Acentrosomal Pole Stability and Maintenance in Caenorhabditis elegans Oocyte Meiotic Spindles

Cavin-Meza, Gabriel Joseph

doi:https://doi.org/10.21985/n2-3vvf-2n52

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Acentrosomal Pole Stability and Maintenance in Caenorhabditis elegans Oocyte Meiotic Spindles

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Meiosis is a specialized form of cell division that occurs to generate sperm and eggs with unique sets of paternal or maternal DNA; this process shuffles genetic information to promote the amazing variation that we observe in living organisms. In order to carry out two rounds of DNA separation with high fidelity, meiosis uses many of the same mechanisms and proteins that are critical in mitosis. However, the meiotic divisions of eggs (oocytes) in many organisms lack protein-rich structures known as centrosomes. These centrosomes serve as organizing centers for microtubules, creating a bipolar structure that forms around chromosomes and facilitates their segregation (the spindle). Notably, acentrosomal oocyte meiotic spindles are still bipolar; the mechanisms that allow these spindles to assemble and stabilize their poles are not well understood. Proteins that organize bipolar spindles during mitosis have been prime candidates for study in meiotic spindles. Multiple microtubule-associated proteins (MAPs) have been well-studied in mitosis and some of their functions have been shown to be conserved in meiosis. However, previous studies with essential proteins have been complicated due to off-target effects generated from traditional depletion methods such as genetic knockouts or RNA interference (RNAi); experimental limitations made it difficult to determine whether the activity of these essential proteins is only important during formation of meiotic spindles or must be maintained throughout the entire process. Here, I have applied the auxin-inducible degron (AID) system to study the functions of three essential mitotic proteins in oocytes. I rapidly removed DHC-1 (dynein), ZYG-9 (XMAP215), or AIR-1 (Aurora A kinase) from C. elegans oocytes, revealing that all three proteins are not only required to assemble stable acentrosomal poles, but also to maintain them throughout meiosis. Despite having similar localizations in the meiotic spindle, each depletion condition yielded distinct phenotypes that manifested quickly after auxin treatment. Further experimentation with dynein depletion uncovered the presence of a redundant outwards sorting force provided by the kinesin-5 homolog BMK-1. We used various experimental approaches to reveal that ZYG-9 is extremely dynamic in the meiotic spindle and likely contributes to microtubule dynamics across the entire spindle rather than locally to acentrosomal poles. Finally, our preliminary work with AIR-1 (Aurora A) suggests that stability of overlapping MT bundles near chromosomes is directly regulated by AIR-1.Our work has expanded our knowledge of the forces and protein interactions that are involved in meiotic spindle organization in C. elegans. Errors in oocyte meiosis have been shown to be correlated to improper segregation of DNA and generate inviable embryos; understanding the fundamental mechanisms of how these meiotic spindles achieve bipolarity is key to building a better model of what factors can lead to loss of faithful segregation.

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