Chromatin

Chromatin is the complex of DNA and protein that makes up chromosomes. In eukaryotes chromatin is found inside the nuclei of eukaryotic cells, while in prokaryotes, the chromatin is held within the nucleoid. The nucleic acids are in the form of double-stranded DNA (a double helix). The major proteins involved in chromatin are histone proteins, although many other chromosomal proteins have prominent roles too. The functions of chromatin are to package DNA into a smaller volume to fit in the cell, to strengthen the DNA to allow mitosis and meiosis, and to serve as a mechanism to control expression. Changes in chromatin structure are affected mainly by methylation (DNA and proteins) and acetylation (proteins). Chromatin structure is also relevant to DNA replication and DNA repair.

Chromatin is easily visualised by staining, hence its name, which literally means coloured material.



Simplistically, there are three levels of chromatin organization (Fig. 1):
 * 1) DNA wrapping around nucleosomes - The "beads on a string" structure.
 * 2) A 30 nm condensed chromatin fiber consisting of nucleosome arrays in their most compact form.
 * 3) Higher level DNA packaging into the metaphase chromosome.

These structures do not occur in all eukaryotic cells; there are examples of more extreme packaging, for example spermatozoa and avian red blood cells.

The different levels of chromatin compaction are clearly visible in cells. In non-dividing cells there are two types of chromatin: euchromatin and heterochromatin. These correspond to uncompacted actively transcribed DNA and compacted untranscribed DNA.

The structure of chromatin varies considerably as the cell progresses through the cell cycle. The changes in structure are required to allow the DNA to be used and managed, whilst minimising the risk of damage.

Interphase Chromatin
The structure of chromatin during interphase (the period of normal cell function between divisions) is optimised to allow easy access of transcription and DNA repair factors to the DNA while compacting the DNA into the nucleus. The structure varies depending on the access required to the DNA, for example expressed genes require regular access by RNA polymerase and so have a looser structure - euchromatin.

Change in chromatin structure
Chromatin undergo various form of changes in its structure. Histone proteins which is foundation block of chromatin get modified by various post-translational modification to alter the DNA packing. Histone proteins when acetylated result in loosening of chromatin and help in replication, transcription of gene as unpacked DNA get easy access to various enzymes like DNA,RNA polymerase). Histone proteins when get methylated strongly holds DNA together and prevents its access to various enzymes.A recent study show that there is a bivalent structure present in the chromatin. This structure is methylated lysine residues at location 4 and 27 on histone 3. It is thought that these methylation may be responsible in some case to development.Its found that there is more methylation of lysine 27 in embryonic cells than in differentiated cells ,whereas lysine 4 methylation is positively regulates transcription by recruiting nucleosome remodeling enzymes and histone acetylases.

DNA structure


The vast majority of DNA within the cell is the normal DNA structure. However in nature DNA can form three structures, A-, B- and Z-DNA. A and B-DNA are very similar, forming right handed helices, while Z-DNA is a more unusual left handed helix with a zig-zag phosphate backbone. Z-DNA is thought to play a specific role in chromatin structure and transcription because of the properties of the junction between B- and Z-DNA.

At the junction of B- and Z-DNA one pair of bases is flipped out from normal bonding. These play a dual role of a site of recognition by many proteins and as a sink for torsional stress from RNA polymerase or nucleosome binding.

The Nucleosome and "Beads-on-a-String"

 * Main articles: Nucleosome, Chromatosome and Histone

The basic repeat element of chromatin is the nucleosome, linked by sections of linker DNA. The is far smaller than can be reached by DNA in solution.

In addition to the core histones there is the linker histone, H1, which contacts the exit/entry of the DNA strand on the nucleosome. The nucleosome, together with histone H1, is known as a chromatosome. Chromatosomes, connected by about 20 to 60 base pairs of linker DNA, form an approximately 10nm "beads-on-a-string" fibre. (Fig. 1-2).

The nucleosomes bind DNA non-specifically, as required by their function in general DNA packaging. There is, however, some preference in the sequences the nucleosomes will bind. This is largely through the properties of DNA, adenosine (A) and thymine (T) bases are more favorably compressed into the inner minor grooves. This means nucleosomes bind preferentially at one position every 10 base pairs - where the DNA is rotated to maximise the number of A and T bases which will lie in the inner minor groove. See mechanical properties of DNA.

30nm chromatin fibre
The "beads-on-a-string" structure in turn coils into a 30nm diameter helical structure known as the 30nm fibre or filament. The precise structure of the chromatin fibre in the cell is not known in detail, and there is still some debate over the exact structure. There are, however, three well established models.

This level of chromatin structure is thought to be the form of euchromatin, which contains actively transcribed genes. EM studies have demonstrated the 30nm fibre is highly dynamic such that it unfold into a 10nm fiber ("beads-on-a-string") structure when transversed by an RNA polymerase engaged in transcription.

