19. Summary
19.19 Summary |
All eukaryotic chromatin consists of nucleosomes. A nucleosome contains a characteristic length of DNA, usually ~200 bp, wrapped around an octamer containing two copies each of histones H2A, H2B, H3, and H4. A single H1 protein is associated with each nucleosome. Virtually all genomic DNA is organized into nucleosomes. Treatment with micrococcal nuclease shows that the DNA packaged into each nucleosome can be divided operationally into two regions. The linker region is digested rapidly by the nuclease; the core region of 146 bp is resistant to digestion. Histones H3 and H4 are the most highly conserved and an H3 PH4 tetramer accounts for the diameter of the particle. The H2A and H2B histones are organized as two H2A PH2B dimers. Octamers are assembled by the successive addition of two H2A PH2B dimers to the H32 PH42 kernel, but it is not known whether this occurs before or after association with DNA.
The path of DNA around the histone octamer creates V1.65 supercoils. The DNA "enters" and "leaves" the nucleosome in the same vicinity, and could be "sealed" by histone H1. Removal of the core histones releases V1.0 supercoils. The difference can be largely explained by a change in the helical pitch of DNA, from an average of 10.2 bp/turn in nucleosomal form to 10.5 bp/turn when free in solution. There is variation in the structure of DNA from a periodicity of 10.0 bp/turn at the nucleosome ends to 10.7 bp/turn in the center. There are kinks in the path of DNA on the nucleosome.
Nucleosomes are organized into a fiber of 30 nm diameter which has 6 nucleosomes per turn and a packing ratio of 40. Removal of H1 allows this fiber to unfold into a 10 nm fiber that consists of a linear string of nucleosomes. The 30 nm fiber probably consists of the 10 nm fiber wound into a solenoid. The 30 nm fiber is the basic constituent of both euchromatin and heterochromatin; nonhistone proteins are responsible for further organization of the fiber into chromatin or chromosome ultrastructure.
RNA polymerase displaces histone octamers during transcription. The octamers reattach to DNA after the polymerase has passed, unless transcription is very intensive (such as in rDNA) when they may be displaced completely.
Two types of changes in sensitivity to nucleases are associated with gene activity. Chromatin capable of being transcribed has a generally increased sensitivity to DNAase I, reflecting a change in structure over an extensive region that can be defined as a domain containing active or potentially active genes. Hypersensitive sites in DNA occur at discrete locations, and are identified by greatly increased sensitivity to DNAase I. They occur at sites that regulate transcription, at origins for replication, at centromeres, and other locations. A hypersensitive site consists of a sequence of ~200 bp from which nucleosomes are excluded by the presence of other proteins. A hypersensitive site forms a boundary that may cause adjacent nucleosomes to be restricted in position. Nucleosome positioning may be important in controlling access of regulatory proteins to DNA.
Formation of heterochromatin may be initiated at certain sites and then propagated for a distance that is not precisely determined. When a heterochromatic state has been established, it is inherited through subsequent cell divisions. This gives rise to a pattern of epigenetic inheritance, in which two identical sequences of DNA may be associated with different protein structures, and therefore have different abilities to be expressed. This explains the occurrence of position effect variegation in Drosophila.
Inactive chromatin at yeast telomeres and silent mating type loci appears to have a common cause, and involves the interaction of certain proteins with the N-terminal tails of histones H3 and H4. Formation of the inactive complex may be initiated by binding of one protein to a specific sequence of DNA; the other components may then polymerize in a cooperative manner along the chromosome.
Inactivation of one X chromosome in female (eutherian) mammals occurs at random. The Xic locus is necessary and sufficient to count the number of X chromosomes. The n-1 rule ensures that all but one X chromosome are inactivated. Xic contains the gene Xist, which codes for an RNA that is expressed only on the inactive X chromosome. Stabilization of Xist RNA is the mechanism by which the inactive X chromosome is distinguished.
Methylation of DNA is inherited epigenetically. Replication of DNA creates hemimethylated products, and a maintenance methylase restores the fully methylated state. Some methylation events depend on parental origin. Sperm and eggs contain specific and different patterns of methylation, with the result that paternal and maternal alleles are differently expressed in the embryo. This is responsible for imprinting, in which the nonmethylated allele inherited from one parent is essential because it is the only active allele; the allele inherited from the other parent is silent. Patterns of methylation are reset during gamete formation in every generation.
Prions are proteinaceous infectious agents that are responsible for the disease ofscrapie in sheep and for related diseases in man. The infectious agent is a variant of a normal cellular protein. The PrPSc form has an altered conformation that is self-templating: the normal PrPC form does not usually take up this conformation, but does so in the presence of PrPSc. A similar effect is responsible for inheritance of the PSI element in yeast.
