Duro, E. | Search bioRxiv

bioRxiv2020

The proteomic landscape of centromeric chromatin reveals an essential role for the Ctf19CCAN complex in meiotic kinetochore assembly

Kinetochores direct chromosome segregation in mitosis and meiosis. Faithful gamete formation through meiosis requires that kinetochores take on new functions that impact homolog pairing, recombination and the orientation of kinetochore attachment to microtubules in meiosis I. Using an unbiased proteomics pipeline, we determined the composition of centromeric chromatin and kinetochores at distinct cell-cycle stages, revealing extensive reorganisation of kinetochores during meiosis. The data uncover a network of meiotic chromosome axis and recombination proteins that replace the microtubule-binding outer kinetochore sub-complexes during meiotic prophase. We show that this kinetochore remodelling in meiosis requires the Ctf19cCCAN inner kinetochore complex. Through functional analyses, we identify a Ctf19cCCAN-dependent kinetochore assembly pathway that is dispensable for mitotic growth, but becomes critical upon meiotic entry. Therefore, extensive kinetochore remodelling and a distinct assembly pathway direct the specialization of meiotic kinetochores for successful gametogenesis.

cell biology↗

bioRxiv2018

The Molecular Basis of Monopolin Recruitment to the Kinetochore

In budding yeast meiosis I, the kinetochores of each sister chromatid pair are fused by the monopolin complex to mediate their monoorientation on the meiosis I spindle, enabling the biorientation and segregation of homologs. Monopolin forms a V-shaped complex with binding sites for the kinetochore protein Dsn1 at the apices of the V, suggesting that monopolin forms a physical bridge between the two sister kinetochores. Here, we reveal the molecular basis of the monopolin-kinetochore interaction and identify the key interfaces required for monopolin function at the kinetochore. The disordered N-terminus of budding-yeast Dsn1 unexpectedly possesses two binding motifs for the monopolin subunit Csm1, encompassing the previously-identified \"Box 1\" and \"Box 2-3\" regions of Dsn1. Strikingly, Dsn1 Box 1 and Box 2-3 bind the same conserved hydrophobic cavity on the monopolin complex subunit Csm1, suggesting that they are mutually exclusive for Csm1 binding, yet both regions are critical for monopolin function in Saccharomyces cerevisiae meiosis I. We find that Dsn1 Box 1 is an ancestral monopolin-binding motif that is conserved throughout fungi, including in the fission yeast Schizosaccharomyces pombe. In contrast, Box 2-3 is found only in species with sequence-defined point centromeres (S. cerevisiae and its close relatives), suggesting that this region contributes specifically to sister kinetochore crosslinking in meiosis I. Finally, we propose that phosphorylation of two conserved serine residues in Box 3 may stabilize monopolin at the kinetochore, providing a potential mechanism for enforcing specific sister kinetochore crosslinking in meiosis I.

cell biology↗