1. DNA Organization Lec 2

2. Aims The aims of this lecture is to investigate how cells organize their DNA within the cell nucleus, how is the huge amount of genomic DNA packaged to fit within the cell nucleus, and still keeping specific sequences accessible for transcription?. We will discuss genome organization, satellites, repetitive DNAetc We will discuss the structure of the nucleosome and higher levels of chromatin organization and packaging.

3. Interactions with proteins All the functions of DNA depend on interactions with proteins. These protein interactions can be non-specific, or the protein can bind specifically to a single DNA sequence. Enzymes can also bind to DNA for example the polymerases that copy the DNA sequence in transcription and DNA replication.

4. DNA-binding proteins Within chromosomes, DNA is held in complexes with structural proteins. These proteins organize the DNA into a compact structure called chromatin. In eukaryotes this structure involves DNA binding to a complex of small basic proteins called histones, while in prokaryotes multiple types of proteins are involved. The histones form a disk-shaped complex called a nucleosome. These non-specific interactions are formed through basic residues in the histones making ionic bonds to the acidic sugar-phosphate backbone of the DNA.

6. Chromatin Chromatin is the complex combination of DNA and protein that makes up chromosomes. It is found inside the nuclei of eukaryotic cells. The major components of chromatin are DNA and histone proteins. The functions of chromatin are to package DNA into a smaller volume to fit in the cell.

7. Chromatin is the substance which becomes visible chromosomes during cell division. Its basic unit is nucleosome, composed of 146 bp DNA and eight histone proteins. The structure of chromatin is dynamically changing, at least in part, depending on the need of transcription. In the metaphase of cell division, the chromatin is condensed into the visible chromosome. At other times, the chromatin is less condensed, with some regions in a "Beads-On-a-String" conformation.

9. Histones are the proteins closely associated with DNA molecules. They are responsible for the structure of chromatin and play important roles in the regulation of gene expression. Five types of histones have been identified: H1 (or H5), H2A, H2B, H3 and H4. H1 and its homologous protein H5 are involved in higher- order structures of chromatin. The other four types of histones associate with DNA to form nucleosomes.

10. Histones (H1, H2A, H2B, H3, H4, and H5) organized into two super classes as follows: Core histones – H2A, H2B, H3 and H4 and linker histones – H1 and H5. Histones contain a high proportion of basic amino acids (arginine and lysine) that facilitate binding to the negatively charged DNA molecule.

11. Two of each of the core histones (H2A, H2B, H3 and H4) assemble to form one nucleosome core particle by wrapping 146 base pairs of DNA around the protein spool in 1.65 left- handed super-helical turn. The linker histone H1 binds the nucleosome and the entry and exit sites of the DNA, thus locking the DNA into place and allowing the formation of higher order structure.

14. each nucleosome is associated with an H1 (or H5) to form a solenoid structure. H1 and H5 are called linker histones.

16. Chromosomes A chromosome is an organized structure of DNA and protein that is found in cells. It is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences. Chromosomes also contain DNA-bound proteins, which serve to package the DNA and control its functions.

17. Chromosomes in prokaryotes The prokaryotes – bacteria and archaea – typically have a single circular chromosome, but many variations do exist. Most bacteria have a single circular chromosome that can range in size from only 160,000 base pairs in the endosymbiotic bacterium Candidatus Carsonella ruddii, to 12,200,000 base pairs in the soil-dwelling bacterium Sorangium cellulosum. Spirochaetes of the genus Borrelia are a notable exception to this arrangement, with bacteria such as Borrelia burgdorferi, ontaining a single linear chromosome.

18. Repetitive DNA Sequences A stretch of DNA sequence often repeats several times in the total DNA of a cell. For example, the following DNA sequence is just a small part of telomere located at the ends of each human chromosome:

19. An entire telomere, about 15 kb, is constituted by thousands of the repeated sequence "GGGTTA".

20. DNA sequences are divided into three classes: Highly repetitive: About 10-15% of mammalian DNA fragments reassociate very rapidly. This class includes tandem repeats. Moderately repetitive: Roughly 25-40% of mammalian DNA fragments reassociate at an intermediate rate. This class includes interspersed repeats (also known as mobile elements or transposable elements). Single copy (or very low copy number): This class accounts for 50-60% of mammalian DNA.

21. Tandem repeats are an array of consecutive repeats. They include three subclasses: satellites, minisatellites and microsatellites. The name "satellites" comes from their optical spectra.

22. Satellites The size of a satellite DNA ranges from 100 kb to over 1 Mb. In humans, a well known example is the alphoid DNA located at the centromere of all chromosomes. Its repeat unit is 171 bp and the repetitive region accounts for 3-5% of the DNA in each chromosome. Other satellites have a shorter repeat unit. Most satellites in humans or in other organisms are located at the centromere.

23. Minisatellites The size of a minisatellite ranges from 1 kb to 20 kb. One type of minisatellites is called variable number of tandem repeats (VNTR). Its repeat unit ranges from 9 bp to 80 bp. They are located in non-coding regions. The number of repeats for a given minisatellite may differ between individuals. This feature is the basis of DNA fingerprinting. Another type of minisatellites is the telomere. In a human germ cell, the size of a telomere is about 15 kb. In an aging somatic cell, the telomere is shorter. The telomere contains tandemly repeated sequence GGGTTA.

24. Microsatellites Microsatellites are also known as short tandem repeats (STR), because a repeat unit consists of only 1 to 6 bp and the whole repetitive region spans less than 150 bp. Similar to minisatellites, the number of repeats for a given microsatellite may differ between individuals. Therefore, microsatellites can also be used for DNA fingerprinting. In addition, both microsatellite and minisatellite patterns can provide information about paternity.

25. Interspersed Repeats Interspersed repeats are repeated DNA sequences located at dispersed regions in a genome. They are also known as mobile elements or transposable elements. A stretch of DNA sequence may be copied to a different location through DNA recombination. After many generations, such sequence (the repeat unit) could spread over various regions. Mobile elements are found in all kinds of organisms. In mammals, the most common mobile elements are LINEs and SINEs.

26. Thanks for your attention