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.

Grant Number: 5R00HD102584-04
NIH Institute/Center: NIH
Principal Investigator: Chen Cao