MIS12 Is Required for Kinetochore‐Microtubule Attachment in Oocyte Meiosis

ABSTRACT Accurate kinetochore‐microtubule (K‐MT) attachment is crucial for chromosome segregation in oocyte meiosis. The NDC80 complex, a subcomplex of the outer kinetochore KNL1‑MIS12‑NDC80 (KMN) network, is canonically responsible for K‑MT attachment. Here, by employing a conditional knockout (cKO) mouse model, combined with protein localization analysis for MIS12 following chromosome spreading, we demonstrate that endogenous MIS12 localizes to kinetochores in both mouse and human oocytes, and depleting MIS12 severely disrupts K‐MT attachment in both meiotic divisions, independent of outer kinetochore protein NDC80. Crucially, we identified that MIS12 directly interacts with β‐tubulin (including TUBB3 and TUBB5), which is essential for microtubule attachment to kinetochores, revealing a previously unknown mechanism of K‐MT interaction. Furthermore, we definitively show that MIS12 is required for spindle assembly checkpoint (SAC) signaling by stabilizing the KNL1 assembly. Our findings not only resolve the previous controversy by establishing the canonical and essential role of kinetochore‐localized MIS12 in oocytes but also redefine its molecular function through its direct binding to microtubules, providing a new paradigm for KMN network‐mediated K‐MT attachment in mammalian oocyte meiosis.