Final Report Summary - STRUCTMITO (Structural Basis for the Molecular Mechanisms Involving the Ska Complex in Establishing Stable Kinetochore - Microtubule Attachments.)
We had previously shown that the Ska complex (Ska1-Ska2-Ska3) forms a ‘W’-shaped dimer made of triple helical bundles with microtubule-binding domains flanking at the ends of each bundle and that this architecture is critical for stabilizing kinetochore-microtubule attachments (Jeyaprakash et al., 2012, Mol Cell). We further demonstrated that unlike the Ndc80 complex, the Ska complex binds surface-exposed regions of tubulin-monomers that are not affected by microtubule-polymerisation/depolymerisation, thus allowing it to bind dynamic MTs, a key requirement for chromosome segregation (Abad et al., 2014, Nat Commun). Recently, we identified the Ska complex subunit Ska3 as a direct binder of MTs whose activity is required for timely mitotic progression (Abad et al., 2016, Sci Rep).
Error-correction mediated by the CPC depends on its timely inner centromere localisation of during prometaphase mediated by Histone-H3 (HH3) and hSgo1. We had previously demonstrated that the N-terminal tails of both HH3 and hSgo1 are capable of binding Survivin (Jeyaprakash et al., 2011, Structure). Our recent work unprecedentedly showed that CPC-HH3 and CPC-hSgo1 interactions are mutually exclusive in-vitro and might happen in a temporally and/or spatially restricted manner (Gupta et al., MS in prep).
Correct kinetochore assembly and function rely on the maintenance of the centromeric chromatin underlying the kinetochore. Mis18 is a central player regulating this process. We showed that Mis18 centromere localisation and function requires its ‘Yippee-like’ domain mediated oligomerisation (Subramanian et al., 2016, EMBO Rep).
To explore RNA-mediated centromere regulation, we structurally characterised CENP-32, originally identified as a kinetochore-associated RNA-methyltransferase. While CENP-32 kinetochore role is unclear, it is required to integrate centrosomes into mitotic spindle. Ongoing CENP-32 substrate identification and functional characterization will provide novel-insights into RNA-mediated regulation of chromosome segregation.
Over all our work during the funding period provided key mechanistic insights into the regulation of kinetochore function essential for chromosome segregation.