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Xiuqian Mu, Yichen Ge, Fuguo Wu, Jonathan Bard, Julien Kann, Donald Yergeau, Darshan Sapkota, Zihua Hu, jie wang, Tao Liu; Investigating the developmental trajectory of retinal ganglion cells by scRNA-seq and and scATAC-seq. Invest. Ophthalmol. Vis. Sci. 2020;61(7):4018.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal ganglion cells (RGCs) are the first retinal cell type to form during development, arising from multipotent retinal progenitor cells (RPCs). The purpose of this study is to understand the transcriptomic and epigenetic changes along the developmental trajectory from naive RPCs to differentiated RGCs.
To achieve our objectives, we first performed regular RNA-seq with E14.5 wild-type and Atoh7-null mouse retinas . We then performed scRNA-seq and scATAC-seq on E13.5/E14.5 mouse retinal cells. We initially performed scRNA-seq with wild-type and Atoh7-null cells at E13.5. We then purified specific wild-type and Atoh7-null retinal cell populations by FACS using two fluorescent protein labeled mouse lines and performed scRNA-seq and scATAC-seq.
By comparing the transcriptomes of wild-type and Atoh7-null retinas, we were able to identify significantly more genes altered in expression due to Atoh7 deletion and expand our understanding of the scope of downstream events. We then performed scRNA-seq on E13.5 wild-type and Atoh7-null retinal cells. Clustering analysis correctly grouped known cell types at this developmental stage and provided clear developmental trajectories of each retinal cell type born at this stage. Analysis of the Atoh7-null retina not only confirmed that the RGC lineage was affected but also identified the affected genes/pathways involved in the different cell states. Using the sorted cells, we have now performed scATAC-seq to assess shifts of the epigenetic landscape along the RGC developmental trajectory. Our preliminary analysis indicated that scATAC-seq data matched well with the scRNA-seq data, and provide insights into the changes in chromatin structure and enhancer activities that underly the progression of RGC differentiation.
Using single-cell technologies and mouse genetics, we are investigating the transcriptomic and epigenetic changes along the RGC developmental trajectory. Our study provides an unprecedented and detailed view of retinal cell differentiation at cellular, transcriptomics, and epigenomics levels.
This is a 2020 ARVO Annual Meeting abstract.
Pseudotime analysis of the developmental trajectories in the wild-type and Atoh7-null (Mutant) retinas based on scRNA-seq data.
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