Soma-germline communication drives sex maintenance in the Drosophila testis.

In adult gonads, disruption of somatic sexual identity leads to defective gametogenesis and infertility. However, the underlying mechanisms by which somatic signals regulate germline cells to achieve proper gametogenesis remain unclear. In our previous study, we introduced the chinmo^{Sex Transformation} (chinmo^ST) mutant Drosophila testis phenotype as a valuable model for investigating the mechanisms underlying sex maintenance. In chinmo^ST testes, depletion of the Janus Kinase-Signal Transducer and Activator of Transcription downstream effector Chinmo from somatic cyst stem cells (CySCs) feminizes somatic cyst cells and arrests germline differentiation. Here, we use single-cell RNA sequencing to uncover chinmo^ST -specific cell populations and their transcriptomic changes during sex transformation. Comparative analysis of intercellular communication networks between wild-type and chinmo^ST testes revealed disruptions in several soma-germline signaling pathways in chinmo^ST testes. Notably, the insulin signaling pathway exhibited significant enhancement in germline stem cells (GSCs). Chinmo cleavage under targets and tagmentation (CUT&Tag) assay revealed that Chinmo directly regulates two male sex determination factors, doublesex (dsx) and fruitless (fru), as well as Ecdysone-inducible gene L2 (ImpL2), a negative regulator of the insulin signaling pathway. Further genetic manipulations confirmed that the impaired gametogenesis observed in chinmo^ST testes was partly contributed by dysregulation of the insulin signaling pathway. In summary, our study demonstrates that somatic sex maintenance promotes normal spermatogenesis through Chinmo-mediated conserved sex determination and the insulin signaling pathway. Our work offers new insights into the complex mechanisms of somatic stem cell sex maintenance and soma-germline communication at the single-cell level. Additionally, our discoveries highlight the potential significance of stem cell sex instability as a novel mechanism contributing to testicular tumorigenesis. [ABSTRACT FROM AUTHOR]