental obstacles of comprehensively understanding the parental genetic inheritance and its regulation during development are limitations in incomplete coverage of each cell within developing embryos, and the accuracy of differentiation between allele sequences. As the closest living relatives of vertebrates, the ascidians C. intestinalis and C. savignyi provide the right degree of simplicity among multicellular organisms and extreme divergence of the maternal and paternal genomes for overcoming these limitations. Here I propose to use hybrid ascidians to elucidate the regulatory grammar of allele-specific gene expression during development at single cell resolution. During the K99 phase, I will work on Aim 1 and initiate Aim 2. Based on my preliminary results, I have reconstructed the reference lineage map of temporal expression profiles for hybrid embryos using single cell RNA-Seq technology. Due to the small cell numbers of ascidians, I expect to obtain comprehensive coverage of every cell type during development, including rare neuronal subtypes. In Aim 1, based on the extreme divergence between parental genomic sequences in hybrid embryos, I will characterize allele-specific expression of each defined cell lineage, and develop novel approaches that are able to simultaneously profile gene expression and chromatin accessibility in single cells, which will link parental expression patterns with putative cis-regulatory DNA sequences. Upon the completion of Aim 1, I expect to establish potential associations of allelic-specific expression and the specification and morphogenesis of individual cells and cell lineages. During the R00 phase, based on my training gained during the K99 phase, I will work on the highly innovative Aim 2, in which I plan to develop the spatial transcriptomics technique in order to add another dimension to dissect the allele-specific expression pattern and its regulation in the context of cell-cell contacts. Upon successful completion of Aim 1 and Aim 2, this data-rich approach will greatly enhance our understanding of parental genetic inheritance and its regulation, and eventually let us move closer to a holistic and quantitative understanding of embryonic development. It also outlines an extensive career development plan for me to complete my training under the mentorship of Prof. Levine and the transition to an independent academic position by establishing a multi-disciplinary research program.