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Gilles Fischer
Comparative genomics
GENOME EVOLUTION:
DNA duplications
Chromosome dynamics
Nucleotide composition
A brief introduction to the field of Comparative Genomics
DNA carries the genetic information: Avery (1943) and Hershey-Chase (1952)
"Il ne fait aucun doute que l'tude systmatique de la teneur absolue du noyau en
acide dsoxyribonuclique, travers de nombreuses espces animales puisse
fournir des suggestions intressantes en ce qui concerne le problme de
l'volution"
Jacques Monod:
"Tout ce qui est vrai pour le colibacille est vrai pour l'lphant"
A brief introduction to the field of Comparative Genomics
time
or
quantity of evolutionary changes
time
or
quantity of evolutionary changes
Bio-informatics
Experimental Biology
Genetic screens
Molecular Mechanistic
mechanisms hypotheses
functional genomics
A brief introduction to the field of Comparative Genomics
Bio-informatics
SMALL GENOMES
AND
EXPERIMENTALLY TRACTABLE
Experimental Biology
Genetic screens
Molecular Mechanistic
mechanisms hypotheses
functional genomics
A brief introduction to the field of Yeast Genomics
Saccharomyces cerevisiae used in baking and fermenting alcoholic beverages for thousands of
years
Other species of yeast, such as Candida albicans, are opportunistic human pathogens
Yeasts have recently been used to generate electricity in microbial fuel cells and produce
ethanol for the biofuel industry.
Schizosaccharomyces pombe
Saccharomycotina
A brief introduction to the field of Yeast Genomics
Saccharomyces paradoxus
Saccharomyces mikatae
Saccharomyces cerevisiae
Saccharomyces kudriavzevii
Whole Genome Duplication Saccharomyces bayanus
Saccharomyces pastorianus
Saccharomyces exiguus
Saccharomyces servazzii
Gain of Megasatellites Saccharomyces castellii
Candida glabrata
Vanderwaltozyma polyspora
Zygosaccharomyces rouxii
Gain of HO gene Lachancea thermotolerans
Lachancea waltii
Lachancea kluyveri
Kluyveromyces lactis
Gain of mating type cassettes Kluyveromyces marxianus
and small centromeres Eremothecium gossypii
Saccharomycodes ludwigii
Brettanomyces bruxellensis
Pichia angusta
frequent tandem duplications Candida lusitaniae
Debaryomyces hansenii
Pichia stipitis
Pichia sorbitophila
Candida guilliermondii
Extensive loss of transposable Candida tropicalis
elements and spliceosomal Candida parapsilosis
introns Lodderomyces elongisporus
Candida albicans
Candida dubliniensis
Arxula adeninivorans
Yarrowia lipolytica
Schizosaccharomyces pombe
A brief introduction to the field of Yeast Genomics Genome annotation
# chr size (Mb) # genes # tRNA # introns
100 *
100 MYr
Candida glabrata 65
Homo sapiens
Zygosaccharomyces rouxii -
450 MYr
100 - 300 MYr
90
550 MYr
300 - 1000 MYr
Lachancea thermotolerans -
70
Takifugu rubripes
Kluyveromyces lactis 60
Tetraodon negroviridis
Debaryomyces hansenii 51
1.25
Candida glabrata
1.20
1.15
Zygosaccharomyces rouxii
1.10
Lachancea kluyveri
(WashU seq center M. Jonhston)
Lachancea thermotolerans
- important level of redundancy (in all
eukaryotic phyla)
Kluyveromyces lactis
- Gene order changes (differential loss of
duplicates, translocation breakpoints)
Debaryomyces hansenii
S. c. A. t. C. e. D. m. H. s. s.
duplication
Duplicated Genes 43% 65% 49% 40% 50%
Degeneration
Pseudogenization Neofunctionalization Conservation Complementation
Loss of function Gain of a new Gene dosage increase Specialization of
(most frequent fate) function Genetic robustness the 2 copies
CGH
A duplication assay:
XV
RPL20B and so on
XIII
???
