A novel inducible Cre mouse model for genetic manipulation in nonciliated cells of efferent ductules†

The efferent ductules (ED) function as essential conduits for spermatozoa transport from the rete testis to the epididymis. The nonciliated and ciliated cells within the efferent ductal epithelium are responsible for fluid reabsorption and stirring the luminal fluid to prevent sperm agglutination, respectively. Dysfunction in either cell type can result in obstructive azoospermia. To systematically investigate the molecular mechanisms underlying efferent ductal development and function, we successively developed two novel knock-in mouse models via CRISPR/Cas9-mediated insertion of Cre-P2A or CreERT2-P2A cassettes into the Adgrg2 locus, enabling Adgrg2 promoter-driven co-expression of endogenous Adgrg2 and Cre recombinase. Cre-active tissues were examined in Cre-positive males crossed to Rosa26LacZ or Rosa26tdTomato reporter mice. Adgrg2-Cre mice exhibited embryonic Cre activity, as evidenced by tdTomato fluorescence in embryonic ED, proximal epididymis, and precursor cells, while postnatal males showed widespread genetic recombination across multiple tissues. In contrast, in postnatal Adgrg2-CreERT2 males under tamoxifen administration, Cre activity was prominently present in nonciliated cells within the efferent ductule epithelium and proximal epididymis, with minimal activity in other tissues. These models provide precise tools for cell type- and stage-specific genetic manipulation, facilitating studies on efferent ductal development, fluid homeostasis, and obstructive male infertility. The Adgrg2-CreERT2 line, in particular, offers a unique platform for nonciliated cell-specific genetic studies. This study opens new avenues for understanding the genetic and molecular basis of male reproductive tract function and associated pathologies.