Ensemble-Level Organization of Human Kinetochores and Evidence for Distinct Tension and Attachment Sensors

Summary

Kinetochores are multi-protein machines that form dynamic attachments to microtubules and control chromosome segregation. High fidelity is ensured because kinetochores can monitor attachment status and tension, using this information to activate checkpoints and error-correction mechanisms. To explore how kinetochores achieve this, we used two- and three-color subpixel fluorescence localization to define how proteins from six major complexes (CCAN, MIS12, NDC80, KNL1, RZZ, and SKA) and the checkpoint proteins Bub1, Mad1, and Mad2 are organized in the human kinetochore. This reveals how the outer kinetochore has a high nematic order and is largely invariant to the loss of attachment or tension, except for two mechanical sensors. First, Knl1 unravels to relay tension, and second, NDC80 undergoes jackknifing and loss of nematic order under microtubule detachment, with only the latter wired up to the checkpoint signaling system. This provides insight into how kinetochores integrate mechanical signals to promote error-free chromosome segregation.

Graphical Abstract

Highlights

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  3D mapping of kinetochore architecture in human RPE1 cells

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  Outer kinetochore (NDC80, Mad1, and RZZ) has high nematic order

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  NDC80 jackknives and KNL1 unravels upon loss of attachment and tension, respectively

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  Recruitment of Mad2 is only coupled to the occupancy sensor (NDC80)

Abstract

Roscioli et al. use subpixel imaging and computational methods to determine the ensemble-level 3D organization of the human kinetochore. They show how kinetochores undergo distinct rearrangements in response to the loss of attachment and tension.