DNA Sequence-Specific Binding of CENP-B Enhances the Fidelity of Human Centromere Function

Abstract

SUMMARY Human centromeres are specified by a stably inherited epigenetic mark that maintains centromere position and function through a two-step mechanism relying on self-templating centromeric chromatin assembled with the histone H3 variant CENP-A, followed by CENP-A-dependent nucleation of kinetochore assembly. Nevertheless, natural human centromeres are positioned within specific megabase chromosomal regions containing α-satellite DNA repeats, which contain binding sites for the DNA sequence specific binding protein CENP-B. We now demonstrate that CENP-B directly binds both CENP-A’s amino-terminal tail and CENP-C, a key nucleator of kinetochore assembly. DNA sequence-dependent binding of CENP-B within α-satellite repeats is required to stabilize optimal centromeric levels of CENP-C. Chromosomes bearing centromeres without bound CENP-B, including the human Y chromosome, are shown to missegregate in cells at rates several fold higher than chromosomes with CENP-B containing centromeres. These data demonstrate a DNA sequence-specific enhancement by CENP-B of the fidelity of epigenetically defined human centromere function.

Unexpected conformational variations of the human centromeric chromatin complex

“…This suggests that maintenance of strong and dense CENP-B boxes increases the efficiency of CENP-A/B/C binding to α-satellite centromeres. Our evidence that CENP-B boxes within homogeneous functional α-satellite arrays have evolved to stabilize the resident CENP-A/B/C particles provides support for the proposal that CENP-B contributes to segregation fidelity by stabilizing CENP-C (Fachinetti et al 2015).…”

Alteration/Deficiency in Activation 3 (ADA3) Protein, a Cell Cycle Regulator, Associates with the Centromere through CENP-B and Regulates Chromosome Segregation

“…Whereas studies of human artificial chromosome formation have revealed an essential role of CENP-B in de novo centromerization (12,13), the less severe mitotic defects in Cenpb knock-out mice (39 -41) and lack of CENP-B boxes in Y chromosome and neocentromeres (42,43) have led to the idea that CENP-B might be dispensable in centromere function. However, the two recent ne plus ultra studies (16,17) by Cleveland and colleagues have provided several lines of evidence that uncover the role of CENP-B in centromere function. They observed an increased rate of chromosome mis-segregation in Cenpb null MEFs and CENP-B-devoid Y and neocentromeres, a new concept according to which CENP-B ensures the highest fidelity of chromosome segregation.…”

“…protein is important in ensuring faithful chromosome segregation during mitosis and thus assuring the highest fidelity of centromere function (16,17) because deletion of CENP-B causes significant elevation in chromosome mis-segregation (17). Because ADA3 knockdown/deletion in cells causes significant reduction in CENP-B recruitment onto centromeres, we speculated that deletion of Ada3 might cause chromosome mis-segregation in cells as seen upon depletion of CENP-B.…”

“…Yeast CENP-B homolog acts as a sitespecific nucleation factor for the formation of centromeric heterochromatin by heterochromatin-specific modifications of histone tails (14). The centromere function mainly entails CENP-A, -B, and -C, in which CENP-B plays a crucial role by recruiting CENP-A and stabilizing CENP-C at centromeres (15)(16)(17).…”

Genomic variation within alpha satellite DNA influences centromere location on human chromosomes with metastable epialleles

“…Historically, CENPB was not thought to play a functional role in centromeric chromatin, since it is present at both active and inactive alpha satellite arrays (Earnshaw et al 1989;Sullivan and Schwartz 1995). However, de novo centromere assembly of HACs depends on CENPB-box-containing alpha satellite DNA (Ohzeki et al 2002;Okada et al 2007), and CENPB is thought to position CENPA nucleosomes and stabilize CENPA and CENPC within centromeric chromatin (Yoda et al 1998;Okada et al 2007;Hasson et al 2013;Fachinetti et al 2015). CENPB binding sites (i.e., the number of CENPB boxes) within an array might determine how well an array can recruit centromere proteins and achieve the three-dimensional structure required for kinetochore assembly and centromere function.…”