Kinetochores read detyrosinated tubulin to sustain microtubule attachments

ABSTRACT: Post-translational cycles of α-tubulin detyrosination/tyrosination generate microtubule diversity, with broad physiological and clinical implications. However, the mechanistic understanding of how this microtubule diversity is decoded into specific cellular functions remains largely unknown. Here we show that the core microtubule-binding site at kinetochores ‘reads’ and responds to the α-tubulin detyrosination/tyrosination code. By combining CRISPR-Cas9 gene editing of enzymatic ‘writers’ of α-tubulin detyrosination/tyrosination in human cells, with biochemical purifications, mass spectrometry and super-resolution microscopy, we identified over a hundred proteins that preferentially associate with either detyrosinated or tyrosinated microtubules during mitosis. Among these, the conserved KNL-1/MIS12 complex/NDC80 complex (KMN) network at kinetochores showed a preference for tyrosinated microtubules. In addition to established roles in error correction and polar chromosome alignment, live-cell imaging uncovered a previously overlooked link between the α-tubulin detyrosination/tyrosination code and the formation of functional kinetochore-microtubule attachments, necessary for normal metaphase chromosome oscillations and timely spindle assembly checkpoint satisfaction. In vitro reconstitution experiments revealed that α-tubulin detyrosination specifically promotes Ndc80 diffusion along the microtubule lattice. We propose that the gradual accumulation of detyrosinated α-tubulin after initial microtubule capture at kinetochores facilitates the biased diffusion of core-binding proteins along the microtubule lattice, establishing a positive feedback loop that sustains kinetochore-microtubule attachments.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Hugo Osorio  

LAB HEAD: Helder Maiato

PROVIDER: PXD047133 | Pride | 2024-10-03

REPOSITORIES: Pride