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Single-Cell Study Explores Transcriptome in Developing Mouse Cerebellum

NEW YORK (GenomeWeb) – Using single-cell RNA sequencing, a team led by researchers at St. Jude Children's Research Hospital has profiled transcriptome patterns in tens of thousands of cells in the cerebellum of the developing mouse brain and has made the data available through an interactive web-based tool called Cell Seek.

"We wanted to build an easy-to-use interface that allowed labs without bioinformatics capability to mine the data," co-senior and co-corresponding author Charles Gawad, a researcher in the departments of oncology and computational biology at St. Jude, said in a statement. "With Cell Seek, they can easily track the developmental processes they are interested in and use the insights to inform their experiments to study cerebellar development and disease."

For their analysis, described online today in Current Biology, Gawad and his colleagues performed RNA-seq on 39,245 individual cells isolated from the mouse cerebellum across a dozen time points during embryonic development or shortly after birth. With these data, they began teasing out shifting gene expression and gene regulatory features in the cerebellum.

The cerebellum is involved in motor coordination, executive functions, language processing, and other higher functions in humans, the team explained. It is altered or affected in conditions ranging from Joubert syndrome to pediatric cancers, such as medulloblastoma, astrocytoma, or ependymoma.

"If we can capture cells in these different developmental states, we can begin to understand pediatric diseases that happen as a result of abnormal cerebellar development," Gawad said. "Also, since most brain tumors in young children occur in the cerebellar region, this will help us identify the cells of origin for different brain tumors and brain tumor subtypes."

The researchers relied on on 10x Genomics Chromium and Illumina HiSeq 2500 instruments to sequence the RNA of single cerebellum cells isolated at eight embryonic stages, between embryonic days 10 and 17, and four postnatal time points, reaching out to postnatal day 10.

Using hierarchical clustering and other analytical approaches, the team explored transcriptional features in 39,245 cells that met the quality control criteria, representing a wide range of cerebellar cell lineages with previously reported gene expression markers. In particular, the data offered a look at the activity of known transcriptional regulators, and insights into potentially new regulatory features.

In their proof-of-principle analyses, the researchers used the single-cell RNA-seq data to track glutamatergic progenitor cell differentiation into distinct mature cell types in the mouse cerebellum. In particular, they found surges of transcription factor cluster activation as the progenitor cells morphed into glutamatergic cerebellar nuclei or granule neuron progenitors.

"This analysis showed us that the developmental program for these cells was even more complex than we previously appreciated," Gawad said.

Using the data and Cell Seek interface, the authors conducted similar analyses on GABAergic cell populations in the mouse cerebellum. They noted that "[b]y focusing our analyses on specific cell-fate decisions, novel candidate [transcription factors] expressed during cerebellar development add to the knowledge acquired by previous lineage tracing and gene expression studies performed on bulk populations, to provide a more complete view of cell-type-specific transcriptional landscapes governing cerebellar development."