DNA ORGANIZATION IN

EUKARYOTIC CELLS

CONTENTS• Facts about DNA• Eukaryotic chromosomes• Chemical composition of eukaryotic chromosomes• Histones• Non-histone chromosomal protein• Scaffold proteins• Folded fibre model• Nucleosome model• H1 proteins• Histone modification• Chromatosome• Higher order of chromatin structure• Mechanism of DNA packaging• Conclusion

FACTS ABOUT DNA..!• DNA stores all the information that makes up

an organism.

• DNA are made up of four building blocks,which are- cytosine(C), Thymine(T), Guanine (G) and adenine(A).

• Each DNA strand is about 1.8 meters long but squeezed into a space of 0.09 micrometers.

• 99.9% of DNA is identical in all humans on this Earth. The remaining 0.1% is what helps us to differentiate between DNA sequences allowing us to tell which DNA belongs to whom.

• Human DNA is 95% identical to DNA of chimpanzees and interestingly it is 50% identical to the DNA of the banana.

THE EUKARYOTIC

CHROMOSOME • Individual eukaryotic chromosome contain enormous amount of DNA.

• For all of these DNA to fit into the nucleus, tremendous packaging and folding are required.

• The chromosomes are in an elongated, relatively uncondensed state during interphaseof the cell cycle.

MEIOSIS

MITOSIS

THE EUKARYOTIC CHROMOSOME

• In the course of the cell cycle , the level of DNA packaging changes

• The packaging of eukaryotic DNA is not static.

The Eukaryotic Chromosome

• The haploid chromosome complement, or genome, of a human contains about 1000mm of DNA (or 2000mm per diploid cell).

• This meter of DNA is subdivided among 23 pair chromosomes.

• Chromosomes are of variable shape and size.

• Depending on shape and size, each chromosome contain approximately 15 to 85mm of DNA.

CHEMICAL COMPOSITION OF

EUKARYOTIC CHROMOSOME

• DNA is closely associated with proteins, creating chromatin.

• The two basic types of chromatins are:

-Euchromatin ,which undergoes the normal process of condensation and decondensationin the cell cycle.

-Heterochromatin , which remain in a highly condensed state throughout the cell , even during interphase.

CHEMICAL COMPOSITION OF

EUKARYOTIC CHROMOSOME• Chemical analysis of isolated chromatin shows

that it consists primarily of DNA and proteinswith lesser amount of RNA.

• The proteins are of two major classes:

o Basic (positively charged at neutral pH) proteins called histones.

o A heterogenous , largely acidic (negatively charged at neutral pH) group of proteins collectively referred to as non-histonechromosomal proteins.

HISTONES

H1

H2AH2B

H3

H4

HISTONES• Histones are found in the nucleus of the eukaryotic cell.

• Histones play a major structural role in chromatin.

• They are present in the chromatin of all eukaryotes in amounts equivalent to the amount of DNA.

• This relationship suggests an interaction occurs between histones and DNA that is conserved in eukaryotes.

• Four of the five types of histones are similar in all eukaryotes.

HISTONES

• The histones of all plants and animals consists of five classes of proteins.

• These five major histones types, called H1, H2A, H2B, H3, H4, are present in almost all cell types.

• The five histone type are present in molar ratios of approximately 1:H1,2:H2A,2:H2B, 2:H3,2:H4.

HISTONES • Four of the five types of histonesare specifically complexed with DNA to produce the basic structural subunits of chromatin, small (approximately 11nm in diameter by 6nm high) ellipsoidal beads called nucleosome.

• The histones are basic. • Because they contain 20% to 30% arginine

and lysine, two positively charged amino acids.

• The exposed –NH3+ groups on histones are

important in their interaction with the DNA, which is polyanionic because of the negatively charged phosphate group.

HOW DNA IS PACKAGED?

• To explain how 2meters of DNA is packaged into a nucleus of 10 micro-meter and how the stability of the DNA is maintained

• There were many models given by different scientist out of which two models stand out.

FOLDED FIBRE MODEL

• It was proposed by E.J. Dupraw.According to the model:• Chromosome consist of tightly folded fibre

of 20-30nm diameter.• Folded fibre consist of DNA histone helix

of 3nm in a supercoiled condition.• Histones were attached on the outside of

the DNA coils that is histone shells around DNA.

NUCLEOSOME MODEL

• It was proposed by R.D. Kornberg and confirmed by P.Outdet.

The model states that:• Nucleosome is the lowest level of

organization.• Nucleosome consist of a disc shaped

structure of 11nm in diameter.• Comprising of two parts: a core particle

and a small spacer or linker DNA.

NUCLEOSOME- basic repeating unit of chromatin.

THE NUCLEOSOME

• The basic repeating unit of chromatin.

• It provides the lowest level of compaction of double-stranded DNA into the cell nucleus.

• In an electron microscope, isolated chromatin frequently looks like “beads on a string”.

EXPERIMENT WITH NUCLEASE• If small amount of nuclease is added to this beads

on string structure, enzyme cleaves the string between the beads , leaving the individual beads attached to about 200bp of DNA.

• If more nuclease is added , the enzyme chews up all of the DNA between the beads and leaves a core protein attached to a fragment of DNA.

• Such experiments demonstrated that chromatin is not a random association of proteins and has a fundamental repeating structure.

• The repeating core of proteins and DNA produced by digestion with nuclease enzymes is the simplest level of chromatin structure, the nucleosome.

NUCLEOSOMES• Nucleosome is a core

particle consisting of DNA wrapped about two times around the octamer of eight histone proteins (two copies each of H2A, H2B, H3 and H4) and H1 as linker DNA.

