Download Genome Shock in Polyploid Plants

Genome Shock in Polyploid Plants
Matt Hegarty
The importance of polyploidy
• It is predicted that ~70% of higher plants have undergone at
least one round of genome duplication over their evolutionary
history.
• Approximately 2-4% of speciation events involve a change in
ploidy.
• It is estimated that the majority of polyploid species are the
result of interspecific hybridisation.
• Polyploids represent a source of genetic novelty upon which
selection may act.
• Many of the world's most successful crop species are polyploids
(bread wheat, coffee, cotton, sugarcane, maize) and often
significantly outperform their diploid relatives.
Hybrid speciation in Senecio: the
allopolyploid origin of S. cambrensis
X
Senecio squalidus
Diploid, SI
Known to have
occurred at least
TWICE
Senecio vulgaris
Tetraploid, SC
Senecio x baxteri
Triploid, sterile
Chromosome doubling
Senecio cambrensis
Hexaploid, fertile, SC
What are the genetic consequences of polyploidy?
• Genetic changes as a consequence of uniting 2+ divergent genomes
within a single hybrid nucleus:
1) new combinations of gene regulatory networks.
Arabidopsis
thaliana
2n=2x=10
X
A. arenosa
2n=4x=32
Unreduced
gamete
A. suecica
2n=4x=26
FRI
FLC
FRI
FLC
A. thaliana
A. arenosa
Wang et al. (2006) Genetics 173: 965-974
2) Recombination: rapid recombination is often seen in hybrids
- serves to resolve chromosomal incompatibilities in homoploids
and reinforce reproductive isolation.
- in allopolyploids, helps prevent mispairing of homoeologous
chromosomes.
Chromosome reorganisation: sequence loss
• Allopolyploid genome sizes are often less than just the sum
of their parts - the observed C values don‟t match predictions!
• Studies in resynthesised wheat polyploids showed that, almost
immediately, the hybrids lose certain DNA sequences:
GSS are lost in the initial
F1 hybrid.
CSS are lost immediately
after genome doubling.
Sequence loss is reproducible
and can involve up to 15% of
genomic DNA!
Also see rapid loss of noncoding, repetitive DNA.
Feldman et al. (1997) Genetics 147: 1381.
Transposon activation
• McClintock predicted widespread activation of transposable elements
as a consequence of the “genomic shock” of hybridisation.
• Transposon activity would facilitate rapid recombination by increasing
the likelihood of chromosome breakage.
• Also has the potential to alter gene expression.
S. aethnensis
S. chrys.
S. squalidus
S. aethnensis
S. chrys.
S. squalidus
Transposon activation
• Followup assays using quantitative PCR to assess copy number of Ty1
and Ty3 retroelements in resynthesised S. squalidus hybrid lines.
• Preliminary data seems to indicate an increase in copy number in
initial hybrids but a slow decrease after segregation.
700
Amount of template (pg)
600
500
400
Ty1
Ty3
300
200
100
0
F1
F2
F3
Generation
F4
F5
Transposon activation
• Followup assays using quantitative PCR to assess copy number of Ty1
and Ty3 retroelements in resynthesised S. squalidus hybrid lines.
• Preliminary data seems to indicate an increase in copy number in
initial hybrids but a slow decrease after segregation.
4.7
4.6
600
4.5
4.4
500
4.3
400
4.2
300
4.1
4
200
3.9
3.8
100
3.7
0
3.6
F1
F2
F3
Generation
F4
F5
C-value
Amount of template (pg)
700
Ty1
Ty3
C-value
Changes to gene expression in the allopolyploid
hybrids Senecio x baxteri and S. cambrensis
10
1
0.1
SS
SB
SC
SV
Does genome duplication „rein in‟ non-additive expression?
Hegarty et al. (2005) Mol. Ecol. 14: 2493-2510.
Hegarty et al. (2008) Phil. Trans. Roy. Soc. 363: 3055-3069.
