Single-cell transcriptomics identifies Keap1-Nrf2 regulated collective invasion in a Drosophila tumor model

Apicobasal cell polarity loss is a founding event in epithelial-mesenchymal transition and epithelial tumorigenesis, yet how pathological polarity loss links to plasticity remains largely unknown. To understand the mechanisms and mediators regulating plasticity upon polarity loss, we performed single-cell RNA sequencing of Drosophila ovaries, where inducing polarity-gene l(2)gl-knockdown (Lgl-KD) causes invasive multilayering of the follicular epithelia. Analyzing the integrated Lgl-KD and wildtype transcriptomes, we discovered the cells specific to the various discernible phenotypes and characterized the underlying gene expression. A genetic requirement of Keap1-Nrf2 signaling in promoting multilayer formation of Lgl-KD cells was further identified. Ectopic expression of Keap1 increased the volume of delaminated follicle cells that showed enhanced invasive behavior with significant changes to the cytoskeleton. Overall, our findings describe the comprehensive transcriptome of cells within the follicle cell tumor model at the single-cell resolution and identify a previously unappreciated link between Keap1-Nrf2 signaling and cell plasticity at early tumorigenesis. Overall design: Whole-tissue RNA-Seq (2 replicates per genotype; 2 time-points - 24h and 96h) of tjTS>GFP (Control) and tjTS>lglRNAi (Sample) and single-cell RNA-Seq (1 replicate; 1 time-point - 72h) of w1118 cells (Control) of the Drosophila ovary and tjTS>lglRNAi cells (Sample) of the Drosophila ovary. Single-cell RNA-Seq: 10x Genomics CHROMIUM Single Cell 3' Solution V2 Chemistry and Sequencing platform: Illumina NovaSeq 6000.