Tandemly repeated satellite DNA in the Artiodactyla - a lecture
Tandemly repeated, satellite, DNA sequences are an abundant component of the genome of most species, including the Artiodactyla. Multiple DNA familes are present, each in long tandem arrays, with members of each family present on one or more chromosomes at characteristic positions. In particular, several familes are located at the centromeres of most chromosomes, including acrocentrics, metacentrics and the sex chromosomes. Individual arrays are made up of variants of particular sequence motifs, which may be longer than 1,500 bp. In this presentation, we will discuss aspects of the evolution of repetitive sequences within and between chromosomes, with comparative data between different species. With pig, we will show details of the localization of tandem repeats at meiosis, and how these sequences relate to sequence amplification and loss, as well as the epigenetic behaviour of the resulting heterochromatin. In the Bovinae, we will show how molecular cytogenetic methods are essential to build up a full picture of the behaviour and distribution of satellite DNA where current sequencing methods are unable to assemble the sequences blocks accurately.
P. Heslop-Harrison1, T. Schwarzacher1 and R. Chaves2 (Phh4@le.ac.uk)
University of Leicester, Biology, Leicester LE1 7RH UK; 2Institute for Biotechnology and Bioengineering, Centre of Genomics and Biotechnology, University of Trás-os-Montes and Alto Douro, Vila Real, Portugal
Tandem Repeats and Satellite DNA in Bovideae - Colloquium on Animal Cytogenetics
1. Tandemly repeated
satellite DNA in the
Artiodactyla
Pat Heslop-Harrison
Trude Schwarzacher
Raquel Chaves
University of Leicester, UK
Universidade de Trás-os-Montes e Alto Douro, Portugal
www.molcyt.com
Twitter/slideshare Pathh1
phh@le.ac.uk
2. Chromosomal changes are one of the
most important features of evolution
Bovideae: 58 autosomal arms
Cow: 2n=60 (29 pairs of
acrocentric autosomes + X, Y)
Sheep: 2n=54 (25 acrocentric + 2
submetacentric pairs + X, Y)
3. Bos taurus taurus vs Bos taurus indicus:
2n=60, XY
But: B. taurus submetacentric Y
B. indicus acrocentric Y
4. Robertsonian Fusion of 1 and 29 to
give 2n=58 or 59: Gustavson 1964
Heterozygous rob(1;29) example in Portuguese
cattle Barrosa Chaves et al. Chromosome Research
5. Repetitive DNA sequences –
LINE/SINE transposons and satellite
DNA – are the most abundant genome
component
- Often ‘masked’ (ignored) during
sequence assembly
- Satellites ‘collapse’ from hundreds
of tandem repeats to a few
- Often functional regarding centromeric
behaviour and methylation/
heterochromatinization
8. Complex satellite DNA reshuffing in the polymorphic t(1;29) Robertsonian translocation and
evolutionarily derivedchromosomes in cattle Chaves, Adega, Heslop-Harrison et al. 2003
10. Barrosa¬
Order Artiodactyla
(Even-toed ungulates)
3 groups:
1. Suiformes (pigs, peccaries,
hippopotamuses),
2. Tylopoda (camels, llamas)
3. Ruminantia (cattle, goats, sheep, deer,
antelopes, giraffes)
9 families (13 tribes) including Bovidinae
Family Bovidae
c. 137 species
Last species (new genus) discovered in 1992
11. Domestic pig Sus scrofa domestica
Centromeric
satellites
METACENTRIC CLONES:
GC rich centromeric
heterochromatin
• Clone pAL7.5 (“Al”): present
in all metacentric
chromosomes (SSC1 – SSC12
and X ); 294bp
• Clone pAv1.5 (“Av”): present
only in SSC1; 313bp
ACROCENTRIC CLONES:
AT rich centromeric
heterochromatin
• Clone pMb3.5 (“3.5”):
present in all acrocentrics
(SSC13-SSC18); 309bp
Karyotype: Jantsch et al., 1990
12. Domestic pig Sus scrofa domestica
Centromeric
satellites
METACENTRIC CLONES:
GC rich centromeric
heterochromatin
diverse
ACROCENTRIC CLONES:
AT rich centromeric
heterochromatin
homogeneous
Bouquet at meiotic pachytene promotes clustering of
acrocentric centromeres and homogeneisation
Schwarzacher et al., 1984
13. XY
SSC1
The synaptonemal complex at meiotic pachytene
SCP1: central element protein
FISH probe for centromere of chromosome 1 SSC1
Defria and Schwarzacher 2014
Diagram: 2004
Page and Hawley
14. Acrocentric
chromosomes
cluster and
are associated
via their
repetitive DNA
sequences not
the SC itself
SSC1
Ac
Ac
Ac
Ac
Ac
Ac
SCP1: central element protein
FISH probe for centromeres of all Ac and SSC1
Alnajar and Schwarzacher 2010
15. DNA methylation
Immunostaining with anti-SCP1 (red)
and anti-methyl-5-cytosine (green) on SC spreads.
The methylation signal is amplified towards ends of the
chromosomes (yellow tips) and more methylation occurs in the
chromatin loops.
16. The Ac
chromocentre
stains strongly
with DAPI and is
not methylated
5MeC Mc1
5MeC Ac2Sheperd and Schwarzacher 2013 (unpub.)
conventionally spread
pachytene nuclei
20. Dotplot of bovine satellite I against a region
of goat chromosome 5
Dotplot of ovine satellite I repetitive unit
against a region of goat chromosome 10
24. SINE A2/tA is part of Satellite IV and
hybridizes to euchromatin and centromeric
heterochromatin
25. SINE A2/tA is part of Satellite IV and
hybridizes to euchromatin and centromeric
heterochromatin
But it is
outcompeted
when
hybridized
together with
SatIV probe
and appears on
euchromatin
only
26. Conventional and synaptonemal
complex spread of male sheep
BtSatII
Cluster of some
acrocentric
centromeres
BtSatII
Schwarzacher, Chaves, Heslop-Harrison & students 2014
27. Cattle Sat I organisation
Clone pBtKB5 is part of BtSatI and indicates
subrepeats and higher order structures
Hughes and Heslop-Harrison 2014; Chaves et al 2004 Chromosome Research
94.3%.
29. Satellite I Satellite II
Satellite III Satellite IV SINE A2/tA
A element
Cattle Sheep Cattle Sheep Cattle Sheep Cattle Sheep
Shared by
ruminants.
Density Gradient
(g/cm3)
1.715 1.714 1.723 1.723 1.706 X 1.709 X
Length (bp)
1402 820 700 700 X X 3808 X
Pairwise identity
(%)
54.7 % 61.3% X X
32. 1.715 satellite I
Divergence between cattle and sheep/goat
Less cross hybridization in FISH experiments
But strong homogeneisation within each species
acrocentric association during meiosis
33. Tandemly repeated satellite DNA in
the Artiodactyla
Pat Heslop-Harrison phh4@le.ac.uk
www.molcyt.com
Trude Schwarzacher &
Raquel Chaves
Molecular cytogenetic approaches
build a full picture of the behaviour of
chromosomes (translocations/fusions) and
satellite DNA organization and evolution
Current sequencing methods are unable to
assemble the sequences blocks or cope with
chromosomal rearrangements