Genome organisation and
evolution
Level 3 Molecular Evolution and
Bioinformatics
Jim Provan
Page and Holmes: Sections 3.1.4/5 and 3.3
The eukaryotic genome
Single-copy protein
coding genes
Coding DNA
Dispersed
Multigene families
Regulatory sequences
Tandemly repeated
Satellite DNA
Non-coding
DNA
Tandemly repeated
DNA
Transposable elements
And retroviruses
Spacer DNA
Minisatellites
Microsatellites
The C-value paradox
The amount of DNA per
haploid genome is known
as the C-value
Contrary to expectation,
the amount of DNA is not
correlated with complexity:
The protist, Amoeba dubia
has about 200 times more
DNA (670,000,000 kbp) than
humans (3,300,000 kbp)
Cannot be explained by
differences in gene number
12
10
8
6
4
2
0
The structure of genes
There are many forms of genes:
Those which produce a protein, a tRNA or an rRNA are referred
to as structural genes
Those which control how and when genes are expressed are
called regulatory genes
Some housekeeping genes need to be expressed in all tissues
e.g. those involved in protein synthesis
Other, tissue-specific genes, are only expressed in a particular
cell or tissue type e.g. the insulin gene is only expressed in the
pancreatic β-cells
Whatever their function, all genes contain a coding
region which specifies a polypeptide or an RNA molecule
Regulation of gene expression
Coding regions of genes are usually flanked by
regulatory regions which control gene expression
through transcription and translation
Upstream promoter regions:
– In bacteria, there is a Pribnow box (TATAAT) about 10 bp
upstream from where transcription starts, the ‘-35 site’
(TTGACA) about 35 bp upstream and the Shine-Dalgarno box
(AGGAGG) about 7 bp before the initiation codon
– In eukaryotes, as well as the TATA box, some promoter regions
contain a CAAT box about 40 bp before initiation codon and a
GC box (GGGCGG) about 110 bp upstream
Downstream elements such as the polyadenylation signal
(AATAA) signify the end of transcription and increase
stability of RNA transcripts
Structure of a typical gene - alcohol
dehydrogenase (Adh)
Promoter region
• TATA box
• CAAT box (in mammals)
• GC box (GGGCGGG)
Exon 1
Polyadenylation
signal
AATAA
Eukaryote
Exon 2
Exon 3
Exon 4
5’
3’
Intron 1
Intron 2
Intron 3
Initiation codon
Stop codon
Promoter region
• Shine-Dalgarno box (AGGAGG)
• Pribnow box (TATAAT)
Prokaryote
• -35 site (TTGACA)
5’
3’
Initiation codon
Stop codon
Introns
Occur frequently within eukaryotic genomes and
make up most of the length of very long genes
Number, size and organisation of introns varies:
Histones have no introns: chicken pro-a2-collagen gene has
over fifty
SV40 virus contains an intron of 31 bp: human dystrophin
gene has an intron of over 210,000 bp
Some introns have genes contained within them - the Adh
gene in Drosophila is located within the intron of the
outspread gene
Strong conservation of intron-exon boundaries nearly always begin with GT and end with AG
Types of introns
Most introns in eukaryotes are spliceosomal
introns (‘nuclear introns’) because they are
spliced by a spliceosome of proteins and RNA
Some introns can splice without the aid of
proteins (“self-splicing introns”):
One class - group I introns - are sometimes mobile because
they encode proteins such as DNA endonucleases. They are
found in mitochondrial and chloroplast genomes, rRNAs of
some eukaryotes and in T4 bacteriophage
Group II introns are found in organelles and their bacterial
ancestors and contain reverse transcriptase-like sequences
Group III introns are found in a few protists and are similar
to group II introns with the central portion removed
The evolution of introns
There are two competing hypotheses for the
evolution of spliceosomal introns:
The introns-early hypothesis, proposed by Walter Gilbert,
suggests that introns mark the boundaries between ancient
genes which encoded distinct proteins.
Throughout evolution these once-independent proteins have
been put together in new combinations to produce more
complex proteins by exon shuffling
An alternative hypothesis (introns-late) suggests that introns
only invaded eukaryote genomes fairly recently
The evolution of introns (continued)
A crucial prediction of the introns-early hypothesis is
that spliceosomal introns delineate structural or
functional units within proteins:
Introns are found in the same places in all known globin
genes, including myoglobin and plant leghaemoglobins
More frequently, however, introns do not appear to separate
functionally distinct parts of proteins
Other problem with introns-early hypothesis is
absence from Archaea and Bacteria:
Massive intron loss has been postulated but does not explain
why they are found in nuclear copies of organelle genes but
not in the genes of the organelles or their precursors
Exon shuffling has probably been a factor in later eukaryotes
Multigene families
Many genes are found not as individual copies but as
part of multigene families, larger families of related
genes:
Important evolutionary innovation: proteins with similar
function can be arranged so that they are regulated efficiently
Vertebrates have a variety of multipolypeptide globin genes,
produced by gene duplication, which are adapted to varying
oxygen requirements of different developmental stages
Not all genes are functional:
Pseudogenes arise through gene duplications but acquire
mutations since only one copy is required
Processed pseudogenes, which lack promoters and introns,
have been produced by reverse transcription of mRNA
Multigene families (continued)
e
Foetal
g1
g2
Pseudogene
yh
Adult
d
b
0
100
200
Millions of years ago
Embryonic
Evolution of multigene families
Most obvious way in which gene number can change
between species is through gene duplication:
Can arise through unequal crossing-over
May occur by duplication of entire genomes (polyploidy):
– Common in plants: around 50% of angiosperms are polyploid
– Xenopus laevis is tetraploid: normal meiosis is possible
– Other members of the genus Xenopus have chromosome numbers
ranging from 20 to 108
Another mechanism of gene duplication is transposition
Fate of new gene depends on function: redundancy vs.
