Identifying And Profiling Modalities Of Alternative Polyadenylation Usage In Single Cells

A large number of 3’ end sequencing or RNA-seq transcriptome studies have found that as many as 70%of genes in animals and plants can undergo alternative polyadenylation(APA),which promotes the diversity and complexity of transcriptome.A variety of APA usage modalities and tissue-specific usages of APA have been found in the process of human disease occurrence,embryonic development and differentiation,which provides important information for the study of mechanisms of gene expression regulation.At present,APA dynamics are mainly analyzed at the overall level of cell populations while rarely at the single-cell level.Existing related studies are either limited to specific cell types or a small number of cells,or focus on the identification of APA sites between cell types;there is a lack of in-depth research on APA usage modalities and related effects at the single-cell level.This paper proposes a scAPAmod analysis framework for identifying and analyzing APA usage modalities at the single-cell level.The framework adopts the polyA usage index to characterize the usage of proximal APA sites at the single-cell level.The scAPAmod framework identifies usage modalities(Unimodal,Bimodal and Multimodal)based on expression of single-cell APA sites using Gaussian mixture model.Using single-cell mouse sperm data,the dynamic changes of APA usage modalities in different cell stages and development stages were analyzed.Moreover,APA usage modalities in different genomic regions and preference of specific polyadenylation sites in single cells were also characterized.In addition,cell-type specific modalities and changes in APA usage modalities among cell types were profiled.Results show that compared with the existing tools,scAPAmod can accurately identify tissue specific APA modalities in single cells and discovered different APA usage modalities at different stages.At the same time,it was also found that the APA usage modalities of same gene in 3’ UTR and non 3’ UTR were different,and relevant genes were enriched in different regulatory mechanisms and functional pathways.In summary,scAPAmod provides a new framework for characterizing modalities of APA usages at the single-cell level,which helps to understand the differences in APA usage modalities at different cell differentiation stages and to cell-cell heterogeneity of gene expression.