RESUMO
Female meiosis provides an opportunity for selfish genetic elements to violate Mendel's law of segregation by increasing the chance of segregating to the egg [1]. Centromeres and other repetitive sequences can drive in meiosis by cheating the segregation process [2], but the underlying mechanisms are unknown. Here, we show that centromeres with more satellite repeats house more nucleosomes that confer centromere identity, containing the histone H3 variant CENP-A, and bias their segregation to the egg relative to centromeres with fewer repeats. CENP-A nucleosomes predominantly occupy a single site within the repeating unit that becomes limiting for centromere assembly on smaller centromeres. We propose that amplified repetitive sequences act as selfish elements by promoting expansion of CENP-A chromatin and increased transmission through the female germline.
Assuntos
Proteína Centromérica A/genética , Centrômero/metabolismo , Meiose , Repetições de Microssatélites , Animais , Linhagem Celular , Proteína Centromérica A/metabolismo , Feminino , CamundongosRESUMO
The histone H3 variant CENP-A is incorporated into nucleosomes that mark centromere location. We have recently reported that CENP-A nucleosomes, compared with their H3 counterparts, confer an altered nucleosome shape. Here, using a single-molecule fluorescence resonance energy transfer (FRET) approach with recombinant human histones and centromere DNA, we found that the nucleosome shape change directed by CENP-A is dominated by lateral passing of two DNA gyres (gyre sliding). A nonhistone centromere protein, CENP-C, binds and reshapes the nucleosome, sliding the DNA gyres back to positions similar to those in canonical nucleosomes containing conventional histone H3. The model that we generated to explain the CENP-A-nucleosome transition provides an example of a shape change imposed by external binding proteins and has notable implications for understanding of the epigenetic basis of the faithful inheritance of centromere location on chromosomes.
Assuntos
Autoantígenos/metabolismo , Centrômero/química , Centrômero/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Nucleossomos/química , Nucleossomos/metabolismo , Proteína Centromérica A , Transferência Ressonante de Energia de Fluorescência , Humanos , Modelos Biológicos , Modelos MolecularesRESUMO
Inheritance of each chromosome depends upon its centromere. A histone H3 variant, centromere protein A (CENP-A), is essential for epigenetically marking centromere location. We find that CENP-A is quantitatively retained at the centromere upon which it is initially assembled. CENP-C binds to CENP-A nucleosomes and is a prime candidate to stabilize centromeric chromatin. Using purified components, we find that CENP-C reshapes the octameric histone core of CENP-A nucleosomes, rigidifies both surface and internal nucleosome structure, and modulates terminal DNA to match the loose wrap that is found on native CENP-A nucleosomes at functional human centromeres. Thus, CENP-C affects nucleosome shape and dynamics in a manner analogous to allosteric regulation of enzymes. CENP-C depletion leads to rapid removal of CENP-A from centromeres, indicating their collaboration in maintaining centromere identity.
Assuntos
Autoantígenos/metabolismo , Centrômero/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Nucleossomos/metabolismo , Autoantígenos/química , Autoantígenos/genética , Centrômero/química , Centrômero/ultraestrutura , Proteína Centromérica A , Proteínas Cromossômicas não Histona/química , Proteínas Cromossômicas não Histona/genética , DNA/química , DNA/metabolismo , Epigênese Genética , Transferência Ressonante de Energia de Fluorescência , Técnicas de Silenciamento de Genes , Humanos , Nucleossomos/química , Nucleossomos/ultraestrutura , Estrutura Secundária de ProteínaRESUMO
Structural dynamics of nucleic acid and protein is an important physical basis of their functions. These motions are often very difficult to synchronize and too fast to be clearly resolved with the currently available single molecule methods. Here we demonstrate a novel hybrid single molecule approach combining stochastic data analysis with fluorescence correlation that enables investigations of sub-ms unsynchronized structural dynamics of macromolecules. Based on the method, we report the first direct evidence of spontaneous DNA motions at the nucleosome termini. The nucleosome, comprising DNA and a histone core, is the fundamental packing unit of eukaryotic genes that must be accessed during various genome transactions. Spontaneous DNA opening at the nucleosome termini has long been hypothesized to enable gene access in the nucleosome, but has yet to be directly observed. Our approach reveals that DNA termini in the nucleosome open and close repeatedly at 0.1-1 ms(-1). The kinetics depends on salt concentration and DNA-histone interactions but not much on DNA sequence, suggesting that this dynamics is universal and imposes the kinetic limit to gene access. These results clearly demonstrate that our method provides an efficient and robust means to investigate unsynchronized structural changes of DNA at a sub-ms time resolution.
Assuntos
DNA/química , Transferência Ressonante de Energia de Fluorescência/métodos , Nucleossomos/química , Corantes Fluorescentes , Funções Verossimilhança , Movimento (Física) , FótonsRESUMO
The centromere is the locus that directs chromosomal inheritance at cell division. While centromeres in diverse eukaryotes are commonly found at sites of repetitive DNA, their location is epigenetically specified. The histone H3 variant CENP-A is the prime candidate for epigenetically marking the centromere, and recent work has uncovered several additional proteins that play key roles in centromere assembly and maintenance. We describe advances in the identification and characterization of proteins that form the centromere, and focus on recent findings that have advanced our understanding of the assembly of functional centromeric chromatin. This article is part of a Special Issue entitled: Histone chaperones and chromatin assembly.
Assuntos
Centrômero/metabolismo , Cromatina/metabolismo , Animais , Ciclo Celular/fisiologia , Cromatina/química , Montagem e Desmontagem da Cromatina/fisiologia , Proteínas Cromossômicas não Histona/fisiologia , Epigênese Genética/fisiologia , Humanos , Transdução de Sinais/fisiologiaRESUMO
The centromere is the locus that directs chromosomal inheritance at cell division. While centromeres in diverse eukaryotes are commonly found at sites of repetitive DNA, their location is epigenetically specified. The histone H3 variant CENP-A is the prime candidate for epigenetically marking the centromere, and recent work has uncovered several additional proteins that play key roles in centromere assembly and maintenance. We describe advances in the identification and characterization of proteins that form the centromere, and focus on recent findings that have advanced our understanding of the assembly of functional centromeric chromatin. This article is part of a Special Issue entitled: Histone chaperones and chromatin assembly.