Genome duplication has a distinct, secondary
effect on gene expression in allopolyploids
58%
17%
Novel expression in synthetics
SB
S1
S2
S3
S4
S5
SC
SB
S1
S2
S3
S4
S5
SC
Hegarty et al. (2006) Current Biology 16: 1652-1659.
Epigenetic regulation of gene expression
• Studies in allopolyploid systems show rapid changes to DNA
methylation and histone acetylation relative to parental taxa.
- Early work in Arabidopsis using cDNA-AFLP showed silencing
of a number of genes in the allotetraploid hybrid.
- This was tested further using MSAP and 8.3% of loci showed
differential methylation between the parents and an F3 hybrid.
• Similar findings have been made in systems such as Spartina (up to
30% of loci affected!) and wheat (13%).
• These changes may serve to prevent expression of duplicated gene
copies where this would be counterproductive.
• In an extreme example, the genes affected in A. suecica hybrids
are the rRNA genes of the A. thaliana parent, causing almost total
suppression of A. thaliana gene expression - nucleolar dominance.
• Very little information on the role of miRNA/siRNAs in hybrids...
Investigating DNA methylation in Senecio using MSAP
• MSAP = modified AFLP using EcoRI and either HpaII or MspI.
• If a locus is methylated will see a shift in AFLP profile.
HpaII
In this example the peak in the
MspI sample but not HpaII
indicates methylation at the CCGG
recognition sequence of this locus.
MspI
Currently analysing data from an
analysis of three synthetic lines
of S. x baxteri and two generations
of their allohexaploid derivatives
plus the parental taxa.
Subfunctionalisation - a basis for heterosis?
• Studies in allotetraploid cotton (and recently wheat) showed that
gene copies from different parents had different expression
patterns.
• What they observed was organspecific reciprocal silencing of
one parental gene copy.
• When assessed in new hybrids,
they found that these patterns
were replicated.
A genome
copy
D genome
copy
• More importantly, they observed
that relative expression of the
parental gene copies could vary based on environmental stress.
• This could allow polyploids to „select‟ the best gene copy to express
in a given environment - maybe why polyploids are so successful?
cDNA-SSCP
PCR is used to amplify a small
segment of the gene of interest
containing a SNP that can be used
to identify parental origin.
These secondary structures
migrate at different rates, so
sequence variants can be
identified by differences in
banding patterns.
S. vulgaris
S. squalidus
…CACCG…GGTTC…
...CAGCG…GGGTC…
hfr2 - Hessian Fly Resistance gene
gDNA
cDNA
SV
SS
AC
Bax
SV
SS
AC
Bax
• Upregulated in S. squalidus relative to all other taxa according to
array data.
• S. vulgaris specific band, while inherited in the hybrids, is silenced.
• SSCP hybrid expression pattern favours S. squalidus.
Future work
• Next-generation sequencing in Senecio - funded for a single run
using CRoPS to identify polymorphisms between allopolyploids and
parents. Also awaiting decision on NERC proposal.
• Other allopolyploid systems - festulolium, white clover.
Festulolium = grass hybrid of
Lolium perenne X Festuca pratensis
IBERS have a number of hybrid/
introgression lines
Genetics well-characterised
Natural hybrids exist, may have
improved flood tolerance
Images from Humphreys & Harper (2008)
Crop Wild Relative 6:1-3.
Synteny with other grasses
Acknowledgements
University of Bristol:
University of St. Andrews:
Simon Hiscock (PI)
Richard Abbott (Co-I)
Keith Edwards (Co-I)
Adrian Brennan (Postdoc)
Gary Barker (Bioinformatics)
Jane Coghill (Transcriptomics)
Chris Thorogood & Sacha Allen
(Ph.D students)
Tom Batstone (Technician)
Aberystwyth University:
Wayne Powell (Director)
Charly Morgan (Technician)
Research Funding:
NERC
Linnean Society