Chapter 12 DNA Organization in Chromosomes PDF

Chapter 12 DNA Organization in Chromosomes Genetic information in viruses, bacteria, mitochondria, and chloroplasts is typically contained in short, circular DNA molecules with minimal associated proteins. Eukaryotic cells have large amounts of DNA organized into nucleosomes, exist...

Chapter 12 DNA Organization in Chromosomes Genetic information in viruses, bacteria, mitochondria, and chloroplasts is typically contained in short, circular DNA molecules with minimal associated proteins. Eukaryotic cells have large amounts of DNA organized into nucleosomes, existing as uncoiled chromatin fibers or more condensed structures during most of the cell cycle. During eukaryotic cell division, the uncoiled chromatin fibers of interphase coil up and condense into chromosomes. Bacterial genomes primarily consist of unique DNA sequences coding for proteins, while eukaryotic genomes contain both unique and repetitive DNA sequences. Eukaryotic genomes are predominantly composed of noncoding DNA sequences. Learning Objectives Bacterial chromosomes are typically circular and smaller, while viral chromosomes can be either DNA or RNA and vary in size and structure. This reflects the simpler complexity of viruses compared to bacteria. To determine if viral DNA fits inside a viral particle, compare the length and diameter of the DNA with the size of the viral head. The number of base pairs in a DNA molecule can be calculated from its length. Research findings have led to the development of the chromatin structure model. Chromatin organization follows a hierarchy, affecting replication and transcription through histone/DNA associations. The number of nucleosomes required to pack DNA into a specific nanometer fiber can be predicted. Euchromatin is less condensed and transcriptionally active, while heterochromatin is more condensed and transcriptionally inactive. Epigenetic marks like DNA methylation and histone acetylation affect gene transcription and can be predicted based on the current state of transcription. C-banding and G-banding are techniques for staining chromosomes, with G-banding used for human chromosome nomenclature. The eukaryotic genome is more complex than bacterial or viral genomes. Only a small percentage of the eukaryotic genome codes for proteins. 12.1 Viral and Bacterial Chromosomes Are Relatively Simple DNA Molecules Viral and bacterial chromosomes are simpler than eukaryotic chromosomes, typically consisting of a single nucleic acid molecule. Bacterial chromosomes lack associated proteins and contain less genetic information, simplifying genetic analysis. Viral chromosomes can be DNA or RNA, single or double-stranded, and can exist as circular or linear molecules. Examples include the single-stranded DNA of the ϕX174 bacteriophage and the double-stranded DNA of the polyoma virus, both forming closed circles. The bacteriophage lambda (λ) has a linear double-stranded DNA that forms a ring upon infecting a host cell. T-even bacteriophages have linear double-stranded DNA that does not form circles inside the host. Circularity is not essential for viral replication.  Viruses, bacteria, and eukaryotic cells share the ability to package long DNA molecules into small volumes. Bacterial chromosomes are composed of double-stranded DNA molecules. These chromosomes are compacted into a structure known as the nucleoid. Escherichia coli, a well-studied bacterium, has a circular chromosome approximately 1.2 mm in length. The chromosome can be visualized under an electron microscope when the cell is gently lysed. 12.2 Supercoiling Facilitates Compaction of the DNA of Viral and Bacterial Chromosomes Supercoiled DNA is a characteristic of closed-circular DNA molecules, discovered through studies on the polyoma virus. In 1963, high-speed centrifugation of polyoma DNA revealed three components of different densities and compactness. The least dense component was identified as linear DNA, while the denser components were circular DNA molecules. Closed-circular DNA molecules are more compact and sediment faster during centrifugation than linear DNA molecules of the same molecular weight. Jerome Vinograd proposed that the denser circular DNA molecules were covalently closed helices that are slightly underwound, leading to supercoiling. Supercoiling occurs to maintain normal base pairing and results in tighter packing and increased density. Underwound DNA molecules form negative supercoils to achieve a more energetically favorable conformation, resulting in a more compact and stable structure. In closed-circular DNA, such as in bacteria and their phages, the DNA helix is often slightly underwound. E. coli exhibits a higher number of supercoils, aiding in chromosome condensation in the nucleoid region. Topoisomerases are enzymes that convert one topoisomer to another by cutting and resealing DNA strands. They are classified as type I (cleaving one strand) or type II (cleaving both strands). In E. coli, topoisomerase I reduces negative supercoils, while topoisomerase II introduces them. In eukaryotes, supercoils occur when DNA regions are embedded in protein lattices, creating anchored ends that maintain supercoils introduced by topoisomerases. DNA replication and transcription in both prokaryotes and eukaryotes generate supercoils, which are managed by topoisomerases. Topoisomerases also play roles in separating sister chromatids' DNA following replication. 