Stepwise unfolding supports a subunit model for vertebrate kinetochores

Abstract

During cell division, interactions between microtubules and chromosomes are mediated by the kinetochore, a proteinaceous structure located at the primary constriction of chromosomes. In addition to the centromere histone centromere protein A (CENP-A), 15 other members of the constitutive centromere associated network (CCAN) participate in the formation of a chromatin-associated scaffold that supports kinetochore structure. We performed a targeted screen analyzing unfolded centrochromatin from CENP-depleted chromosomes. Our results revealed that CENP-C and CENP-S are critical for the stable folding of mitotic kinetochore chromatin. Multipeak fitting algorithms revealed the presence of an organized pattern of centrochromatin packing consistent with arrangement of CENP-A-containing nucleosomes into up to five chromatin "subunits"-each containing roughly 20-30 nucleosomes. These subunits could be either layers of a boustrophedon or small loops of centromeric chromatin.centromere | kinetochore | CENP-A | centrochromatin | boustrophedon

Distinct Roles of the Chromosomal Passenger Complex in the Detection of and Response to Errors in Kinetochore-Microtubule Attachment

“…Proteins at the inner kinetochore not only provide a platform for outer kinetochore assembly, but have also been implicated in the response to tension (Ribeiro et al, 2010; Suzuki et al, 2014; Vargiu et al, 2017). Therefore, we hypothesized that the localization of the CPC to centromeres may play a role in the localization of inner kinetochore components (Figure 1B).…”

“…However, we also show that BubR1 recruitment to unattached kinetochores does not depend on a fully formed inner kinetochore (Figure S5C). A likely explanation for this separation of function is that a fully formed inner kinetochore is required to prevent “hyper-stretching” within kinetochores by microtubule pulling forces (a defect that is not reflected in inter-kinetochore distances) (Ribeiro et al, 2010; Suzuki et al, 2014; Vargiu et al, 2017; Suzuki et al, 2011). We propose that kinetochores built in the presence of non-centromeric CPC are “hyper-stretched” and thus there is no differentiation between high and low tension states.…”

Centromere inheritance through the germline

“…Indeed, a single point mutation of the nucleosome interaction surface of CENP-C retains binding to CENP-A but eliminates structural stability and hinders its ability to retain CENP-A nucleosomes at the centromere (Guo et al 2017). CENP-C and CENP-S have also been shown to be important for resisting unfolding of centromeric chromatin in low ionic strength solutions (Vargiu et al 2017). In addition, flies with impaired CENP-C function have reduced CENP-A at centromeres in spermatids, indicating an extrinsic mechanism for maintaining CENP-A during early meiosis in males (Kwenda et al 2016).…”

Structural mechanisms of centromeric nucleosome recognition by the kinetochore protein CENP-N

“…Thus, the valency of CENP-C/CENP-N/CENP-L interactions could facilitate clustering of sparse and nonadjacent CENP-A nucleosomes (Fig. 4C) (9, 26), which might help to establish the folding of centromeric chromatin (27, 28) and/or the integrity of the kinetochore (29, 30). …”