Tandemly repeated
satellite DNA in the
Artiodactyla
Pat Heslop-Harrison
Trude Schwarzacher
Raquel Chaves
University of Lei...
Chromosomal changes are one of the
most important features of evolution
Bovideae: 58 autosomal arms
Cow: 2n=60 (29 pairs o...
Bos taurus taurus vs Bos taurus indicus:
2n=60, XY
But: B. taurus submetacentric Y
B. indicus acrocentric Y
Robertsonian Fusion of 1 and 29 to
give 2n=58 or 59: Gustavson 1964
Heterozygous rob(1;29) example in Portuguese
cattle Ba...
Repetitive DNA sequences –
LINE/SINE transposons and satellite
DNA – are the most abundant genome
component
- Often ‘maske...
Robertsonian Fusion
(+ )
Chaves et al.
Chromosome
Research
Complex satellite DNA reshuffing in the polymorphic t(1;29) Robertsonian translocation and
evolutionarily derivedchromosom...
Barrosa¬
Barrosa¬
Order Artiodactyla
(Even-toed ungulates)
3 groups:
1. Suiformes (pigs, peccaries,
hippopotamuses),
2. Tylopoda (c...
Domestic pig Sus scrofa domestica
Centromeric
satellites
METACENTRIC CLONES:
GC rich centromeric
heterochromatin
• Clone p...
Domestic pig Sus scrofa domestica
Centromeric
satellites
METACENTRIC CLONES:
GC rich centromeric
heterochromatin
diverse
A...
XY
SSC1
The synaptonemal complex at meiotic pachytene
SCP1: central element protein
FISH probe for centromere of chromosom...
Acrocentric
chromosomes
cluster and
are associated
via their
repetitive DNA
sequences not
the SC itself
SSC1
Ac
Ac
Ac
Ac
A...
DNA methylation
Immunostaining with anti-SCP1 (red)
and anti-methyl-5-cytosine (green) on SC spreads.
The methylation sign...
The Ac
chromocentre
stains strongly
with DAPI and is
not methylated
5MeC Mc1
5MeC Ac2Sheperd and Schwarzacher 2013 (unpub....
Sheep satellite I OaSatI
Sheep satellite I OaSatI
Hughes and Heslop-Harrison 2014
BtSatI homology in sheep
73.6%
Dotplot of bovine satellite I against a region
of goat chromosome 5
Dotplot of ovine satellite I repetitive unit
against a...
BtSatI BtSatIV
Gaspar, Hughes, Chaves and Schwarzacher 2014
FISH on
cattle
(Brakman)
chromosomes
Satellite I and II collocalize,
Satellite IV has separate arrays
BtSatII BtSatI BtSatII BtSatIV
Gaspar and Schwarzacher 20...
pBtKB
5
BtSatI-2
BtSatI-4
SINE A2/tA is part of Satellite IV and
hybridizes to euchromatin and centromeric
heterochromatin
SINE A2/tA is part of Satellite IV and
hybridizes to euchromatin and centromeric
heterochromatin
 But it is
outcompeted
w...
Conventional and synaptonemal
complex spread of male sheep
BtSatII
Cluster of some
acrocentric
centromeres
BtSatII
Schwarz...
Cattle Sat I organisation
Clone pBtKB5 is part of BtSatI and indicates
subrepeats and higher order structures
Hughes and H...
BtSatI homology between cattle
and sheep
Laetita Gaspar
Pairwise identity 54.7%
Satellite I Satellite II
Satellite III Satellite IV SINE A2/tA
A element
Cattle Sheep Cattle Sheep Cattle Sheep Cattle She...
BtSatI homology in goat
Hughes and Heslop-Harrison 2014
60.3%
1.715 satellite I
Divergence between cattle and sheep/goat
Less cross hybridization in FISH experiments
But strong homogen...
Tandemly repeated satellite DNA in
the Artiodactyla
Pat Heslop-Harrison phh4@le.ac.uk
www.molcyt.com
Trude Schwarzacher &
...
Tandem Repeats and Satellite DNA in Bovideae - Colloquium on Animal Cytogenetics
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Tandem Repeats and Satellite DNA in Bovideae - Colloquium on Animal Cytogenetics

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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

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Tandem Repeats and Satellite DNA in Bovideae - Colloquium on Animal Cytogenetics

  1. 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. 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. 3. Bos taurus taurus vs Bos taurus indicus: 2n=60, XY But: B. taurus submetacentric Y B. indicus acrocentric Y
  4. 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. 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
  6. 6. Robertsonian Fusion (+ )
  7. 7. Chaves et al. Chromosome Research
  8. 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
  9. 9. Barrosa¬
  10. 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. 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. 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. 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. 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. 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. 16. The Ac chromocentre stains strongly with DAPI and is not methylated 5MeC Mc1 5MeC Ac2Sheperd and Schwarzacher 2013 (unpub.) conventionally spread pachytene nuclei
  17. 17. Sheep satellite I OaSatI
  18. 18. Sheep satellite I OaSatI
  19. 19. Hughes and Heslop-Harrison 2014 BtSatI homology in sheep 73.6%
  20. 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
  21. 21. BtSatI BtSatIV Gaspar, Hughes, Chaves and Schwarzacher 2014 FISH on cattle (Brakman) chromosomes
  22. 22. Satellite I and II collocalize, Satellite IV has separate arrays BtSatII BtSatI BtSatII BtSatIV Gaspar and Schwarzacher 2014
  23. 23. pBtKB 5 BtSatI-2 BtSatI-4
  24. 24. SINE A2/tA is part of Satellite IV and hybridizes to euchromatin and centromeric heterochromatin
  25. 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. 26. Conventional and synaptonemal complex spread of male sheep BtSatII Cluster of some acrocentric centromeres BtSatII Schwarzacher, Chaves, Heslop-Harrison & students 2014
  27. 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%.
  28. 28. BtSatI homology between cattle and sheep Laetita Gaspar Pairwise identity 54.7%
  29. 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
  30. 30. BtSatI homology in goat Hughes and Heslop-Harrison 2014 60.3%
  31. 31. 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
  32. 32. 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

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