Regulation of outer kinetochore assembly during meiosis I and II by CENP-A and KNL-2/M18BP1 in C. elegans oocytes.

During cell division, chromosomes build kinetochores that attach to spindle microtubules. Kinetochores usually form at the centromeres, which contain CENP-A nucleosomes. The outer kinetochore, which is the core attachment site for microtubules, is composed of the KMN network (Knl1c, Mis12c, and Ndc80c complexes) and is recruited downstream of CENP-A and its partner CENP-C. In C. elegans oocytes, kinetochores have been suggested to form independently of CENP-A nucleosomes. Yet kinetochore formation requires CENP-C, which acts in parallel to the nucleoporin MEL-28^ELYS. Here, we used a combination of RNAi and Degron-based depletion of CENP-A (or downstream CENP-C) to demonstrate that both proteins are in fact responsible for a portion of outer kinetochore assembly during meiosis I and are essential for accurate chromosome segregation. The remaining part requires the coordinated action of KNL-2 (ortholog of human M18BP1) and of the nucleoporin MEL-28^ELYS. Accordingly, co-depletion of CENP-A (or CENP-C) and KNL-2^M18BP1 (or MEL-28^ELYS) prevented outer kinetochore assembly in oocytes during meiosis I. We further found that KNL-2^M18BP1 and MEL-28^ELYS are interdependent for kinetochore localization. Using engineered mutants, we demonstrated that KNL-2^M18BP1 recruits MEL-28^ELYS at meiotic kinetochores through a specific N-terminal domain, independently of its canonical CENP-A loading factor activity. Finally, we found that meiosis II outer kinetochore assembly was solely dependent on the canonical CENP-A/CENP-C pathway. Thus, like in most cells, outer kinetochore assembly in C. elegans oocytes depends on centromeric chromatin. However, during meiosis I, an additional KNL-2^M18BP1 and MEL-28^ELYS pathway acts in a non-redundant manner and in parallel to canonical centromeric chromatin.