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1. Dynamic cell cycle–dependent phosphorylation modulates CENP-L–CENP-N centromere recruitment

   https://doi.org/10.1091/mbc.E22-06-0239  

   Navarro, Alexandra P.; Cheeseman, Iain M. (September 2022, Molecular Biology of the Cell)

   Bloom, Kerry (Ed.)

   The kinetochore is a macromolecular structure that is needed to ensure proper chromosome segregation during each cellular division. The kinetochore is assembled upon a platform of the 16-subunit constitutive centromere-associated network (CCAN), which is present at centromeres throughout the cell cycle. The nature and regulation of CCAN assembly, interactions, and dynamics needed to facilitate changing centromere properties and requirements remain to be fully elucidated. The CENP-LN complex is a CCAN component that displays unique cell cycle–dependent localization behavior, peaking in the S phase. Here, we demonstrate that phosphorylation of CENP-L and CENP-N controls CENP-LN complex formation and localization in a cell cycle–dependent manner. Mimicking constitutive phosphorylation of either CENP-L or CENP-N or simultaneously preventing phosphorylation of both proteins prevents CENP-LN localization and disrupts chromosome segregation. Our work suggests that cycles of phosphorylation and dephosphorylation are critical for CENP-LN complex recruitment and dynamics at kinetochores to enable cell cycle–dependent CCAN reorganization. 

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2. The Sequence Dependent Nanoscale Structure of CENP-A Nucleosomes

   https://doi.org/10.3390/ijms231911385  

   Stormberg, Tommy; Lyubchenko, Yuri L. (October 2022, International Journal of Molecular Sciences)

   CENP-A is a histone variant found in high abundance at the centromere in humans. At the centromere, this histone variant replaces the histone H3 found throughout the bulk chromatin. Additionally, the centromere comprises tandem repeats of α-satellite DNA, which CENP-A nucleosomes assemble upon. However, the effect of the DNA sequence on the nucleosome assembly and centromere formation remains poorly understood. Here, we investigated the structure of nucleosomes assembled with the CENP-A variant using Atomic Force Microscopy. We assembled both CENP-A nucleosomes and H3 nucleosomes on a DNA substrate containing an α-satellite motif and characterized their positioning and wrapping efficiency. We also studied CENP-A nucleosomes on the 601-positioning motif and non-specific DNA to compare their relative positioning and stability. CENP-A nucleosomes assembled on α-satellite DNA did not show any positional preference along the substrate, which is similar to both H3 nucleosomes and CENP-A nucleosomes on non-specific DNA. The range of nucleosome wrapping efficiency was narrower on α-satellite DNA compared with non-specific DNA, suggesting a more stable complex. These findings indicate that DNA sequence and histone composition may be two of many factors required for accurate centromere assembly. 

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3. Ccp1 Homodimer Mediates Chromatin Integrity by Antagonizing CENP-A Loading

   https://doi.org/10.1016/j.molcel.2016.08.022  

   Dong, Qianhua; Yin, Feng-Xiang; Gao, Feng; Shen, Yuan; Zhang, Faben; Li, Yang; He, Haijin; Gonzalez, Marlyn; Yang, Jinpu; Zhang, Shu; et al (October 2016, Molecular Cell)
4. CENP-N promotes the compaction of centromeric chromatin

   https://doi.org/10.1038/s41594-022-00758-y  

   Zhou, Keda; Gebala, Magdalena; Woods, Dustin; Sundararajan, Kousik; Edwards, Garrett; Krzizike, Dan; Wereszczynski, Jeff; Straight, Aaron F.; Luger, Karolin (April 2022, Nature Structural & Molecular Biology)

   Abstract The histone variant CENP-A is the epigenetic determinant for the centromere, where it is interspersed with canonical H3 to form a specialized chromatin structure that nucleates the kinetochore. How nucleosomes at the centromere arrange into higher order structures is unknown. Here we demonstrate that the human CENP-A-interacting protein CENP-N promotes the stacking of CENP-A-containing mononucleosomes and nucleosomal arrays through a previously undefined interaction between the α6 helix of CENP-N with the DNA of a neighboring nucleosome. We describe the cryo-EM structures and biophysical characterization of such CENP-N-mediated nucleosome stacks and nucleosomal arrays and demonstrate that this interaction is responsible for the formation of densely packed chromatin at the centromere in the cell. Our results provide first evidence that CENP-A, together with CENP-N, promotes specific chromatin higher order structure at the centromere. 

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5. Heterochromatin and RNAi regulate centromeres by protecting CENP-A from ubiquitin-mediated degradation

   https://doi.org/10.1371/journal.pgen.1007572  

   Yang, Jinpu; Sun, Siyu; Zhang, Shu; Gonzalez, Marlyn; Dong, Qianhua; Chi, Zhongxuan; Chen, Yu-hang; Li, Fei; Malik, Harmit S. (August 2018, PLOS Genetics)