Genetics, DNA Packaging

Article Author:
Hessah Al Aboud
Article Editor:
Nora Al Aboud
Updated:
11/8/2018 8:35:41 PM
PubMed Link:
Genetics, DNA Packaging

Introduction

Eukaryotic genomes consist of a nuclear genome that is packaged into chromatin in chromosomes. It also consists of mitochondria and chloroplast genomes. A eukaryotic chromosome is one long strand of DNA from tip-to-tip. Eukaryotic chromosomes contain many origins of replication approximately 100,000 base pairs apart. They have at the middle of the chromosome a centromere, which is attached at kinetochore, a complex of proteins that aid in ensuring the 2 sister chromatids go to opposite poles during mitosis and meiosis. Eukaryotic chromosomes also have telomeric regions on the ends of the chromosomes which prevent shortening during DNA replication.[1][2]

Development

All DNA is wrapped around structures called nucleosomes. Nucleosomes are composed of double-stranded DNA wrapped around an octamer of 8 histone proteins H2A, H2B, H3, and H4. It is a dimer of 2 each of these proteins.

The DNA wraps around this ball of proteins, and there are a linker region and another ball of proteins wrapping the DNA around nucleosomes. This creates a fiber that is 10 to 11 nanometers in diameter. A nucleosome consists of the 8 histone proteins and approximately 146 or 147 base pairs of DNA. The linker region in between the 2 nucleosomes can vary from 20 to 100 base pairs, depending on species and cell type, and the region of the chromosome that is being either transcribed or not transcribed. Each nucleosome contains 2 molecules of H2A, H2B, H3, and H4. The DNA is wrapped around this. There are long tails sticking out from the histone protein. These proteins tails can be modified by acetylation, methylation, and phosphorylation, and these modifications will affect gene regulation.[3][4]

Cellular

Roger Kornberg, a prominent DNA and chromatin biologist, proposed the model of nucleosome structure in 1974. The model was based on his biochemical experiments, x-ray diffraction studies, and electron microscopy images. Markus Noll's experiment, however, gave an easily visually interpretable result to understand how DNA wraps around the nucleosomes. Therefore, his experiment began with nuclei not extracted DNA, but actual nuclei. One can remove nuclei from cells and work them into individual intestines.

A nucleus has DNA that is associated with proteins. These orange balls in the image below represent a single nucleosome, and the blue line is DNA. Noll treated these nuclei with 2 concentrations of DNase. His idea was that the nucleosomes protect the DNA, but a high concentration of DNase will digest the regions in between the nucleosomes. DNase would cut every linker region of this chromatin, but at very low concentrations of DNase, only some of these regions would be cut. This would leave different lengths of DNA at the end of the digestion. NExt, Noll treated the nuclei with the 3 different concentrations of DNase. He treated the nuclei with detergent and phenol to extract the DNA. He ran the DNA out on an agarose gel. Agarose gel is a matrix. If an electric field is applied across the agarose gel, the DNA will migrate toward a positive pole because DNA is negatively charged. With this technique, the smaller fragments will migrate quicker, and the larger fragments will migrate slower; therefore, the DNA can separate based on its size.[5][6]

Biochemical

After the DNA has been run on the gel, Noll stained it with ethidium bromide. This intercalates into the double strands of DNA. It fluoresces under UV light, at which point a  picture can be taken. These images allow Noll's data interpretation. When the DNA is cut by DNase, one can end up with a length of DNA that is one nucleosome worth of DNA.

At high concentrations of DNase, all the chromosomal DNA digested into fragments that are approximately 200 base pairs long. Noll interpreted the data was that every single linker region was cut by DNase and that the length of DNA that was wrapped around a single nucleosome was 200 base pairs. At low concentrations, however not all the linker DNA was cut, resulting in DNA fragments that were 1 nucleosome long or 2 nucleosomes long, 400 base pairs, or 3, 4, 5, and so on nucleosomes long. In this way, Markus Noll determined that the DNA in the nucleus wrapped in chromatin was approximately 200 base pairs long around each nucleosome[7][8]

Molecular

In addition to the core nucleosomes, nucleosomal proteins, and a histone octamer there are additional proteins that bind to DNA in the nucleus. There is another histone called H1 that binds to the DNA just next to the nucleosome. There are also other DNA binding proteins known as non-histone proteins that play a role in organization and compaction of the chromosome, and this is a very large group of heterogeneous proteins. Many different proteins are called non-histone proteins. Without each one, the chromatin is a 10-nanometer fiber; an addition of histone H1 creates a 30-nanometer fiber. The 30-nanometer fiber is a coil of chrome with the 10-nanometer fiber chromatin. There are 2 models that likely exist in the cell. The Solenoid model is very compact, and the nucleosome is wound tightly around (regular, spiral configuration containing 6 nucleosomes per turn). The Zigzag model is a little bit looser form of chromatin (irregular configuration where nucleosomes have little face-to-face contact), but they are both 30-nanometer fibers.[9][10]

Function

There are many proteins in the nucleus. Nuclei are not an empty bag of fluid. They contain something called the nuclear matrix which consists of a lot of different types of proteins. The nuclear lamina is just under the inner membrane of the nucleus, and here there are scaffolding proteins and matrix attachment proteins. Eukaryotic DNA is organized into loops. The loops can be quite variable in length from 25 to 200 base pairs long, and there are matrix attachment regions (MARs) or scaffold attachment regions (SARs) where the DNA is bound to the matrix or scaffold of the chromosome, and the MARs are attached to the nuclear matrix creating these radial loops.[11] 

Mechanism

Chromosomes have different regions called heterochromatin regions and euchromatin regions. Heterochromatin regions are tightly compacted there at the telomeres and centromeres, these regions of the chromosome are always heterochromatin, and they are always tightly packaged where the DNA is very tightly coiled around proteins.[12]

Pathophysiology

Euchromatin exists in the other sections of the arms of the chromosomes where genes that are expressed are found. The euchromatin (30-nm fibers) are anchored in radial loops in these regions of chromosomes. Greater compaction of the radial loops constitutes the heterochromatin. In eukaryotic nuclei, the DNA from individual chromosomes is not intertwined with other chromosomes but remains in specific regions of the nucleus.[13][14]

Clinical Significance

The DNA double helix, which is approximately 2 nm in diameter. Adding the histone core created an 11-nm fiber; adding histone H1 creates a 30-nm fiber, and this 30-nm fiber is anchored to form radio loops to the nuclear matrix. Further compaction of radial loops ends up with a metaphase chromosome with a final diameter of a metaphase chromosome of about 1400 nm. Euchromatin is loosely packaged; heterochromatin is more tightly packaged. The most tightly packaged is a metaphase chromosome. If this metaphase chromosome is treated with high salt to remove the proteins, the DNA splays out from the scaffold. The 2 multiprotein complexes help to form and organize metaphase chromosomes and cohesin, which keeps the 2 sister chromatid aligned.[15][16][17][18][1]



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      Contributed by Nora Al Aboud, PhD (Genetics)