Tension directly stabilizes reconstituted kinetochore-microtubule attachments

Abstract

Kinetochores are macromolecular machines that couple chromosomes to dynamic microtubule tips during cell division, thereby generating force to segregate the chromosomes1,2. Accurate segregation depends on selective stabilization of correct ‘bi-oriented’ kinetochore-microtubule attachments, which come under tension due to opposing forces exerted by microtubules3. Tension is thought to stabilize these bi-oriented attachments indirectly, by suppressing the destabilizing activity of a kinase, Aurora B4,5. However, a complete mechanistic understanding of the role of tension requires reconstitution of kinetochore-microtubule attachments for biochemical and biophysical analyses in vitro. Here we show that native kinetochore particles retaining the majority of kinetochore proteins can be purified from budding yeast and used to reconstitute dynamic microtubule attachments. Individual kinetochore particles maintain load-bearing associations with assembling and disassembling ends of single microtubules for >30 min, providing a close match to the persistent coupling seen in vivo between budding yeast kinetochores and single microtubules6. Moreover, tension increases the lifetimes of the reconstituted attachments directly, via a catch bond-like mechanism that does not require Aurora B7-10. Based on these findings, we propose that tension selectively stabilizes proper kinetochore-microtubule attachments in vivo through a combination of direct mechanical stabilization and tension-dependent phosphoregulation.

Merotelic attachments allow alignment and stabilization of chromatids in meiosis II oocytes

“…Syntelic attachments (analogous to the monopolar attachments for chromatids) are effectively replaced by amphitelic attachments (analogous to the merotelic attachments for chromatids) in somatic cells during mitosis 19,20 . This is likely a consequence of the fact that amphitelic, but not syntelic attachments, generates intra-and inter-kinetochore tension, tension that increases the number of microtubules bound to kinetochores and stabilize their interactions [21][22][23] . We propose that the merotelic attachments observed at chromatids in Sycp3 À / À MII oocytes fulfil a role similar to that of amphitelic attachments for chromosomes, generating tension across the single kinetochore of chromatids and contributing to microtubule recruitment and stabilization.…”

Dynamic catch of a Thy-1–α5β1+syndecan-4 trimolecular complex

“…Surprisingly, we find a complex mechanical regulation of adhesive bonds at the single-molecule level: tensile force prolongs the bond lifetime (that is, catch bonds). In contrast to the well-known catch bonds of bimolecular systems, such as cell adhesion receptors (integrins 28,38,39 , selectins 40,41 , glycoprotein Iba 30,42 , E-cadherin 43 and FimH 44 ), the T-cell receptor 31 and cytoskeletal linkages 45,46 , the Thy-1 catch bond is strongly correlated with a bond-stiffening phenotype, termed 'dynamic catch', which partially shifts force to the already engaged but unstretched coreceptor Syn4. Thus, the data reveal a unique, previously unreported class of receptor-ligand bonds whereby force tightens co-receptor engagement that is required for forcemediated adhesion signalling.…”

Keeping kinetochores on track

“…The study of kinetochore proteins has been difficult for a long time for two main reasons: firstly the proteins are only present in low abundance, and until very recently the purification of entire kinetochores was elusive (Akiyoshi et al, 2010). This delayed the functional study of kinetochore proteins, as only a few of them were known.…”