Endoplasmic reticulum membranes are continuously required to maintain mitotic spindle size and forces.

Membrane organelle function, localization, and proper partitioning upon cell division depend on interactions with the cytoskeleton. Whether membrane organelles also impact the function of cytoskeletal elements remains less clear. Here, we show that acute disruption of the ER around spindle poles affects mitotic spindle size and function in Drosophila syncytial embryos. Acute ER disruption was achieved through the inhibition of ER membrane fusion by the dominant-negative cytoplasmic domain of atlastin. We reveal that when centrosome-proximal ER membranes are disrupted, specifically at metaphase, mitotic spindles become smaller, despite no significant changes in microtubule dynamics. These smaller spindles are still able to mediate sister chromatid separation, yet with decreased velocity. Furthermore, by inducing mitotic exit, we found that nuclear separation and distribution are affected by ER disruption. Our results suggest that ER integrity around spindle poles is crucial for the maintenance of mitotic spindle shape and pulling forces. In addition, ER integrity also ensures nuclear spacing during syncytial divisions.

Astral microtubule cross-linking safeguards uniform nuclear distribution in the Drosophila syncytium.

The early insect embryo develops as a multinucleated cell distributing the genome uniformly to the cell cortex. Mechanistic insight for nuclear positioning beyond cytoskeletal requirements is missing. Contemporary hypotheses propose actomyosin-driven cytoplasmic movement transporting nuclei or repulsion of neighbor nuclei driven by microtubule motors. Here, we show that microtubule cross-linking by Feo and Klp3A is essential for nuclear distribution and internuclear distance maintenance in Drosophila. Germline knockdown causes irregular, less-dense nuclear delivery to the cell cortex and smaller distribution in ex vivo embryo explants. A minimal internuclear distance is maintained in explants from control embryos but not from Feo-inhibited embryos, following micromanipulation-assisted repositioning. A dimerization-deficient Feo abolishes nuclear separation in embryo explants, while the full-length protein rescues the genetic knockdown. We conclude that Feo and Klp3A cross-linking of antiparallel microtubule overlap generates a length-regulated mechanical link between neighboring microtubule asters. Enabled by a novel experimental approach, our study illuminates an essential process of embryonic multicellularity.