natural selection
Genes can also acquire new functions without
duplication e.g. e-crystallin and LDH
Gene duplication in the Hox gene family
Homeotic genes control
the development of
body plan in animals
In both vertebrate Hox
and invertebrate HOM
genes, there is a highly
conserved protein motif
known as a homeobox
Mutations in Hox/HOM
genes can drastically
affect the organisation
of body parts
Gene duplication in the Hox gene family
Although Hox/HOM genes are related, their
organisation differs between organisms:
In vertebrates, there are multiple clusters of Hox genes: the
mouse has four clusters, each located on a different
chromosome and covering over 100 kb
HOM genes in Drosophila are found in two clusters,
Antennipedia and Bithorax, on the same chromosome
In amphioxus – a class of marine invertebrates which are the
closest relatives to the vertebrates – there is a single cluster of
at least 10 Hox genes each of which is homologous to a
different Hox gene in vertebrates: origin of vertebrates
coincided with a series of gene duplications
Example of a dispersed gene family in vertebrates
Gene duplication in the Hox gene family
Gene
Duplications
(four clusters)
Amphioxus
Hypothetical
Common
Ancestor
Drosophila
lab
pb
Dfd
Scr
Antp
Ubx
AbdA
AbdB
Tandem arrays
Tandem arrays contain
multiple copies of genes
with the same function
Good example is the rDNA
array:
Large quantities of rRNA
required
Genes and spacers cotranscribed and separated by
non-transcribed spacer
Variation in size of arrays:
– 1 copy in Tetrahymena
– 19,300 copies in Amphiuma
ETS
ITS1 ITS2
NTS
18S
5.8S
28S
Evolution of rDNA arrays
Because they contain both highly conserved (18S) and
highly variable (NTS) regions, rDNA sequences have
been used frequently in molecular systematics
Despite this, they do not evolve in a simple manner:
Although there is a high degree of sequence similarity within
species, there is great divergence between them
Due to unequal crossing-over and gene conversion, concerted
evolution can take place which allows genes to evolve together
by spreading mutations throughout members
This makes phylogenetic analysis difficult since it is not easy to
discern which genes are truly homologous
Often leads to “mosaics” of sequences, each with different
phylogenetic history
Non-coding repetitive DNA
Class
Copy number
Organisation
Satellite DNA
Highly repetitive (>104)
Tandemly repeated
Mini-/microsatellite
Moderately repetitive
Tandemly repeated
Transposable elements
Moderately/highly repetitive Dispersed
Tandemly repeated DNA
Much of the non-coding repetitive DNA in eukaryotes
consists of tandem repeats of short sequence motifs:
Satellite DNA is located mainly in the heterochromatin and
consists of motifs up to 40 kb in length:
– The a-satellite DNA of primates based on a 171 bp motif repeated
for hundreds of kilobases
– Over 60% of the genome of Drosophila nasutoides is satellite DNA
Minisatellites and microsatellites are comprised of shorter motifs
duplicated through unequal crossing over and DNA slippage:
– Minisatellites motifs are 11 – 60 bp in length and contain a G-rich
“core” sequence
– Microsatellites are shorter, generally dinucleotide repeats
– Both exhibit extremely high mutation rates and multiple alleles are
usually found in populations
– Used in population genetics / forensics
Transposable elements
Transposable elements increase copy number by moving
around the genome making additional copies:
Around 50% of the maize genome may be transposable elements
10-20% of the Drosophila genome
Three groups of transposable elements:
Class I (retroelements) transpose through an intermediate RNA
stage via reverse transcriptase cf. retroviruses
Class II (DNA elements) transpose directly from DNA to DNA
Little is known about miniature inverted-repeat transposable
elements (MITEs): around 100 – 400 bp in length and transpose
by as yet unknown means
Transposable elements
Class I transposable elements (retroelements)
Retrotransposons
LTR
Retroposons
Reverse transcriptase
LTR
Reverse transcriptase
AAAAAA
Class II transposable elements (DNA elements)
Ac-like elements
Transposase
Miniature inverted-repeat transposable elements (MITEs)
Short repeat
e.g. Tourist and Stowaway
Terminal repeat
Retroelements
Two subgroups:
Retrotransposons contain long terminal repeats at both ends:
example is copia element which is found 20 – 60 times in the
genome of D. melanogaster
Retroposons have no LTR and have a poly-A tail:
– Long interspersed nuclear elements (LINEs) are 6 – 8 kb in length
and present in thousands of copies: the L1 family is present in
590,00 copies in the human genome (17% of total)
– Short interspersed nuclear elements (SINEs) do not produce reverse
transcriptase and so are not considered true retroelements: they
vary in size from 130 – 300 bp and have copy numbers from 50,000
to over 1,000,000
– Originally derived from RNA transcripts
Endogenous retroviruses are proviruses which have been
integrated into the germ-line of eukaryotes
Class II (DNA) elements
Possess terminal repeats but unlike retrotransposons
these are short (generally < 100 bp) and usually inverted
Encode a special transposase protein
Best known types:
Mariner elements in animals
Hobo and P elements in Drosophila:
– P elements can move between species and affect host phenotype
– Increased infertility due to chromosome breakage (hybrid
dysgenesis) occurs in D. melanogaster. P elements are not found in
closely related species (D. simulans, D. sechellia, D. mauritania) but
are found in more distantly related species e.g. D. willistoni group:
transferred after D. melanogaster split from sibling species
– Insertion can have “knock-out” effect on phenotype e.g. white gene
in flies lacking red eye pigment