Rae Brown
Ph.D. Student in Biochemistry, admitted Autumn 2020
All Publications
-
Phosphorylation of Xenopus M18BP1 governs centromeric localization and CENP-A nucleosome assembly.
EMBO reports
2026
Abstract
Eukaryotic chromosome segregation requires attachment of chromosomes to microtubules through the kinetochore so that chromosomes can align and move in mitosis. Kinetochores assemble on the centromere, which is epigenetically defined by the histone H3 variant CENtromere Protein A (CENP-A). During DNA replication, CENP-A is equally divided between replicated chromatids, and new CENP-A nucleosomes are re-assembled during the subsequent G1 phase. How cells regulate the cell cycle timing of CENP-A assembly is a central question in the epigenetic maintenance of centromeres. CENP-A nucleosome assembly requires the Mis18 complex (Mis18α, Mis18β, and M18BP1), whose localization to centromeres occurs between metaphase and G1. Here, we define a new regulatory mechanism that works through phosphorylation of Xenopus laevis M18BP1 between metaphase and interphase. Phosphorylation disrupts binding of M18BP1 to CENP-A nucleosomes in metaphase, and when relieved, enables M18BP1 binding to CENP-A nucleosomes in interphase. We show that this phosphorylation-dependent mechanism regulates CENP-A nucleosome assembly. We propose that the phospho-regulated binding of M18BP1 to CENP-A nucleosomes restricts new CENP-A assembly to interphase.
View details for DOI 10.1038/s44319-026-00714-7
View details for PubMedID 41680291
View details for PubMedCentralID 6702948
-
Regulation of X. laevis M18BP1 centromeric localization and CENP-A assembly.
bioRxiv : the preprint server for biology
2025
Abstract
Eukaryotic chromosome segregation requires attachment of chromosomes to microtubules of the mitotic spindle through the kinetochore so that chromosomes can align and move in mitosis. Kinetochores are assembled on the centromere which is a unique chromatin domain that is epigenetically defined by the histone H3 variant CENtromere Protein A (CENP-A). During DNA replication CENP-A is equally divided between replicated chromatids and new CENP-A nucleosomes are re-assembled during the subsequent G1 phase of the cell cycle. How cells regulate the strict cell cycle timing of CENP-A assembly is a central question in the epigenetic maintenance of centromeres and kinetochores. One essential assembly factor for CENP-A nucleosomes is the Mis18 complex (Mis18α, Mis18β, and M18BP1) which is regulated in its localization to centromeres between metaphase and G1 when CENP-A assembly occurs. Here, we define a new regulatory mechanism that works through cell cycle dependent phosphorylation of Xenopus laevis M18BP1 between metaphase and interphase. This phosphoregulatory switch disrupts binding of M18BP1 to CENP-A nucleosomes in metaphase, and when relieved enables M18BP1 binding to CENP-A nucleosomes in interphase. We show that this phosphorylation dependent switching mechanism regulates CENP-A nucleosome assembly. We propose that the phospho-regulated binding of M18BP1 to CENP-A nucleosomes is an important control mechanism that restricts the timing of new CENP-A assembly.
View details for DOI 10.1101/2025.07.15.664882
View details for PubMedID 40791504
View details for PubMedCentralID PMC12338635
-
Repression of CENP-A assembly in metaphase requires HJURP phosphorylation and inhibition by M18BP1.
The Journal of cell biology
2023; 222 (6)
Abstract
Centromeres are the foundation for mitotic kinetochore assembly and thus are essential for chromosome segregation. Centromeres are epigenetically defined by nucleosomes containing the histone H3 variant CENP-A. CENP-A nucleosome assembly is uncoupled from replication and occurs in G1, but how cells control this timing is incompletely understood. The formation of CENP-A nucleosomes in vertebrates requires CENP-C and the Mis18 complex which recruit the CENP-A chaperone HJURP to centromeres. Using a cell-free system for centromere assembly in X. laevis egg extracts, we discover two activities that inhibit CENP-A assembly in metaphase. HJURP phosphorylation prevents the interaction between HJURP and CENP-C in metaphase, blocking the delivery of soluble CENP-A to centromeres. Non-phosphorylatable mutants of HJURP constitutively bind CENP-C in metaphase but are not sufficient for new CENP-A assembly. We find that the M18BP1.S subunit of the Mis18 complex also binds to CENP-C to competitively inhibit HJURP's access to centromeres. Removal of these two inhibitory activities causes CENP-A assembly in metaphase.
View details for DOI 10.1083/jcb.202110124
View details for PubMedID 37141119
-
DiMeLo-seq: a long-read, single-molecule method for mapping protein-DNA interactions genome wide.
Nature methods
2022
Abstract
Studies of genome regulation routinely use high-throughput DNA sequencing approaches to determine where specific proteins interact with DNA, and they rely on DNA amplification and short-read sequencing, limiting their quantitative application in complex genomic regions. To address these limitations, we developed directed methylation with long-read sequencing (DiMeLo-seq), which uses antibody-tethered enzymes to methylate DNA near a target protein's binding sites in situ. These exogenous methylation marks are then detected simultaneously with endogenous CpG methylation on unamplified DNA using long-read, single-molecule sequencing technologies. We optimized and benchmarked DiMeLo-seq by mapping chromatin-binding proteins and histone modifications across the human genome. Furthermore, we identified where centromere protein A localizes within highly repetitive regions that were unmappable with short sequencing reads, and we estimated the density of centromere protein A molecules along single chromatin fibers. DiMeLo-seq is a versatile method that provides multimodal, genome-wide information for investigating protein-DNA interactions.
View details for DOI 10.1038/s41592-022-01475-6
View details for PubMedID 35396487