SPE-55 and VHA-18 are essential for FB-MO complex assembly and transport during spermatogenesis in C. elegans.

The spatiotemporal control of asymmetric positioning of cellular cargoes, such as macromolecules and organelles, during spermatogenesis is essential for producing functional sperm in most animal species. However, the regulatory mechanisms that regulate cargo sorting and transporting remain largely unknown. Here, we demonstrate that the casein-like kinase SPE-55 localized on the surface of a membranous organelle (MO) is required for the SPE-18 localization/puncta formation and facilitates the assembly of major sperm protein (MSP) condensate, named the fibrous body (FB), in spermatocytes during spermatogenesis in Caenorhabditis elegans. Furthermore, we discover that the H subunit of v-ATPase, which is localized on MOs, played a crucial role in maintaining MO morphology and function, promoting FB-MO connections, and thus ensuring efficient MSP sorting during spermatogenesis. Therefore, the interplay between the membraneless condensate FBs mediated by SPE-55 and the membrane-bound MOs regulated by VHA-18 provides a coordinated mechanism for cargo sorting and directional transport during spermatogenesis in C. elegans.