Single-cell transcriptomic profiling of bam mutant tumor reveals germline heterogeneity and gcrf1 as a modulator in Drosophila germ cells

The bam mutant ovary of Drosophila represents a classic tumor model caused by germline stem cell (GSC) differentiation defects. To date, its molecular and genetic features have rarely been characterized in detail at the single-cell resolution. Here, we performed single-cell RNA sequencing (scRNA-seq) to comprehensively delineate the transcriptomic landscape and identify distinct germline cell types in bam mutant ovaries by using in situ hybridization. Differentially expressed gene analysis and PAGA plots reveal different transcriptional profiles and developmental relationships in ovarian cells. Based on the expression pattern of eggpl, a useful marker for undifferentiated germ cell identity, and morphological differences in bam mutant ovarioles, two potentially distinct germ cell states are distinguished. Comparative single-cell analysis reveals the potential regulatory network and cellular communication in subclusters of undifferentiated germ cells, and contributes to the identification of gcrf1 as a novel marker gene for female GSC, which involves in the regulation of early germ cell proliferation and Drosophila fertility. Collectively, our study not only provides insights into tumorigenesis caused by GSC differentiation defects but also offers a valuable transcriptomic resource that can be mined for the reproductive features of bam mutant tumors by community.