Acetylation of histone H4 lysine 5 and 12 is required for CENP-A deposition into centromeres

Centromeres are specified epigenetically through the deposition of the centromere-specific histone H3 variant CENP-A. However, how additional epigenetic features are involved in centromere specification is unknown. Here, we find that histone H4 Lys5 and Lys12 acetylation (H4K5ac and H4K12ac) primarily occur within the pre-nucleosomal CENP-A-H4HJURP (CENP-A chaperone) complex, before centromere deposition. We show that H4K5ac and H4K12ac are mediated by the RbAp46/ 48-Hat1 complex and that RbAp48-deficient DT40 cells fail to recruit HJURP to centromeres and do not incorporate new CENP-A at centromeres. However, C-terminally-truncated HJURP, that does not bind CENP-A, does localize to centromeres in RbAp48-deficient cells. Acetylation-dead H4 mutations cause mis-localization of the CENP-A-H4 complex to non-centromeric chromatin. Crucially, CENP-A with acetylation-mimetic H4 was assembled specifically into centromeres even in RbAp48-deficient DT40 cells. We conclude that H4K5ac and H4K12ac, mediated by RbAp46/ 48, facilitates efficient CENP-A deposition into centromeres

Chromosomal Instability Affects the Tumorigenicity of Glioblastoma Tumor-Initiating Cells.

UNLABELLED: Tumors are dynamic organs that evolve during disease progression with genetic, epigenetic, and environmental differences among tumor cells serving as the foundation for selection and evolution in tumors. Tumor-initiating cells (TIC) that are responsible for tumorigenesis are a source of functional cellular heterogeneity, whereas chromosomal instability (CIN) is a source of karyotypic genetic diversity. However, the extent that CIN contributes to TIC genetic diversity and its relationship to TIC function remains unclear. Here, we demonstrate that glioblastoma TICs display CIN with lagging chromosomes at anaphase and extensive nonclonal chromosome copy-number variations. Elevating the basal chromosome missegregation rate in TICs decreases both proliferation and the stem-like phenotype of TICs in vitro Consequently, tumor formation is abolished in an orthotopic mouse model. These results demonstrate that TICs generate genetic heterogeneity within tumors, but that TIC function is impaired if the rate of genetic change is elevated above a tolerable threshold. SIGNIFICANCE: Genetic heterogeneity among TICs may produce advantageous karyotypes that lead to therapy resistance and relapse; however, we found that TICs have an upper tolerable limit for CIN. Thus, increasing the chromosome missegregation rate offers a new therapeutic strategy to eliminate TICs from tumors. Cancer Discov; 6(5); 532-45. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 461

Small changes in phospho-occupancy at the kinetochore-microtubule interface drive mitotic fidelity

Kinetochore protein phosphorylation promotes the correction of erroneous microtubule attachments to ensure faithful chromosome segregation during cell division. Determining how phosphorylation executes error correction requires an understanding of whether kinetochore substrates are completely (i.e., all-or-none) or only fractionally phosphorylated. Using quantitative mass spectrometry (MS), we measured phospho-occupancy on the conserved kinetochore protein Hec1 (NDC80) that directly binds microtubules. None of the positions measured exceeded ∼50% phospho-occupancy, and the cumulative phospho-occupancy changed by only ∼20% in response to changes in microtubule attachment status. The narrow dynamic range of phospho-occupancy is maintained, in part, by the ongoing phosphatase activity. Further, both Cdk1–Cyclin B1 and Aurora kinases phosphorylate Hec1 to enhance error correction in response to different types of microtubule attachment errors. The low inherent phospho-occupancy promotes microtubule attachment to kinetochores while the high sensitivity of kinetochore–microtubule attachments to small changes in phospho-occupancy drives error correction and ensures high mitotic fidelity