12.4 DNA Is Organized into Chromatin in Eukaryotes Eukaryotic chromosomes are highly condensed structures visible during mitosis and decondense into chromatin during interphase. Chromatin is dispersed throughout the nucleus during interphase and condenses back into chromosomes as cells reenter mitosis. Chromatin condensation represents a length contraction of approximately 10,000 times for each chromatin fiber. Eukaryotic DNA organization is more complex than in viruses or bacteria due to the greater amount of DNA and the presence of numerous associated proteins. E. coli chromosome DNA is about 1200 μm long, whereas human chromosome DNA ranges from 19,000 to 73,000 μm in length. The total DNA in a single human nucleus can extend almost 2 meters, contained within a nucleus measuring about 5 to 10 μm in diameter. Chromatin Structure and Nucleosomes Viral and bacterial genetic material consists of DNA or RNA with minimal protein. Eukaryotic chromatin has significant protein associated with chromosomal DNA throughout the cell cycle. Chromatin-associated proteins are either positively charged histones or less positively charged nonhistone proteins. Histones play a crucial structural role in chromatin. Histones are rich in positively charged amino acids, lysine and arginine, allowing them to bond electrostatically with the negatively charged phosphate groups of nucleotides. There are five main types of histones. Categories and Properties of Histone Proteins Chromatin structure involves extensive coiling and folding of DNA and protein within the cell nucleus. Histones are crucial for chromatin structure, as their removal disrupts the regular diffraction pattern of chromatin. Chromatin consists of repeating structural units, as evidenced by regularly spaced diffraction rings. Electron microscopy shows chromatin fibers as linear arrays of spherical particles, resembling beads on a string, known as nucleosomes. Nucleosomes, initially called v-bodies, are regularly spaced along the chromatin strand and support the model of repeating units in chromatin. Each nucleosome unit consists of one of each tetramer, forming an octamer, associated with approximately 200 base pairs of DNA. Extended nuclease digestion removes some of the 200 base pairs, resulting in a nucleosome core particle with 147 base pairs of DNA. The DNA lost during digestion is linker DNA, which connects nucleosomes and is associated with histone H1. A detailed model of the nucleosome, based on X-ray and neutron-scattering analyses, shows a 147-bp DNA segment coiling around a histone octamer in a left-handed superhelix, completing about 1.7 turns per nucleosome. Each nucleosome is ellipsoidal, measuring about 11 nm at its longest point. The formation of nucleosomes reduces the DNA helix to about one-third of its original length by winding around histones. Chromatin fibers in the nucleus are rarely in an extended form; instead, they are packed into a 30-nm fiber, also known as a solenoid, which is dependent on histone H1. The 30-nm fiber consists of nucleosomes coiled and stacked, providing a sixfold increase in DNA compaction and is characteristic of uncoiled chromatin in interphase. During the transition to mitotic chromosomes, the 30-nm fibers fold into looped domains, further condensing into a 300-nm structure. These coiled chromatin fibers compact into chromosome arms, forming chromatids, with a typical diameter of 700 nm, though this can vary among organisms. A pair of sister chromatids in a chromosome measures about 1400 nm. Human cells store genetic material in a nucleus about 5 to 10 µm in diameter, with the haploid genome containing over 3 billion base pairs of DNA across 23 chromosomes. The diploid cell contains twice the DNA amount, totaling almost 2 meters in length when fully extended. DNA in a human nucleus is complexed with approximately 25 × 10^6 nucleosomes. The transition from an extended DNA helix to a mitotic chromosome requires a packing ratio of about 500 to 1, though current models account for a ratio of about 50 to 1, indicating further condensation is necessary. Chromatin Remodeling Histone proteins are crucial for packaging DNA into nucleosomes, which form chromatin. DNA in chromatin is often inaccessible to DNA-binding