Human DNA is 2 billion nanometers in length and fits into a small nucleus which is about 2000-6000 nanometers in diameter. DNA passes through various stages to get accommodated within the nucleus. Let us explore the stages of organization of DNA.
Flow of genetic informATION
Central dogma was proposed by Francis Crick in 1958 and it proves that the information flows from DNA to RNA and then to proteins.
The first step of this dogma is the replication where the information within the DNA gets replicated. The second step is transcription wherein the information within the DNA gets copied to RNA. The third step is translation where the information within the RNA is utilized to synthesize proteins
The first step of this dogma is the replication where the information within the DNA gets replicated. The second step is transcription wherein the information within the DNA gets copied to RNA. The third step is translation where the information within the RNA is utilized to synthesize proteins
Packing ratio
Definition- The ratio obtained when the length of DNA is divided by the length into which it is packaged.
Example – There are 4.6 x 107 bp present in the shortest human chromosome. To obtain the length of DNA the number of base pairs is multiplied with .34nm (distance between the two base pairs). The length of DNA so obtained comes to 14,000 µm. The most condensed form of DNA during mitosis measures 2 µm in length. Hence the packing ratio comes to 7000 (14,000/2).
Hence packing ratio gives us an idea about the level to which DNA gets condensed. To attain this packing ratio DNA moves through several hierarchies of organization.
Organization of DNA within prokaryotes -
The prokaryotic organisms lack defined nucleus however the DNA is organized with the help of some positively charged groups into structure named as nucleoid. The DNA is negatively charged due to the phosphate groups and the repulsion due to this negative charge is counteracted by the association of DNA with positively charged polyamines such as spermine and spermidine. These positively charged groups shield the negative charges of the DNA phosphate groups.
Definition- The ratio obtained when the length of DNA is divided by the length into which it is packaged.
Example – There are 4.6 x 107 bp present in the shortest human chromosome. To obtain the length of DNA the number of base pairs is multiplied with .34nm (distance between the two base pairs). The length of DNA so obtained comes to 14,000 µm. The most condensed form of DNA during mitosis measures 2 µm in length. Hence the packing ratio comes to 7000 (14,000/2).
Hence packing ratio gives us an idea about the level to which DNA gets condensed. To attain this packing ratio DNA moves through several hierarchies of organization.
Organization of DNA within prokaryotes -
The prokaryotic organisms lack defined nucleus however the DNA is organized with the help of some positively charged groups into structure named as nucleoid. The DNA is negatively charged due to the phosphate groups and the repulsion due to this negative charge is counteracted by the association of DNA with positively charged polyamines such as spermine and spermidine. These positively charged groups shield the negative charges of the DNA phosphate groups.
Structures of polyamines which interact with DNA
Along with polyamines there are abundant small proteins which give the DNA a compact structure (ex. H-NS). The DNA finally attains a supercoiled structure which gets opened with the help of enzymes like DNA gyrase during replication.
Organization of DNA within eukaryotes
Chromosomes - In eukaryotic organisms DNA is present along with some basic proteins in the form of chromosomes within the nucleus.
Chromatin – It is a unit of analysis of a chromosome and gives a general idea of the nature of a chromosome. Chromatin is not unique to any single chromosome.
Histones:
The association of the DNA with histone proteins starts from the formation of nucleosomes.
Chromosomes - In eukaryotic organisms DNA is present along with some basic proteins in the form of chromosomes within the nucleus.
Chromatin – It is a unit of analysis of a chromosome and gives a general idea of the nature of a chromosome. Chromatin is not unique to any single chromosome.
Histones:
- They are the positively charged basic proteins.
- They are of 5 major types and contain amino acids residues like lysine and arginine.
- The five major types are H1, H2A, H2B, H3 and H4.
The association of the DNA with histone proteins starts from the formation of nucleosomes.
- Histone octamer acts like a core and consists of two copies of each of these histone proteins H2A, H2B, H3 and H4.
- The duplex DNA which is around 147bp in length wraps around the histone octamer.
- During this coiling process we can see that DNA takes one full turn and in the next round it covers ¾ of the core histone complex.
- This small basic unit is termed as nucleosome. It is the fundamental unit of chromatin. This means that chromatin is made up of repeating units of neucleosomes.
- Nucleosomes together with DNA appear like beads on a string. At this stage when the DNA is observed under electron micrograph the DNA along with histones looks like beads on a string.
- At this stage the DNA is 10nm in diameter and attains a packing ratio of about 6.
Role of Histone 1 or H1-
- It holds the DNA which is wrapped around the nucleosome in a proper position
- The H1 binds to the linker DNA (which is made up of approximately 20-60 bp) giving stability to the next level of organization.
- From the beads on a string stage the DNA coils in such a way that at least 6 nucleosomes are packed per coil.
- In this stage DNA attains a diameter of 30 nm and this level of organization is known as 30 nm fiber or solenoid fiber.
- When the chromatin is extracted from isotonic buffers it appears like a 30 nm fiber.
- At this stage it attains a packing ratio of 10.
- This stage is seen during interphase in the cell cycle.
- Later the solenoid fiber get further coiled and condensed and is organized into loops, scaffolds and domains to obtain cytologically visible threads known as chromatids.
- The looping is such that the base of the loops is attached to the same protein skeletal work.
- Some metallic ions like calcium and copper along with non histone proteins help in the looping process.
- This increases the packing ratio to about 1000 in interphase chromosomes and about 10,000 in mitotic chromosomes.
