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Chromosome organization
Genome size of some of the important model organisms and
human
E. coli
Yeast
Drosophila
E. coli 4.6 Mb
Mouse Human
Yeast 12.1 MB
Drosophila 130 MB
Mouse 3 GB
Human 3.2 GB
DNA content of the haploid genome
C‐value Paradox
The DNA content of the
haploid genome of a range
of phyla. The range of
values within a phylum is
indicated by the shaded
area. The size of the haploid
genome also is known as
the C‐value.
Structural organization of genome in E. Coli
E. Coli Bacterial genome has compact organization with
• 1 circular chromosome very little space between the genes
Packaging
• Nucleoid
• DNA gyrase and topoisomerase
• HU protein tetramers
E. Coli K12
• Non coding DNA accounts for 11% of the 4.64
Mb genome.
• Non coding DNA is dispersed through out the
genome.
• It is possible that such compactness is
required to replicate the genome quickly.
E. coli genome organization
E.coli chromosomes have about 4.7 x 106 base pairs
which results in a length of 1.6 mm. An E.coli cell,
however is only 0.002 mm long . That explains why the
DNA must be folded in order to fit into the cell.
Figure 8.5 Genomes 3 (© Garland Science 2007)
Supercoiling is a good way of packaging of E. coli genome
Reduced turns Negative super coiling
Additional turns Positive super coiling
Figure 8.2 Genomes 3 (© Garland Science 2007)
E. coli genome is packed in several loops
DNA packaging proteins in E. coli
• DNA gyrase and topoisomerase
• HU protein tetramers ( tetramer packs 60bp DNA around it.)
• Several tens of other protein including ribosomal proteins are associated
Figure 8.3 Genomes 3 (© Garland Science 2007)
Experiment: E. coli DNA is packaged in several loops
Trimethylpsoralen cannot
bind to supercoiled DNA
With increasing dose of
radiation supercoiling
reduced.
However it did not occur
in single go, rather in a
gradual manner.
Figure 8.4 Genomes 3 (© Garland Science 2007)
Structural organization of genome in prokaryotes
• Bacteria can have circular or linear
Most prokaryotes contain a single, chromosome (Streptomyces, Borrelia,
double‐stranded, circular DNA
Rhodococcus, etc.).
chromosome. The remaining
prokaryotes have genomes consisting
of one or more chromosomes that may • Agrobacterium tumefaciens, there is one
be circular or linear. In the latter cases, circular chromosome and one non‐
there is typically a main chromosome homologous linear chromosome.
and one or more smaller
chromosomes. • Borrelia sp. has a very complex plasmid
The smaller chromosomes replicate content with 12 linear molecules and 9
autonomously of the main circular molecules.
chromosome and may or may not be
essential to the life of the cell.
Autonomously replicating small
chromosomes not essential to the life Archaea DNA packaging proteins are more
of the cell are known as plasmids. similar to eukaryotic histones, which is one
of main reason they are currently classified
separately from bacteria.
DNA topology
A circular DNA cannon be untangled without breaking a
phosphodiester bond
Covalently linked closed circular DNA
(cccDNA)
e.g. Bacterial chromosome,
plasmid, viral genome
Topological domain
A DNA segment constrained in such a way that it’s ends cannot rotate freely,
is called a topological domain.
Linking number
A fundamental topological parameter of a cccDNA
Important features of Linking number
Twist is another topological parameter
and is a component of linking number
For a circular DNA, twist is the total number of helical turns in
circular DNA under given conditions
Since DNA is a right‐
handed helix with 10.5
base pair per turn, Twist
is a large positive number
for any natural DNA
Twist is represented with negative
number for the left handed helix
Important features of twist
Writhe is another topological parameter
and is a component of linking number
Writhe is the number of times the double helix crosses over itself
(these are the supercoils)
Writhe can be positive or negative
Relationship between
linking number, twist and writhe
Writhe is ‘0’ for a cccDNA which lies flat on the surface
Writhes are of two types
Writhe is positive or negative based on the number required to
compensate the change in twist number
Writhe number for a
right handed helical DNA
Spiral
Spiral
Linking number can be changed by breaking
phosphodiester bond
Linking number change can cause supercoiling
DNA Topoisomers and topoisomerases
Topoisomerase can change linking number
Topoisomerase II Topoisomerase I
Change of linking number by 2
DNA gyrase a special type of Type II
topoisomerase which introduces negative Change of linking number by 1
supercoiling rather than removing them.