proteins necessary for replication and transcription. Chromatin remodeling is the process by which chromatin changes structure to allow protein-DNA interactions. Chromatin must relax to expose DNA for replication and gene expression and then compact again during inactivity. In 1997, Timothy Richmond's team improved X-ray diffraction resolution of nucleosome crystals to 2.8 Å, revealing detailed DNA-histone interactions. The nucleosome structure consists of 147 base pairs of DNA wrapped around four pairs of histone proteins, repeating throughout the chromatin fiber. o Histone tails are targets for chemical modifications that impact chromatin remodeling and gene expression regulation. o Acetylation, mediated by histone acetyltransferase (HAT), neutralizes the positive charge of lysine residues, leading to chromatin fiber remodeling and gene activation. o High acetylation levels correlate with active genes, while low levels are seen in inactive regions, such as the underacetylated histone H4 in Barr bodies. o Methylation and phosphorylation are other key histone modifications, involving enzymes methyltransferases and kinases, respectively. o Methylation of arginine and lysine residues is linked to gene activity, while phosphorylation of serine and histidine introduces negative charges, correlating with chromatin changes during the cell cycle. o These modifications are reversible, controlled by specific enzymes. o Methylation of cytosine in DNA (forming 5-methyl cytosine) usually negatively correlates with gene activity, especially in CpG islands. o Chromatin remodeling is crucial for genetic processes and gene expression regulation, with further details discussed in later chapters on gene expression and epigenetics. Heterochromatin o Eukaryotic chromosomes consist of a continuous double-helical DNA fiber but are not structurally uniform throughout. o Euchromatin and heterochromatin are terms used to describe uncoiled and condensed parts of chromosomes, respectively. o Heterochromatin is genetically inactive, either lacking genes or containing repressed genes, and replicates later in the S phase than euchromatin. o Heterochromatin includes regions like telomeres, which maintain chromosome integrity, and centromeres, which are involved in chromosome movement during cell division. o The presence of heterochromatin is unique to eukaryotes, with some chromosomes, such as the mammalian Y chromosome, being largely heterochromatic. o The inactivated X chromosome in mammalian females forms a condensed heterochromatic Barr body. o In some species, entire haploid sets of chromosomes can be heterochromatic, as seen in mealy bugs. o Translocation of heterochromatic areas can render previously active euchromatic regions genetically inert, demonstrating the position effect, where the gene's location relative to other genetic material affects its expression. 12.5 Chromosome Banding Differentiates Regions along the Mitotic Chromosome o Before 1970, mitotic chromosomes could only be distinguished by size and centromere position, making it difficult to differentiate between similar chromosomes. o Chromosome-banding techniques were developed to allow differential staining along the longitudinal axis of mitotic chromosomes, creating distinct banding patterns. o Mary-Lou Pardue and Joe Gall developed one of the first chromosome-banding techniques, known as C- banding. o C-banding involves heat denaturing chromosome preparations and treating them with Giemsa stain, which results in only the centromeric regions of mitotic chromosomes taking up the stain. o This technique highlights specific areas of the chromosome composed of heterochromatin. o In mice, which have telocentric chromosomes, the stain localizes at the end of each chromosome. o Chromosome-banding techniques were developed to produce differential staining patterns along chromosomes. o G-banding is a key technique that involves digesting mitotic chromosomes with trypsin and then staining with Giemsa. o The resulting G-bands reflect the heterogeneity and complexity of the chromosome's structure. o In 1971, a standardized nomenclature for human chromosome-banding patterns was established using G- banding. o The nomenclature applies to the X chromosome, with specific regions designated on the p and q arms. o The left side of the chromosome diagram shows the organizational levels of banding, while the right side shows the specific region designations. o Banding patterns on chromosomes are unique and allow for the distinction between chromosomes of identical size and centromere placement. o These patterns are crucial in cytogenetic analysis, especially in humans, enabling the identification of specific chromosomes. o The precision of banding patterns allows for the differentiation of homologous chromosomes and the identification of translocated chromosome segments.