RNA velocity quantification of single-cell transcriptomics
Full title
Utilising RNA velocity quantification of single cell transcriptomics to study differential cellular ontogeny in a single stage sample across related species.
Brief project outline
The precise timing of development is essential for the formation of complex traits. This is particularly relevant for regulatory processes, such as transcriptional networks involved in cell-fate specification, as subtle changes in their timing can result in dramatic phenotypes. A key unanswered question in the field of brain evolution is why marsupials have a cerebral cortex that is less densely populated with neurons than in eutherian (placental) mammals of similar size. Our preliminary data suggest that differences in developmental neurogenesis, such as faster cell cycle exit and differentiation in marsupials, might underlie this. Moreover, in eutherians, basal progenitor cells further amplify cortical neurogenesis, however in marsupials, basal progenitors constitute a much smaller proportion of progenitor cells. Recent evidence from the literature has elucidated that the state of a cell's transcriptome and proteome can control and is reflective of the tempo of development between species (Rayon et al. 2020). Here, we propose to utilise this discovery to pioneer new understandings of mammalian brain evolution, by combining scRNA-seq with a novel tool for dynamic mRNA maturation analysis (RNA velocity).
Genomics-based innovative aspect of proposal
In this project, we aim to use the single cell sequencing capabilities of the GIH to generate the world's first marsupial brain scRNA-seq dataset to allow for in-depth molecular and evolutionary analyses. To date, only one single cell study has been performed on marsupials and involved whole embryos (Mahadevaiah et al. 2020). In collaboration with the GIH sequencing facility, and using a novel mammalian model of neurodevelopment that our lab has established, we will optimise protocols for cell isolation and sequencing from flash frozen dunnart and mouse brain tissue. We will subsequently develop a pipeline that performs two sets of analyses. First, calculate RNA velocity of each cell type and second, compare single cell velocity measures between orthologous cell clusters across species, as a measure of cell-specific heterochrony. This analytical pipeline will involve a mathematical comparison of RNA velocity elements, and will involve collaborative work with a GIH bioinformatician as well as consultation with a mathematician or computational scientist. In addition, the pipeline will analyse velocity profiles across developmental stages within each species, to provide a statistical measure of RNA profile dynamics during development of each cell type.
Broad applicability of the technique
This project will help to keep UQ at the forefront of molecular and neurodevelopmental research, given the novel nature of the proposed marsupial dataset as well as the generation of a new bioinformatic tool to advance the study of the roles of developmental timing on complex trait formation. Furthermore, the scripts can also be applied to the study of disease and be used to perform comparative analysis of mRNA maturation dynamics across disease and control conditions. The pipeline will, therefore, be of interest to any UQ groups working not only on development or evolution, but also on genetic or acquired disease datasets.