XV
RPL20B RPL20B
XIII
rpl20A
dltion
==>slow growth ==> WT growth rate
Genome evolution: DNA duplications A duplication assay:
IV - XII
XV RPL20B
143 kb
VII, XV
II
XIV
X
XI direct tandem
V - VIII
III
VI
I
A A C C T A G A G C T T ( G T T ) 14 G T G G A T T G T T T
Despite the selection of a single gene duplication event, only large segmental duplications were recovered
Genome evolution: DNA duplications Molecular mechanisms:
WT 10-7 (1) 42 6 48 52
REPLICATION
pol32
time (min)
Raghuraman
0 et al. Science, 2001
(<0.07) T -T T T T - T - -
0
microsatellites
25 rad52 3 x 10-7 (3) 70 1 0 100
30 a connection with
35 rad52 replication?
rad1 8 x 10-8 (0.8) 15 0 0 100
40
dnl4
Koszul et al. EMBO J., 2004
Replication-based mechanisms
strain rate of SDs type of SDs breakpoint sequences (%)
(/cell/division)
Intra-chromosomal Inter-chromosomal LTRs microhomologies
microsatellites
WT 10-7 (1) 42 6 48 52
pol32
Bloom and Cross, 2007 0 (<0.07) - - - -
Nick McElhinny, Cell 2008
Pol32
Pol32 is required for initiating BIR reaction (Lydeard et al, 2007)
WT 10-7 (1) 42 6 48 52
CPT
Top1 Top1
=>broken forks promote
SD formation
Dnl4
NHEJ Resection
HR
pas dhomologies, Rad52 Rad1
religature simple
Rad51
Pol32 MMEJ
SSA BIR
Microhomologies (5-12pb)
>30pb dhomologies
SDSA DSBR
Two different replication-based mechanisms
WT 10-7 (1) 42 6 48 52
====>
=>
HR-dependent
HR-independent
X
Dnl4
Resection
X
Rad52 X
Rad1
MMIR: microhomology microsatellite-induced replication
WT 10-7 (1) 42 6 48 52
HR requires Rad52
MMEJ requires Rad1 SD are still being formed in the absence of all known DSB repair pathways
NHEJ requires Dnl4
existence of a new DSB repair pathway?
X
Dnl4
Resection
X
Rad52 X
Rad1
The DSB repair pathways
X
Dnl4
Resection
X
Rad52 X
Rad1
A new pathway?
MMIR
Microhomology/microsatellites Induced Replication
- independent from all known DSB repair pathways (HR, NHEJ, MMEJ)
- dependent from Pol32
- Replication template switching between microhomologies and microsatellites
Genome evolution: DNA duplications Conclusions
SDs are spontaneously generated at high frequency: 10-7 SD/cell/division for the RPL20B locus
SDs arise from two alternative replication-based mechanisms: BIR and MMIR
MMIR represents a new mechanism different from known DSB repair pathways (HR, NHEJ):
between microhomologie (between 2 to 11 nt) and microsatellites (poly A/T, trinucleotide
repeats)
independent from Rad52
requires Pol32
Species 1
translocations
Inversions
Species 2 duplications
# deletions
# x
rates of rearrangements
Genome evolution: Chromosome Dynamics
Sensu stricto
S. cerevisiae
S. serevisiae
S. cariocanus S. bayanus
S. paradoxus
Candida glabrata
S. mikatae
S. kudriavzevii
Zygosaccharomyces rouxii
S. bayanus
Lachancea kluyveri
Saccharomyces sensu stricto complex:
- monophyletic group
- very closely related species Lachancea thermotolerans
- hybrids viable but sterile
- 16 chromosomes
Kluyveromyces lactis
Debaryomyces hansenii
Yarrowia lipolytica
Genome evolution: Chromosome Dynamics
S. cerevisiae
only few translocations:
S. cariocanus (4)
low reorganization
S. paradoxus (0) recombination between repeated sequences
S. mikatae (2) no chromosomal speciation
variable rate of rearrangements?