Topoisomerase can resolve or introduce
different entangles
Organization and regulation of
chromatin structure
Eukaryotic Nuclear Genome
Set of DNA molecules each contained in
chromosomes
Linear DNA
At least 2 chromosomes (1 pair)
Number of chromosome varies and it is
not related to genome size
Chromosome structure in metaphase
Mitotic chromosome Interphase nucleus
Figure 7.4 Genomes 3 (© Garland Science 2007)
Chromosome under EM
30 nm
11 nm
Figure 7.2a Genomes 3 (© Garland Science 2007)
Eukaryotic DNA is compacted at the first level by wrapping
around Nucleosome
Figure 7.2b Genomes 3 (© Garland Science 2007)
Nucleosomes are made up of histone octamers
H2A :2
H2B :2
H3 :2
H4 :2
‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐
8
Molecular Biology of the Cell
• Linker Histone H1
stabilizes the
nucleosome structure
by not allowing the
DNA to slide.
• It also helps to pull
nucleosomes together
into a 30 nm fiber.
Figure 7.2c Genomes 3 (© Garland Science 2007)
How to probe the organization of nucleosomes
in 11 nm fiber? Nuclease protection assay.
Chromosome under EM
Figure 7.2a Genomes 3 (© Garland Science 2007)
Zigzag and solenoid model for 30 nm
chromatin fiber
Zigzag Solenoid
From crystal structure of From cryo‐electron microscopy
tetranucleosome of 30 nm structure
Possibly 30 nm structure is fluid mosaic of several different structure
Role of histone tails in the 30 nm fiber
Most notably, H4 tails and
histone H1 plays important role
in stacking up of nucleosomes
in 30 nm structure.
30nm fiber arranged in looped domain on a
chromosome axis
Lampbrush chromosome
in amphibian oocyte
Chromatin:
The 30 nm fiber
undergo higher
level of compaction
with the help of
histone and
nonhistone DNA
binding proteins
Figure 4-72 Molecular Biology of the Cell (© Garland Science 2008)
The packaging of DNA with histones yields a chromatin fiber approximately 10 nm in diameter that is composed of chromatosomes
separated by linker DNA segments averaging about 80 base pairs in length. In the electron microscope, this 10‐nm fiber has the beaded
appearance that suggested the nucleosome model. Packaging of DNA into such a 10‐nm chromatin fiber shortens its length approximately
sixfold. The chromatin can then be further condensed by coiling into 30‐nm fibers, the structure of which still remains to be determined.
Interactions between histone H1 molecules appear to play an important role in this stage of chromatin condensation.
Chromatin fibers. The packaging of DNA into nucleosomes yields a chromatin fiber approximately 10 nm in diameter. The chromatin is
further condensed by coiling into a 30‐nm fiber, containing about six nucleosomes per turn.
The extent of chromatin condensation varies during the life cycle of the cell. In interphase (nondividing) cells, most of the chromatin (called
euchromatin) is relatively decondensed and distributed throughout the nucleus. During this period of the cell cycle, genes are transcribed
and the DNA is replicated in preparation for cell division. Most of the euchromatin in interphase nuclei appears to be in the form of 30‐nm
fibers, organized into large loops containing approximately 50 to 100 kb of DNA. About 10% of the euchromatin, containing the genes that
are actively transcribed, is in a more decondensed state (the 10‐nm conformation) that allows transcription. Chromatin structure is thus
intimately linked to the control of gene expression in eukaryotes.
In contrast to euchromatin, about 10% of interphase chromatin (called heterochromatin) is in a very highly condensed state that resembles
the chromatin of cells undergoing mitosis. Heterochromatin is transcriptionally inactive and contains highly repeated DNA sequences, such as
those present at centromeres and telomeres.
