Biology subjects

Koch, L. B.

Publications and source records attributed to Koch, L. B..

4 recordsLinked to original sources

bioRxiv2026

A Kinetochore-Associated Proteasome Pool Drives a Second Pathway of Cohesin Removal during Meiosis

Accurate chromosome segregation requires the spatiotemporally regulated removal of sister chromatid cohesion. Cohesin cleavage by the endopeptidase separase depends on destruction of its inhibitor securin by the ubiquitin-proteasome system (UPS) and on phosphorylation-mediated priming of the cohesin kleisin subunit. Whether the UPS also contributes to cohesin priming has remained unknown. Here, we show that the 26S proteasome mediates cohesin removal during meiosis II through branches of two parallel pathways. Using separation-of-function mutants targeting either the proteasome's core or regulatory particles, we identify a proteasome function that is required specifically for centromeric cohesin removal during meiosis II but dispensable for separase activation. Defects in this proteasome function causes centromeric accumulation of phosphatase anchor shugoshin (Sgo1), impaired cleavage of meiotic kleisin Rec8, and frequent failure of sister chromatid segregation. Bypassing the requirement for Rec8 priming, either through a phosphomimetic rec8 allele or by separase-independent Rec8 cleavage, restores chromosome segregation, demonstrating that the proteasome mediates cohesin removal independently of its established role in activating separase. Consistent with a direct role, proteasomes localize prominently to kinetochores during meiosis II. Together, these findings identify the proteasome as a dual-function regulator that mediates both separase activation and cohesin priming, revealing how a single proteolytic machine coordinates the two molecular pathways underlying stepwise chromosome segregation during meiosis.

genetics↗

bioRxiv2025

Specialisation of meiotic kinetochores revealed through a synthetic SAC strategy

Meiosis creates haploid gametes through two sequential M phases. While many studies have focused on meiosis I, the molecular events which drive and define meiosis II are largely unknown. Here, we report a novel cell synchronization strategy which allows for collection of yeast cells arrested at metaphase I or metaphase II, enabling better characterisation of meiosis II events. The method relies on chemically-inducible dimerization of ectopic copies of spindle assembly checkpoint (SAC) proteins Mps1 and Spc105. Using this synthetic SAC (SynSAC) approach, we found that the SAC response is weaker in metaphase I compared to metaphase II and that the PP1 binding site within Spc105 contributes to restraining the MI SAC response. Furthermore, we demonstrate the utility of the SynSAC approach by analysing the composition and phosphorylation of kinetochores from metaphase I and metaphase II. This revealed an increase in the abundance of outer kinetochore proteins in meiotic metaphase I and reduced phosphorylation on metaphase II kinetochore proteins. Overall, we present the SynSAC method as a valuable tool for analysis of both meiotic metaphases.

cell biology↗

bioRxiv2023

Rewiring of the phosphoproteome executes two meiotic divisions

The cell cycle is ordered by a controlled network of kinases and phosphatases. To generate gametes via meiosis, two distinct and sequential chromosome segregation events occur without an intervening S phase. How canonical cell cycle controls are modified for meiosis is not well understood. Here, using highly synchronous budding yeast populations, we reveal how the global proteome and phosphoproteome changes during the meiotic divisions. While protein abundance changes are limited to key cell cycle regulators, dynamic phosphorylation changes are pervasive. Our data indicate that two waves of cyclin-dependent kinase and Polo (Cdc5Polo) kinase activity drive successive meiotic divisions. These two distinct waves of phosphorylation are ensured by the meiosis-specific Spo13Meikin protein, which rewires the phosphoproteome. Spo13Meikin binds to Cdc5Polo to promote phosphorylation of a subset of substrates in meiosis I containing a newly identified motif, which we define as the Spo13Meikin -Cdc5Polo consensus phosphorylation motif. Overall, our findings reveal that a master regulator of meiosis redirects the activity of a kinase to change the phosphorylation landscape and elicit a developmental cascade.

cell biology↗

bioRxiv2021

Kinetochore-associated Mps1 regulates the strength of kinetochore-microtubule attachments via Ndc80 phosphorylation

Dividing cells detect and correct erroneous kinetochore-microtubule attachments during mitosis, thereby avoiding chromosome mis-segregation. Most studies of this process have focused on the Aurora B kinase, which phosphorylates microtubule-binding elements specifically at incorrectly attached kinetochores, promoting their release and providing another chance for proper attachments to form. However, growing evidence suggests additional mechanisms, potentially involving Mps1 kinase, that also underlie error correction. Because these mechanisms overlap in vivo, and because both Mps1 and Aurora B function in numerous other vital processes, their contributions to the correction of erroneous kinetochore attachments have been difficult to disentangle. Here we directly examine how Mps1 activity affects kinetochore-microtubule attachments using a reconstitution-based approach that allowed us to separate its effects from Aurora B activity. When endogenous Mps1 that co-purifies with isolated kinetochores is activated in vitro, it weakens their attachments to microtubules via phosphorylation of Ndc80, a major microtubule-binding element of the outer kinetochore. Mps1 phosphorylation of Ndc80 appears to contribute to error correction because phospho-deficient Ndc80 mutants exhibit genetic interactions and segregation defects when combined with mutants in an intrinsic error correction pathway. In addition, Mps1 phosphorylation of Ndc80 is stimulated on kinetochores lacking tension. These data suggest that Mps1 provides an additional mechanism for correcting erroneous kinetochore-microtubule attachments, complementing the well-known activity of Aurora B.

cell biology↗