S. kudriavzevii (0)
S. bayanus (4)
Candida glabrata
chr VIII
Zygosaccharomyces rouxii
Lachancea kluyveri
Lachancea thermotolerans
Kluyveromyces lactis
Debaryomyces hansenii
Yarrowia lipolytica
chr VIII
chr VIII
Fischer
F. Brunet
chr VIII
chr VIII
- comprehensive reshuffling
C. glabrata
Candida glabrata
- 509 translocations, 104 inversions
- no homologous chromosomes
Zygosaccharomyces rouxii
"UNSTABLE" GENOMES
Lachancea kluyveri
S.cerevisiae
Lachancea thermotolerans
-moderate reshuffling
"STABLE" GENOMES
L. kluyveri
Genome evolution: Chromosome Dynamics
Quantitative estimation of the relative genome stability: GOC (gene order conservation)
species 1
=5
# neighboring orthologues
If yes: +1
? If no: 0
GOC =
Total # orthologues
species 2
=5
GOL (
- Rate of rearrangements = ( Dist phylogntique mean rate
2.7 0.7
1.3
0.4 Candida glabrata
D. hansenii
0.6 0.6
Zygosaccharomyces rouxii
1.7
S. cerevisiae
0.3
Lachancea kluyveri C. glabrata 0.5
(WashU seq center M. Jonhston)
0.3
1.7
Debaryomyces hansenii
1.7
Yarrowia lipolytica
Fischer et al. , PLoS Genet 2006
Genome evolution: Chromosome Dynamics
moderate
massive low
Sensu stricto
S. serevisiae differential gene loss
S. bayanus
Zygosaccharomyces rouxii
Lachancea kluyveri
(WashU seq center M. Jonhston)
Lachancea thermotolerans
Stable genomes
Kluyveromyces lactis
TGA expansion
Debaryomyces hansenii
No synteny
Y. lipolytica
Genome evolution: Chromosome Dynamics Conclusions
Highly variable rates of chromosome rearrangements between lineages but also within a given
lineage
few examples of the adaptative role of rearrangements (proliferation of cancer cells (ONeil
and Look, 2007), growth advantage of translocated yeast cells (Colson et al, 2004),
adaptative gene loss (Domergue, 2005).
QuickTime et un
Candida glabrata 38.8 dcompresseur
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41.5
G T transversions : 8-oxo-guanine
Lachancea kluyveri
Shibutani et al., Nature, 1991
Lachancea thermotolerans 47.3
Global AT-enrichment
Kluyveromyces lactis 38.8
60
47.3
40
39.1
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dcompresseur
sont requis pour visionner cette image.
20
1 2 3 4 5 6 7 8 9 10 Mb
A B C D E F G
Zygosaccharomyces rouxii
60 C-left
52.9
40
41.5
20
1 2 3 4 5 6 7 8 9 10 11 Mb
A B C D E F G H
Genome evolution: Nucleotide composition
DNA
RNA 1st 2nd 3rd 1st 2nd 3rd 1st 2nd 3rd AAAAAA
Protein A G P R I N K F
1.3 1.2 1.1 1.2 0.7 0.8 0.9 0.9 relative use in C-left
S. cerevisiae
100
Alignments of universally conserved proteins :
100
100 L. kluyveri
100 0.05
96
100 L. waltii
100
100
L. thermotolerans
K. lactis
100
98
E. gossypii
C-left has the same phylogentic origin than the rest of the genome
670 kb LAKL_C
200bp fragments
G1 DNACy3
S DNACy5
G2
Genome evolution: Nucleotide composition Replication:
ChrA
ChrB
Genome evolution: Nucleotide composition Replication:
ChrC
ChrD
Genome evolution: Nucleotide composition Conclusions
presents a very high level of synteny conservation with sister species genomes
encompasses the MAT locus but has lost the silent cassettes HMR and HML
- Gnolevures consortium:
Jean-Luc Souciet Univ. Louis Pasteur, Strasbourg