Non Histone proteins
Chromosomal proteins which are not histones are grouped under Non histone proteins. They can be acidic, basic or neutral. Their mol. wt varies from 10 KD to many million Daltons.
EM studies have demonstrated that the 30 nm fibre is highly dynamic such that it unfolds into a 10 nm fiber ("beads-on-a-string") structure when transversed by an RNA polymerase engaged in transcription. The association of DNA with histones resists endonuclease action.
Histone modifications
The primary structure of histone proteins remains the same but they vary from each other due to the chemical modifications which occur at a later stage. These modifications include:
Acetylation:
Addition of acetyl groups (CH3CO-) to lysines.
The N terminal residues of the histone proteins which make the core of the nucleosomes are lysine rich and interact with the phosphate groups of the DNA of the neucleosomes. When these lysine residues are acetylated due to enzymes like histone acetyltransferases (HATs) the interaction of lysine residues with the phosphate groups of DNA is inhibited. So high level of acetylation means the DNA will be stopped from obtaining a condensed form. This also leads to better transcription.
In a highly condensed form the DNA is less exposed to enzymes. Upon deacetylation due to enzymes like histone deacetylases (HDACs), the positively charged residues are again free to interact with DNA to attain the fully coiled structure.
Phosphorylation:
Addition of phosphate groups to serines and threonines.
This makes the chromosomes more compact and prepare them for mitosis and meiosis.
Methylation:
Addition of methyl groups to lysines and arginines.
This either stimulates or inhibits gene transcription at that region. That is some residues when methylated stimulate the transcription whereas the remaining residues inhibit the transcription.
Euchromatin and Heterochromatin
The folding of DNA is not uniform throughout. In some regions the folding is highly compact and intricate giving rise to heterochromatic regions and the others are called euchromatin regions.
Some general characters of heterochromatin
The promoter region of the gene is blocked by a nucleosome and the transcription factors cannot access this promoter region. To begin the transcription of a particular gene the nucleosome is expelled or in some other cases it slides along the DNA so that the transcription factors can bind the promoter region.
The enzyme RNA polymerase II (RNAP II) which catalyses the actual transcription travels down the DNA. During this process a set of proteins removes the neucleosome in front of the DNA to be transcribed and replaces it back after the RNA polymerase II has passed through.
Chromosomal proteins which are not histones are grouped under Non histone proteins. They can be acidic, basic or neutral. Their mol. wt varies from 10 KD to many million Daltons.
- There are about 750-2000 different kinds of Non histone proteins and examples of most abundant ones are Topoisomerases and High mobility group of proteins (HMGs).
- Functions mostly served by them are
- Helping in the structural organization of chromatin fibers
- Maintaining stability of the chromatin fibers
- Involved in gene regulation
EM studies have demonstrated that the 30 nm fibre is highly dynamic such that it unfolds into a 10 nm fiber ("beads-on-a-string") structure when transversed by an RNA polymerase engaged in transcription. The association of DNA with histones resists endonuclease action.
Histone modifications
The primary structure of histone proteins remains the same but they vary from each other due to the chemical modifications which occur at a later stage. These modifications include:
Acetylation:
Addition of acetyl groups (CH3CO-) to lysines.
The N terminal residues of the histone proteins which make the core of the nucleosomes are lysine rich and interact with the phosphate groups of the DNA of the neucleosomes. When these lysine residues are acetylated due to enzymes like histone acetyltransferases (HATs) the interaction of lysine residues with the phosphate groups of DNA is inhibited. So high level of acetylation means the DNA will be stopped from obtaining a condensed form. This also leads to better transcription.
In a highly condensed form the DNA is less exposed to enzymes. Upon deacetylation due to enzymes like histone deacetylases (HDACs), the positively charged residues are again free to interact with DNA to attain the fully coiled structure.
Phosphorylation:
Addition of phosphate groups to serines and threonines.
This makes the chromosomes more compact and prepare them for mitosis and meiosis.
Methylation:
Addition of methyl groups to lysines and arginines.
This either stimulates or inhibits gene transcription at that region. That is some residues when methylated stimulate the transcription whereas the remaining residues inhibit the transcription.
Euchromatin and Heterochromatin
The folding of DNA is not uniform throughout. In some regions the folding is highly compact and intricate giving rise to heterochromatic regions and the others are called euchromatin regions.
Some general characters of heterochromatin
- It is densely packed and is found in the regions of chromosomes where there are few or no genes such as
- Centromeres
- Telomere
- It shows a reduced level of crossing over and replicates during the later stages in the S phase of the cell cycle.
- It is enriched with transposons and other junk DNA
- The genes in the heterochromatin are inactive that is they are less transcribed.
- The transcriptionally active regions are known as euchromatin regions
- Most parts of the chromosomes which are rich in transcriptionally active genes are termed as euchromatin regions.
- They are made up of loosely packed 30nm fibers.
- These regions are separated from heterochromatin by insulators.
- The histone proteins in this region show increased acetylation.
The promoter region of the gene is blocked by a nucleosome and the transcription factors cannot access this promoter region. To begin the transcription of a particular gene the nucleosome is expelled or in some other cases it slides along the DNA so that the transcription factors can bind the promoter region.
The enzyme RNA polymerase II (RNAP II) which catalyses the actual transcription travels down the DNA. During this process a set of proteins removes the neucleosome in front of the DNA to be transcribed and replaces it back after the RNA polymerase II has passed through.