Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA 92093, USA.
Division of Biological Sciences & Department of Cellular & Molecular Medicine, University of California San Diego, La Jolla, CA 92093, USA.
Curr Biol. 2022 Jun 6;32(11):2563-2571.e4. doi: 10.1016/j.cub.2022.04.046. Epub 2022 May 23.
During mitosis and meiosis in the majority of eukaryotes, centromeric chromatin comprised of CENP-A nucleosomes and their reader CENP-C recruits components of the outer kinetochore to build an interface with spindle microtubules. One exception is C. elegans oocyte meiosis, where outer kinetochore proteins form cup-like structures on chromosomes independently of centromeric chromatin. Here, we show that the nucleoporin MEL-28 (ortholog of human ELYS) and CENP-C act in parallel to recruit outer kinetochore components to oocyte meiotic chromosomes. Unexpectedly, co-inhibition of MEL-28 and CENP-C resulted in chromosomes being expelled from the meiotic spindle prior to anaphase onset, a more severe phenotype than what was observed following ablation of the outer kinetochore. This observation suggested that MEL-28 and the outer kinetochore independently link chromosomes to spindle microtubules. Consistent with this, the chromosome expulsion defect was observed following co-inhibition of MEL-28 and the microtubule-coupling KNL-1/MIS-12/NDC-80 (KMN) network of the outer kinetochore. Use of engineered mutants showed that MEL-28 acts in conjunction with the microtubule-binding NDC-80 complex to keep chromosomes within the oocyte meiotic spindle and that this function likely involves the Y-complex of nucleoporins that associate with MEL-28; by contrast, the ability to dock protein phosphatase 1, shared by MEL-28 and KNL-1, is not involved. These results highlight nuclear pore-independent functions for a conserved nucleoporin and explain two unusual features of oocyte meiotic chromosome segregation in C. elegans: centromeric chromatin-independent outer kinetochore assembly, and dispensability of the outer kinetochore for constraining chromosomes in the acentrosomal meiotic spindle.
在大多数真核生物的有丝分裂和减数分裂过程中，着丝粒染色质由 CENP-A 核小体及其读取器 CENP-C 组成，它们招募外动粒的成分来构建与纺锤体微管的接口。一个例外是 C. elegans 卵母细胞减数分裂，其中外动粒蛋白在没有着丝粒染色质的情况下独立于染色体形成杯状结构。在这里，我们表明核孔蛋白 MEL-28（人类 ELYS 的同源物）和 CENP-C 平行作用，将外动粒成分募集到卵母细胞减数分裂染色体上。出乎意料的是，MEL-28 和 CENP-C 的共同抑制导致染色体在后期开始前从减数分裂纺锤体中排出，这比切除外动粒观察到的更严重的表型。这一观察结果表明，MEL-28 和外动粒独立地将染色体与纺锤体微管连接起来。与此一致，在 MEL-28 和外动粒的微管偶联 KNL-1/MIS-12/NDC-80（KMN）网络的共同抑制后观察到染色体排出缺陷。使用工程突变体表明，MEL-28 与微管结合的 NDC-80 复合物一起作用，使染色体保持在卵母细胞减数分裂纺锤体内，并且这种功能可能涉及与 MEL-28 相关的核孔蛋白 Y 复合物；相比之下，MEL-28 和 KNL-1 共有的与蛋白磷酸酶 1 对接的能力不参与其中。这些结果突出了保守核孔蛋白的核孔非依赖性功能，并解释了 C. elegans 卵母细胞减数分裂染色体分离的两个异常特征：着丝粒染色质独立的外动粒组装，以及对外动粒的 dispensability 用于约束无中心体减数分裂纺锤体中的染色体。
