Sertoli cells are the epithelial supporting cells of the seminiferous tubules of male testes, producing steroids, nutrients, and signaling molecules and thus playing a critical role in regulating spermatogenesis (Ni et al., 2019; O'Donnell et al., 2022; Wu et al., 2020). The actin cytoskeleton plays a crucial role in supporting spermatogenesis in the mammalian testis (Wang et al., 2020). The blood-testis barrier (BTB) formed by specialized junctions between adjacent Sertoli cells in the seminiferous epithelium is essential for sperm development. Tight junctions connecting Sertoli cells and the BTB are maintained by the actin filament network (Li et al., 2016). Sertoli cells are also responsible for the phagocytosis of apoptotic germ cells (Penberthy et al., 2018; Wu et al., 2022). However, the mechanism underlying the regulation of the cytoskeleton and phagocytosis of Sertoli cells remains largely unknown.

Disheveled associated activator of morphogenesis 2 (DAAM2) belongs to the DAAM family that regulates actin polymerization (Breitsprecher and Goode, 2013; Higashi et al., 2008). DAAM1 and DAAM2 share a conserved formin homology 2 (FH2) domain with actin nucleation activity, suggesting a role in cell motility (Kida et al., 2004). DAAM2 has been reported to be mainly involved in nervous system development. DAAM2 variants may cause nephrotic syndrome via actin dysregulation (Schneider et al., 2020). DAAM2 also regulates myelin structure and the oligodendrocyte actin cytoskeleton (Cristobal et al., 2022). The DAAM2-VHL-Nedd4 axis has been shown to govern oligodendrocyte differentiation (Ding et al., 2020). Interestingly, DAAM2 plays a key role in the Wnt signaling pathway (Lee et al., 2015; Lee and Deneen, 2012). It is known that Wnt signaling is essential for male germ development, spermatogenesis, and fertility. Disruption of this signaling may cause pathological changes in testes and sperm cells (Takase and Nusse, 2016; Xue et al., 2021). Based on these findings, we hypothesized that DAAM2 may be involved in the regulation of the cytoskeleton in spermatogenesis.

To test our hypothesis, we examined DAAM2 expression in rat testis, spermatogenic cells, and Sertoli cells. Knockdown of DAAM2 was performed to investigate the regulatory role of DAAM2 in the cytoskeleton and phagocytosis of Sertoli cells in vitro and in vivo. Further, RNA sequencing (RNA-seq), pull-down mass spectrometry, and bioinformatics analyses were conducted to identify potential genes, proteins, and signaling pathways associated with DAAM2 in rat Sertoli cells. Our study suggests that DAAM2 plays an essential role in spermatogenesis by regulating cytoskeleton organization and phagocytosis of Sertoli cells.