Beyond Differential Expression: Embracing Cell-to-Cell Variability in Single-Cell Gene Expression Data Analysis

Single-cell RNA sequencing (scRNA-seq) has revolutionized our understanding of cellular variability by capturing gene expression profiles of individual cells. The importance of cell-to-cell variability in determining and shaping cell function has been widely appreciated. Nevertheless, differential expression (DE) analysis remains a cornerstone method in analytic practice, by focusing exclusively on mean expressional differences, current computational analyses overlook the rich information encoded by variability within the single-cell gene expression data. Thus, there is a need for more relevant approaches to assess data variability rather than the mean in order to offer a deeper understanding of cellular systems. Here we present spline-DV, a novel statistical framework for differential variability (DV) analysis using scRNA-seq data. The spline-DV method identifies genes exhibiting significantly increased or decreased expression variability among cells derived from two experimental conditions. Case studies show that DV genes identified using spline-DV are representative, and functionally relevant in the tested cellular conditions including obesity, fibrosis, and cancer.