As cells enter mitosis, their chromosomes become highly condensed so that they can be distributed to daughter cells. The loops of 30‐nm
chromatin fibers are thought to fold upon themselves further to form the compact metaphase chromosomes of mitotic cells, in which the
DNA has been condensed nearly 10,000‐fold. Such condensed chromatin can no longer be used as a template for RNA synthesis, so
transcription ceases during mitosis. Electron micrographs indicate that the DNA in metaphase chromosomes is organized into large loops
attached to a protein scaffold, however, currently our understanding very poor about the detailed structure of this highly condensed
chromatin and the mechanism of chromatin condensation.
The Cell, 2nd edition, A Molecular Approach, Geoffrey M Cooper.
Smc proteins forms condensin complexes which
coil long loops of chromatin domains
SMC: Structural Maintenance of Chromosomes
Condensin may provide structural frame work
to maintain compact chromatin organization in
metaphase
Condensin
Cohesin and condensin
Cohesin Condensin
Chromosome assembly and segregation
Replicated chromatids (sister chromatids) are
held together by the action of cohesin. At the
onset of mitosis, bulk cohesin dissociates from
chromosome arms whereas condensin
associates with them to induce condensation.
These processes lead to the formation of
metaphase chromosomes in which sister
chromatids are microscopically distinguishable
from each other (resolution). In late mitosis,
residual cohesin is cleaved, thereby promoting
irreversible separation of sister chromatids.
http://www.asi.riken.jp/en/laboratories/chieflabs/ch‐dyna/
Different types of sequences in
eukaryotic genome
DNA content of the haploid genome
The DNA content of the haploid
genome of a range of phyla. The
range of values within a phylum
is indicated by the shaded area.
The size of the haploid genome
also is known as the C‐value.
C‐value Paradox
Higher organism’s genome can have large percentage of
repetitive DNA.
In general, the length of the non‐repetitive DNA component tends to increase as
we go up the evolutionary tree and reaches a maximum in mammals (2 × 109 bp)
Composition of human genome
Figure 7.13 Genomes 3 (© Garland Science 2007)
Centromeric DNA is repetitive in nature
Arabidopsis: 180 bp repeat, ~1 mb
Do contain interspersed repeats
Do contain genes in lower density
Human: 171 bp alphoid DNA, 1500‐30000 repeats
~0.25 ‐5 mb
Telomeres are repetitive G‐rich DNA
The special structure of
TRF1: Maintains length
telomere differentiates it from
TRF2: Maintain 3’ overhang
other DNA breaks
Figure 7.10 Genomes 3 (© Garland Science 2007)
Telomerase
Gene
A gene is the basic physical and functional unit of heredity. Genes in living organisms are
made up of DNA. Some genes act as instructions to make molecules called proteins.
However, many genes do not code for proteins. In humans, genes vary in size from a few
hundred DNA bases to more than 2 million bases.
5’ UTR ORF 3’ UTR
UTR : Untranslated region
ORF: Open reading frame
Prokaryotes and eukaryotes have different gene organization
Prokaryotes
Transcription
Mature mRNA
Eukaryotes
Eukaryotic and bacterial genes are
organized differently. A bacterial
gene consists of a single stretch of
uninterrupted nucleotide sequence
that encodes the amino acid
Transcription sequence of a protein (or more
than one protein). In contrast, the
protein‐coding sequences of most
eukaryotic genes (exons) are
pre‐mRNA interrupted by noncoding
Splicing sequences (introns). Promoters for
transcription are indicated in green.
Mature mRNA
Organization of genes on eukaryotic chromosomes
Storage and transfer of genetic information
Central dogma of transfer
of genetic information
The central dogma of molecular biology is
an explanation of the flow of genetic
information within a biological system.
https://en.wikipedia.org/wiki/Central_dogma_of_molecular_biology
Updated central dogma
(HIV)
https://en.wikipedia.org/wiki/Central_dogma_of_molecular_biology
