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1、染色体与 DNA,Figure CO:X chromosome in blue,染色体,2.1.1 染色体概述,染色体(chromosome)是细胞核中载有遗传信息的物质，在显微镜下呈丝状或棒状，由核酸和蛋白质组成，在细胞发生有丝分裂时期容易被碱性染料着色，因此而得名。同一物种内每条染色体所带DNA的量是一定的，但不同染色体或不同种中间变化很大。,原核细胞染色体外裹着稀疏的蛋白质，其中一部分与DNA的折叠有关，另一些则参与DNA复制、重组及转录过程。真核细胞的染色体中，DNA与蛋白质完全融合在一起，其蛋白质与相应DNA的质量比约为1:1.,chromosome A discrete unit
2、of the genome carrying many genes.Each consists of a very long molecule of duplex DNA and an approximately equal mass of proteins.It is visible as a morphological entity only during cell division.,A thin section shows the bacterial nucleoid as a compact mass in the center of the cell,原核细胞染色体(类核Nucle
3、oid),The nucleoid spills out of a lysed E.coli cell in the form of loops of a fiber,The bacterial genome consists of a large number of loops of duplex DNA,FIGURE 08:An unrestrained supercoil in the DNA path creates tension,but no tension is transmitted along DNA when a supercoil is restrained by pro
4、tein binding,The sister chromatids of a mitotic pair each consist of a fiber(30 nm in diameter)compactly folded into the chromosome,真核细胞染色体,类核；染色质；包装比例,nucleoid The structure in a prokaryotic cell that contains the genome.The DNA is bound to proteins and is not enclosed by a membrane.chromatin The s
5、tate of nuclear DNA and its associated proteins during the interphase(between mitoses)of the eukaryotic cell cycle.packing ratio The ratio of the length of DNA to the unit length of the fiber containing it.,Specific Sequences Attach DNA to an Interphase Matrix（分裂间期基质）,DNA is attached to the nuclear
6、matrix at specific sequences called MARs or SARs.The MARs are A-T-rich but do not have any specific consensus sequence.chromosome scaffold（支架）A proteinaceous structure in the shape of a sister chromatid pair,generated when chromosomes are depleted of histones.,Chromatin Is Divided into Euchromatin a
7、nd Heterochromatin,Regions of heterochromatin remain densely packed throughout interphase.chromocenter An aggregate of heterochromatin from different chromosomes.,Regions of compact heterochromatin are clustered near the nucleolus and nuclear membrane,常染色质和异染色质Euchromatin and Heterochromatin,DNA Is
8、Organized in Arrays of Nucleosomes（核小体）,The core DNA is the length of 146 bp that is found on the core particles produced by prolonged digestion with MNase.Linker DNA is the region of 8 to 114 bp that is susceptible to early cleavage by nucleases.,FIGURE 05:Micrococcal nuclease initially cleaves bet
9、ween nucleosomes,The Nucleosome Is the Subunit of All Chromatin,A nucleosome contains 200 bp of DNA and two copies of each core histone(H2A,H2B,H3,and H4).DNA is wrapped around the outside surface of the protein octamer（八聚体）.The histone octamer has a structure of an H32-H42 tetramer associated with
10、two H2A-H2B dimers.,FIGURE 06:The nucleosome consists of approximately equal masses of DNA and histones(including H1),The Nucleosome Is the Subunit of All Chromatin,The Nucleosome Is the Subunit of All Chromatin,The nucleosome is a cylinder with DNA organized into 1 2/3 turns around the surface,The
11、Nucleosome Is the Subunit of All Chromatin,Each histone is extensively interdigitated with its partner.All core histones have the structural motif of the histone fold.N-and C-terminal histone tails extend out of the nucleosome.H1 is associated with linker DNA and may lie at the point where DNA enter
12、s or exits the nucleosome.,Photos courtesy of E.N.Moudrianakis,Johns Hopkins University.,FIGURE 10ab:The crystal structure of the histone core octamer is represented in a space-filling model,The Nucleosome Is the Subunit of All Chromatin,The histone fold consists of two short a-helices flanking a lo
13、nger a-helix,Histone pairs(H3+H4 and H2A+H2B)interact to form histone dimers,Structures from Protein Data Bank 1HIO.G.Arents,et al.,Proc.Natl.Acad.Sci.USA 88(1991):10145-10152.,The Nucleosome Is the Subunit of All Chromatin,The histone fold domains of the histones are located in the core of the nucl
14、eosome,Nucleosomes Are Covalently Modified,Histones are modified by methylation,acetylation,phosphorylation,and other modifications.Combinations of specific histone modifications define the function of local regions of chromatin;this is known as the histone code.,Histone tails have many sites of mod
15、ification,Nucleosomes Are Covalently Modified,Photo courtesy of Sean D.Taverna,Johns Hopkins University School of Medicine,and Haitao Li,Memorial Sloan-Kettering Cancer Center.Additional information at S.D.Taverna,et al.,Nat.Struct.Mol.Biol.14(2007):1025-1040.,Numerous protein motifs recognize methy