The three models are based on the accepted facts that the nucleosomes lie perpendicular to the axis of the fibre, the linker histones lie on the inside of the structure and that it readily unwinds into the 10nm "beads-on-a-string" fibre.

A stable 30nm fibre relies on the regular positioning of nucleosomes along the DNA. The mechanical properties of DNA mean linker DNA is relatively resistant to bending and rotation. This makes the length of linker DNA critical to the stability of the fibre, nucleosomes must separated by lengths which allow them to rotate and then fold into the required orientation without significant stress to the DNA.

Spatial Organization of Chromatin in the Cell Nucleus
The layout of the genome within the nucleus is not random - specific regions of the genome are always found in certain areas, and un-transcribed regions clump together into the nucleolus. Specific regions of the chromatin are thought to be bound to the nuclear membrane, while other regions are bound together by protein complexes. The layout of this is not, however, well characterised.

One well characterised aspect of genome layout within nucleus (in mammals) is the compaction of one of the two X chromosomes in females into the barr body. This serves the role of permanently deactivating these genes, which prevents females getting a 'double dose' of these genes relative to males.

Metaphase Chromatin
The metaphase structure of chromatin is vastly different to during interphase. The structure of the chromatin is optimised for physical strength and manageability, forming the classic chromosome structure seen, for example, in karyotypes.

The structure of the condensed chromosome is thought to be loops of 30nm fibre to a central scaffold of proteins. It is, however, not well characterised, and little solid evidence exists.

The physical strength of chromatin is vital for this stage of division to prevent shear damage to the DNA as the daughter chromosomes are separated. To maximise strength the composition of the chromatin changes as it approaches the centromere, primarily through alternative histone H1 anologues.

Non-Histone Chromosomal Proteins
The proteins that are found associated with isolated chromatin fall into several functional categories:
 * chromatin-bound enzymes
 * high mobility group (HMG) proteins
 * transcription factors
 * scaffold proteins
 * Transition proteins (testis specific proteins)
 * protamines (present in mature sperm)

Enzymes associated with chromatin are those involved in DNA replication and repair, in transcription, and in post-translational modification of histones. Examples are various types of nucleases and proteases. Scaffold proteins encompass chromatin proteins such as insulators, domain boundary factors and cellular memory modules (CMMs).

Chromatin: Alternative Definitions

 * 1) Simple & Concise Definition: Chromatin is DNA plus the proteins (and RNA) that package DNA within the cell nucleus.
 * 2) A Biochemists’ Operational Definition: Chromatin is the DNA/protein/RNA complex extracted from eukaryotic lysed interphase nuclei. Just which of the multitudinous substances present in a nucleus will constitute a part of the extracted material will depend in part on the technique each researcher uses. Furthermore, the composition and properties of chromatin vary from one cell type to the another, during development of a specific cell type, and at different stages in the cell cycle.
 * 3) The DNA plus Histone – Equals – Chromatin - Definition: The DNA double helix in the cell nucleus is packaged by special proteins termed histones. The formed protein/DNA complex is called chromatin. The structural entity of chromatin is the nucleosome.

History
In 1882 Walther Flemming used the term Chromatin for the first time. Flemming assumed that within the nucleus there was some kind of a nuclear-scaffold. Further there were nucleoli, the nuclear plasm and the nuclear membranes. He wrote (transl. from German): “The scaffold owes its capability of refraction, the way how it behaves, and in particular its colorability to a substance which, with regard to its latter attribute, I have termed Chromatin. It is possible that this substance is really identical with the Nuclein-bodies. .... I’ll retain the name Chromatin as long as Chemistry has decided about it, and I empirically refer to it as that substance in the cell's nucleus which takes up the dye upon staining the nucleus ("Kerntinktionen").

Nobel Prizes Related to Chromatin
Albrecht Kossel (University of Heidelberg) was awarded the Nobel Prize in Physiology or Medicine 1910 "in recognition of the contributions to our knowledge of cell chemistry made through his work on proteins, including the nucleic substances".

Thomas Hunt Morgan (California Institute of Technology) was awarded the Nobel Prize in Physiology or Medicine 1933 "for his discoveries concerning the role played by the chromosome in heredity".

Francis Crick, James Watson and Maurice Wilkins (MRC Laboratory of Molecular Biology, Harvard University, London University) were awarded the Nobel Prize in Physiology or Medicine 1962 "for their discoveries concerning the molecular structure of nucleic acids and its significance for information transfer in living material".

Aaron Klug (MRC Laboratory of Molecular Biology) was awarded the Nobel Prize in Chemistry 1982 "for his development of crystallographic electron microscopy and his structural elucidation of biologically important nucleic acid-protein complexes".

Roger Kornberg (Stanford University) was awarded the Nobel Prize in Chemistry in 2006 "for his studies of the molecular basis of eukaryotic transcription". In 1974 published the discovery that the nucleosome consists of a tetramer of histones H3 and H4 and two dimers of histones H2A and H2B.