分类: 生物学 >> 生物物理学 >> 细胞生物学 提交时间: 2016-05-12
摘要: The H3 histone variant CENP-A is an epigenetic marker critical for the centromere identity and function. However, the precise regulation of the spatiotemporal deposition and propagation of CENP-A at centromeres during the cell cycle is still poorly understood. Here, we show that CENP-A is phosphorylated at Ser68 during early mitosis by Cdk1. Our results demonstrate that phosphorylation of Ser68 eliminates the binding of CENP-A to the assembly factor HJURP, thus preventing the premature loading of CENP-A to the centromere prior to mitotic exit. Because Cdk1 activity is at its minimum at the mitotic exit, the ratio of Cdk1/PP1 alpha activity changes in favor of Ser68 dephosphorylation, thus making CENP-A available for centromeric deposition by HJURP. Thus, we reveal that dynamic phosphorylation of CENP-A Ser68 orchestrates the spatiotemporal assembly of newly synthesized CENP-A at active centromeres during the cell cycle.
分类: 生物学 >> 生物物理学 >> 细胞生物学 提交时间: 2016-05-12
摘要: Specific recognition of centromere-specific histone variant CENP-A-containing chromatin by CENP-N is an essential process in the assembly of the kinetochore complex at centromeres prior to mammalian cell division. However, the mechanisms of CENP-N recruitment to centromeres/kinetochores remain unknown. Here, we show that a CENP-A-specific RG loop (Arg80/Gly81) plays an essential and dual regulatory role in this process. The RG loop assists the formation of a compact "ladder-like" structure of CENP-A chromatin, concealing the loop and thus impairing its role in recruiting CENP-N. Upon G1/S-phase transition, however, centromeric chromatin switches from the compact to an open state, enabling the now exposed RG loop to recruit CENP-N prior to cell division. Our results provide the first insights into the mechanisms by which the recruitment of CENP-N is regulated by the structural transitions between compaction and relaxation of centromeric chromatin during the cell cycle.
分类: 生物学 >> 生物物理学 >> 生物物理、生物化学与分子生物学 提交时间: 2016-05-11
摘要: Eukaryotic genomes are organized hierarchically into chromatin structures by histones. Despite extensive research for over 30 years, not only the fundamental structure of the 30-nm chromatin fiber is being debated, but the actual existence of such fiber remains hotly contested. In this review, we focus on the most recent progress in elucidating the structure of the 30-nm fiber upon in vitro reconstitution, and its possible organization inside the nucleus. In addition, we discuss the roles of linker histone H1 as well as the importance of specific nucleosome-nucleosome interactions in the formation of the 30-nm fiber. Finally, we discuss the involvement of structural variations and epigenetic mechanisms available for the regulation of this chromatin form. (C) 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
分类: 生物学 >> 生物物理学 提交时间: 2016-05-11
摘要: DNA methylation is an important epigenetic modification that is essential for various developmental processes through regulating gene expression, genomic imprinting, and epigenetic inheritance(1-5). Mammalian genomic DNA methylation is established during embryogenesis by de novo DNA methyltransferases, DNMT3A and DNMT3B(6-8), and the methylation patterns vary with developmental stages and cell types(9-12). DNAmethyltransferase 3-like protein (DNMT3L) is a catalytically inactive paralogue of DNMT3 enzymes, which stimulates the enzymatic activity of Dnmt3a(13). Recent studies have established a connection between DNA methylation and histone modifications, and revealed a histone-guided mechanism for the establishment of DNA methylation(14). The ATRX-DNMT3-DNMT3L (ADD) domain of Dnmt3a recognizes unmethylated histone H3 (H3K4me0)(15-17). The histone H3 tail stimulates the enzymatic activity of Dnmt3a in vitro(17,18), whereas the molecular mechanism remains elusive. Here we show that DNMT3A exists in an autoinhibitory form and that the histone H3 tail stimulates its activity in a DNMT3L-independent manner. We determine the crystal structures of DNMT3A-DNMT3L (autoinhibitory form) and DNMT3A-DNMT3L-H3 (active form) complexes at 3.82 and 2.90 angstrom resolution, respectively. Structural and biochemical analyses indicate that the ADD domain of DNMT3A interacts with and inhibits enzymatic activity of the catalyticdomain (CD) through blocking its DNA-binding affinity. HistoneH3(but not H3K4me3) disrupts ADD-CD interaction, induces a large movement of the ADD domain, and thus releases the autoinhibition of DNMT3A. The finding adds another layer of regulation of DNA methylation to ensure that the enzyme is mainly activated at proper targeting loci when unmethylated H3K4 is present, and strongly supports a negative correlation between H3K4me3 and DNA methylation across the mammalian genome(9,10,19,20). Our study provides a new insight into an unexpected autoinhibition and histone H3-induced activation of the de novo DNA methyltransferase after its initial genomic positioning.
分类: 生物学 >> 生物物理学 提交时间: 2016-05-11
摘要: Although there have been many investigations into how trinucleotide repeats affect nucleosome formation and local chromatin structure, the nucleosome positioning of GAA triplet-repeats in the human genome has remained elusive. In this work, the nucleosome occupancy around GAA triplet-repeats across the human genome was computed statistically. The results showed a nucleosome-depleted region in the vicinity of GM triplet-repeats in activated and resting CD4(+) T cells. Furthermore, the A-tract was frequently adjacent to the upstream region of GM triplet-repeats and could enhance the depletion surrounding GAA triplet-repeats. In vitro chromatin reconstitution assays with GM-containing plasmids also demonstrated that the inserted GAA triplet-repeats destabilized the ability of recombinant plasmids to assemble nucleosomes. Our results suggested that GM triplet-repeats have lower affinity to histones and can change local nucleosome positioning. These findings may be helpful for understanding the mechanism of Friedreich's ataxia, which is associated with GM triplet-repeats at the chromatin level. (C) 2015 Elsevier Inc All rights reserved.