Normal spindle positioning in the absence of EBPs and dynein plus-end tracking inC. elegans

The position of the mitotic spindle is tightly controlled in animal cells, as it determines the plane and orientation of cell division. Interactions between cytoplasmic dynein at the cortex and astral microtubules generate pulling forces that position the spindle. In yeast, dynein is actively delivered to the cortex through microtubule plus-end tracking complexes. In animal cells, an evolutionarily conserved Gα-GPR-1/2 Pins/LGN –LIN-5 NuMA cortical complex interacts with dynein and is required to generate pulling forces, but the mechanism of dynein recruitment to the cortex is unclear. Using CRISPR/Cas9-assisted recombineering, we fluorescently labeled endogenous DHC-1 dynein in C. elegans. We observed strong dynein plus-end tracking, which depended on the end-binding protein EBP-2. Complete removal of the EBP family abolished dynein plus-end tracking but not LIN-5-dependent cortical localization. The ebp-1/2/3 deletion mutant, which was viable and fertile, showed increased cortical microtubule retention; however, pulling forces and spindle positioning were normal. These data indicate that dynein recruited from the cytoplasm creates robust pulling forces.