Abstract
During mitosis, the structure of the Endoplasmic Reticulum (ER) displays a dramatic reorganization and remodeling, however, the mechanism driving these changes is poorly understood. Hairpin-containing ER transmembrane proteins that stabilize ER tubules have been identified as possible factors to promote these drastic changes in ER morphology. Recently, the Reticulon and REEP family of ER shaping proteins have been shown to heavily influence ER morphology by driving the formation of ER tubules, which are known for their close proximity with microtubules. Here, we examine the role of microtubules and other cytoskeletal factors in the dynamics of a Drosophila Reticulon, Reticulon-like 1 (Rtnl1), localization to spindle poles during mitosis in the early embryo. At prometaphase, Rtnl1 is enriched to spindle poles just prior to the ER retention motif KDEL, suggesting a possible recruitment role for Rtnl1 in the bulk localization of ER to spindle poles. Using image analysis-based methods and precise temporal injections of cytoskeletal inhibitors in the early syncytial Drosophila embryo, we show that microtubules are necessary for proper Rtnl1 localization to spindles during mitosis. Lastly, we show that astral microtubules, not microfilaments, are necessary for proper Rtnl1 localization to spindle poles, and is largely independent of the minus-end directed motor protein dynein. This work highlights the role of the microtubule cytoskeleton in Rtnl1 localization to spindles during mitosis and sheds light on a pathway towards inheritance of this major organelle.
Highlights
The organization and dynamics of eukaryotic cells during cell division have been extensively studied over the past several decades with key discoveries made with respect to chromosomal partitioning, and the changes of the cytoskeleton network [1,2,3]
As well as prior studies [34] show a movement of the spindle poles during nuclear envelope breakdown (NEB) and spindle formation, this movement did not compromise our method for regions of interests (ROIs) selection since it occurs when spindles are already morphologically visible (Fig 1A and 1B)
We examined the movement of spindle poles just prior to the start of mitosis through metaphase in RFP-KDEL and Reticulon-like 1 (Rtnl1)-GFP embryos injected with fluorescently labeled tubulin
Summary
The organization and dynamics of eukaryotic cells during cell division have been extensively studied over the past several decades with key discoveries made with respect to chromosomal partitioning, and the changes of the cytoskeleton network [1,2,3]. Lesser understood is the dynamics of the cytoplasmic organelles during cell division. Studies have investigated organelle dynamics during mitosis, including the disassembly and reassembly of the Golgi apparatus [4,5] and the structural changes and localization of the Endoplasmic Reticulum (ER) [6,7]. De Cruz is supported through a National Science Foundation (NSF) CAREER award, 1553695. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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