• The DNA in direct contact with the histone octamer is between 145bp to 147bp.

• The DNA coils around the histones in left-handeddirection, and is supercoiled.

NUCLEOSOME• The tail of the histone

molecules protude from the nucleosome and are accessible to enzymes that add and remove chemical groups such as methyl and acetyl groups.

• The addition of these group can change the level of expression of genes packaged in nucleosome containing modified histones.

THE FIFTH TYPE OF HISTONE, H1

• H1, histone, is not a part of the core particle.

• It plays an important role in the nucleosomestructure.

• The precise location of H1 with respect to the core particle is still uncertain.

• In general view, H1 sits outside the octamer and binds to the DNA where the DNA joins and leaves the octamer.

H1, HISTONE• However, recent

experiment suggest that the H1 histonessits inside the coils of the nucleosome.

• Regardless of its position, H1 helps to lock the DNA into place, acting as clamp around the nucleosome octamer.

CHROMATOSOME

• Together, the core particle and its associated H1 histoneare are called chromatosome, the next level of chromatin organization.

• The H1 protein is attached to between 20 and 22bp of DNA, and the nucleosomeencompasses an additional 145 to 147bp of DNA; so about 167bp of DNA are held within the chromatosome.

CHROMATOSOME• Chromatosome are

located at regular intervals along the DNA molecules

• They are separated from one another by linker DNA , which varies in size among the cell types.

• Non-histonechromosomal protein maybe associated with this linker DNA and a few also appear to bind directly to the core particles.

NON-HISTONE CHROMOSOMAL PROTEIN

Non-histonechromosomal

protein

serve as structural

roles.

takes part in genetic processes

such as transcription and

replication.

SCAFFOLD PROTEINS

• Scaffold means -to provide or supportwith a raised frame-

work or platform.

• A protein whose main function is to bring other proteins together for them to interact.

SCAFFOLD PROTEINS

• Chromatin is treated with a concentrated salt solution.

• It removes histones and most of the other chromosomal proteins,

• Leaving a “skeleton” to which DNA was attached, known as scaffold proteins.

• These scaffold proteins play a role in the folding and packaging of chromosome.

SCAFFOLD PROTEINS• Histone-depleted chromosomes

were studied in the electron microscope.

• Results show that:

• the histone-depleted chromosomes consist of a scaffold or core, which has the shape characteristic of a metaphase chromosome, surrounded by a halo of DNA;

• the halo consists of many loops of DNA, each anchored in the scaffold at its base;

• most of the DNA exists in loops at least 10-30 µm long (30-90 kilobases).

THREE LEVELS OF DNA

PACKAGING

HIGHER-ORDER CHROMATIN STRUCTURE

• The first level of condensation involves packaging DNA as a negative supercoil into nucleosomes , to produce the 10nm interphase chromatin fiber. This clearly involves an octamer of histone molecules, two each of histones H2A, H2B, H3 and H4.

• In chromosomes, adjacent nucleosome are not separated by space equal to the length of the linker DNA.

• Rather nucleosomes fold on themselves to form a 30nm dense, tightly packed structure.

• The next-higher level of chromatin structure is a series of loop of 30nm fibres, each anchored at its base by proteins in the nuclear scaffold.

Nucleosome fold on themselves to form a dense fibre of 30nm in diameter.

HIGHER-ORDER CHROMATIN STRUCTURE

• On average, each loop encompasses some 20,000 to 100,000bp of DNA and is about 300nm in length, but the individual loop vary considerably.

• The 300nm fibres are packed and folded to produce a 250nm wide fibre.

• Tight helical coiling of the 250nm fiber, in turn, produces the structure that appears in metaphase: an individual chromatid approximately 700nm in width.

METAPHASE• Metaphase chromosomes are

the most condensed of normal eukaryotic chromosome.

• The role these highly condensed chromosomes is to organize and package the giant DNA molecules of eukaryotic chromosomes into structures that will facilitate their seggregation to daughter nuclei without the DNA molecules of different chromosomes becoming entangled.

METAPHASE• There is evidence that

the gross structure of metaphase chromosomes is not dependent on histones.

• Electron micrographs of isolated metaphase chromosome from which the histones have been removed revealed a scaffold, which is surrounded by huge pool of DNA.

• This chromosome must be composed of non-histone chromosomal protein.

CONCLUSION• Each DNA strand is about 1.8 meters long but squeezed into a

space of 0.09 micrometers.

• Individual eukaryotic chromosome contain enormous amount of DNA.

• The packaging of eukaryotic DNA is not static.

• Eukaryotic chromosomes are made up two major types of proteins-basic and largely acidic.

• Histones are of five types- H1, H2a, H2b, H3 and H4.

• Two units of each histone proteins wrapped by DNA, except H1, forms the nucleosome.

• Nucleosome along with H1 forms the chromatosome.

• Each chromatosome is attached by linker DNA.

• They coil round to form a 30nm wide fibre, that forms loop averaging from 300nm in length.

• The 300nm fibre are further compressed and folded to form 250nm wide fibre.

• Tight coiling of the 250nm fibre forms chromatids of chromosomes.

REFERENCES

• https://www.youtube.com/watch?v=3wTAEfjo20c&list=WL

• http://giphy.com/search/chromosome

• http://giphy.com/gifs/astronomy-lOe630aMEzo76

• D.Peer Snustad, Michael J. Simmons,Principle of Genetics,Fifth Edition.

• Eldon John Gardner, D.Peer Snustad, Michael J. Simmons,Principle of Genetics,Eight Edition.

• Peter J. Russell, iGenetics,Third Edition.