16、lated lysines,Nucleosomes Are Covalently Modified,Acetylation during replication occurs on specific sites on histones before they are incorporated into nucleosomes,Nucleosomes Are Covalently Modified,Acetylation associated with gene activation occurs by directly modifying specific sites on histones
17、that are already incorporated into nucleosomes,DNA Structure Varies on the Nucleosomal Surface,DNA is wrapped 1.67 times around the histone octamer.DNA on the nucleosome shows regions of smooth curvature and regions of abrupt kinks.The structure of the DNA is altered so that it has an increased numb
18、er of base pairs/turn in the middle,but a decreased number at the ends.,Structures from Protein Data Bank:1P34.U.M.Muthurajan,et al.,EMBO J.23(2004):260-271.,The Path of Nucleosomes in the Chromatin Fiber,10 nm chromatin fibers consist of a string of nucleosomes.30 nm fibers have six nucleosomes/tur
19、n,which are organized into a two-start helix.Histone H1,histone tails,and increased ionic strength all promote the formation of the 30 nm fiber.,The Path of Nucleosomes in the Chromatin Fiber,The 10 nm fiber is a continuous string of nucleosomes,The 30 nm fiber is a two start helix consisting of two
20、 rows of nucleosomes coiled into a solenoid,Levels of chromatin packaging,Replication of Chromatin Requires Assembly of Nucleosomes,Histone octamers are not conserved during replication,but H2A-H2B dimers and H32-H42 tetramers are conserved.There are different pathways for the assembly of nucleosome
21、s during replication and independently of replication.,Replication of Chromatin Requires Assembly of Nucleosomes,Nucleosome assembly in vivo,Nucleosomes Are Displaced and Reassembled During Transcription,Most transcribed genes retain a nucleosomal structure,though the organization of the chromatin c
22、hanges during transcription.Some heavily transcribed genes appear to be exceptional cases that are devoid of nucleosomes.RNA polymerase displaces histone octamers during transcription in vitro,but octamers reassociate with DNA as soon as the polymerase has passed.,Nucleosomes Are Displaced and Reass
23、embled During Transcription,An experiment to test the effect of transcription on nucleosomes shows that the histone octamer is displaced from DNA and rebinds at a new position,Nucleosomes Are Displaced and Reassembled During Transcription,Nucleosomes are reorganized when transcription passes through
24、 a gene.Additional factors are required both for RNA polymerase to displace octamers during transcription and for the histones to reassemble into nucleosomes after transcription.,染色体带型G（Giemsa)-banding generates a characteristic lateral series of bands in each member of the chromosome set,The human
25、X chromosome can be divided into distinct regions by its banding pattern,每条带约10000Kb,A magnified view of bands 87A and 87C shows their hybridization in situ with labeled RNA extracted from heat-shocked cells,Chromosomes are pulled to the poles via microtubules that attach at the centromeres,有丝分裂中期,有
26、丝分裂后期,有丝分裂末期,着丝点,粘结蛋白,微管,A model of the overall structure of a regional centromere,FIGURE 26:A large protein complex assembles at the CDE sequences and connects the chromosome to microtubules,A typical telomere has a simple repeating structure with a G-T-rich strand that extends beyond the C-A-rich
27、strand,端粒有简单的重复序列A typical telomere has a simple repeating structure with a G-T-rich strand that extends beyond the C-A-rich strand,The crystal structure of a short repeating sequence from the human telomere forms three stacked G quartets,端粒封堵线性染色体的末端,A loop forms at the end of chromosomal DNA,染色体末端
28、形成环状结构,A loop forms at the end of chromosomal DNA,端粒的功能保护染色体末端：端粒延长协助减数分裂中同源染色体配对,The meiotic telomere cluster is visualized by telomere FISH,端粒对生存是必需的,1.在分裂细胞中存在端粒酶活性。2.端粒酶突变后，每次分裂，染色体变短。3.端粒消失后，细胞不能分裂。,Mutation in telomerase causes telomeres to shorten in each cell division,Telomere length is main
29、tained at 350 bp in wild-type yeast,Reproduced from T.M.Nakamura,et al.,Science 277(1997):955-959 http:/www.sciencemag.org.Reprinted with permission from AAAS.Photo courtesy of Thomas R.Cech,Howard Hughes Medical Institute.,真核细胞染色体的组成,1.蛋白质,包括组蛋白和非组蛋白。组蛋白是染色体的结构蛋白，它与DNA组成核小体。,组成：DNA，组蛋白，非组蛋白，RNA,富含赖
30、氨酸和精氨酸,组蛋白的特性1.进化上的极端保守性 特别是H3、H4，牛、猪、大鼠的H4完全相同2.无组织特异性 3.肽链上氨基酸分布的不对称性 N端碱性氨基酸 C端疏水氨基酸4.组蛋白的修饰作用 甲基化、乙酰化、磷酸化、多聚ADP糖基化等 5.富含赖氨酸的组蛋白H5 只存在于鸟类、两栖类等含有细胞核的红细胞中非组蛋白包括酶类和细胞分裂有关的蛋白（收缩蛋白、骨架蛋白、核孔复合物蛋白、肌动蛋白、肌球蛋白、微管蛋白、原肌蛋白）高速泳动蛋白（high mobility group protein,HMG蛋白）能与DNA结合，也能与H1作用，可能与超螺旋结构有关。DNA结合蛋白 可能是一些与DNA复制或转录有关的酶或调解物质。A24非组蛋白 C端与H2A相同，功能不详,3.染色质和核小体,染色质DNA的Tm值比自由DNA高。染色质DNA的复制和转录活性比自由DNA低。DNA酶I对染色质DNA的消化慢于纯DNA。,核小体是一种串珠状结构，由核心颗粒和连结线DNA两部分组成，可描述如下：每个核小体单位包括约200bp的DNA、一个组蛋白核心和一个H1；由H2A、H2B、H3、H4各两分子形成八聚体，构成核心颗粒；DNA分子以左手螺旋缠绕在核心颗粒表面，每圈80bp，共1.75圈，约146bp，两端被H1锁合；相邻核心颗粒之间为一段60bp的连接线DNA。,
