Poxviruses and Adaptive Genome Evolution

Aoife McLysaght

Dept. of Genetics

Trinity College Dublin

Genome Evolution

• Evolution of genome arrangement

• Evolution of genome content

– .

Genome Evolution

• Evolution of genome arrangement– Gene order changes

• Inversions, translocations

• Evolution of genome content

• .

Genome Evolution

• Evolution of genome arrangement– Gene order changes

• Inversions, translocations

• Evolution of genome content– Gene gain (sequence divergence, duplication,

recombination, horizontal transfer)– Gene loss (deletion)

• .

Genome Evolution

• Evolution of genome arrangement– Gene order changes

• Inversions, translocations

• Evolution of genome content– Gene gain (sequence divergence, duplication,

recombination, horizontal transfer)– Gene loss (deletion)

• One or more genes per event

Genome Evolution

• Translate knowledge from sequenced or model genomes to organism of interest– Positional cloning of genes– Use probes designed in one genome to detect

a target in another genome

• Improve model parameters for phylogenetic inference from genome arrangement

Genome Structure

• Not just a bag of genes

• Genome organisation contains information– Order of Hox genes corresponds to spatial

pattern of gene expression– Clustering of housekeeping genes

• By observation of ‘allowed’ changes gain understanding of genomic constraints and plasticity

Multiple Genome Comparison

• Greater power to detect change

• Precision– Can infer lineage in which change occurred

• Detect direction and rate of change

• More genomes also increase computational burden

Pox virus genomes

• 20 completely sequenced genomes

• 150-300kb containing ~200 genes

Poxviruses

• Double-stranded DNA viruses, no RNA stage

• Replicate in the host cytoplasm

• Entomopox – insect infecting• Chordopox – vertebrate infecting• Orthopox – subset of chordopox which

includes smallpox (variola) and vaccinia

Questions:

• How are these genomes arranged?

• How has genome content changed?

• Is the rate of change constant?

Questions:

• How are these genomes arranged?

• How has genome content changed?

• Is the rate of change constant?

• Can we detect adaptive genome evolution?

Orthologue detection

Significant sequence similarity– How significant?

over a long stretch of the protein– How long?

e-value threshold

Minimumaligned proportion 1 1e-5 1e-10 1e-20

0.0 0 31 29 19

0.1 0 31 29 19

0.2 4 32 29 19

0.3 7 33 31 20

0.4 10 34 31 20

0.5 17 32 30 20

0.6 29 33 30 19

0.7 28 30 26 18

0.8 26 25 22 14

0.9 15 16 14 10

1.0 0 0 0 0

• Complete linkage

• Single-link clustering

• Our method

Complete linkage

A

C

BD

E

Single-link clustering

A

C

BD

E

F

G

F

G

H

I

J

A

C

B

E

D

C

B

E

D

Orthologues

• 4042 total proteins• 3384 proteins classified into 875 groups

– 813 complete linkage

• 521 groups of 1 member• 150 groups of 2 members• 204 ≥ 3 members

Conserved gene order and spacing

Poxvirus Phylogeny

34 orthologues present in all genomes

Poxvirus Phylogeny

34 orthologues present in all genomes

Orthopox phylogeny

92 orthologues present in all orthopox genomes

Counting gene gain and loss

• Examine phylogenetic spread of a group of orthologues

• Assign gene gain and loss events to branches in the phylogeny

Phylogenomic Approach

Infer gene gain along the branch to the most recent common ancestor

Infer gene loss parsimoniously

Numbers of gain/loss events

Numbers of gain/loss events

Rate of Gene Gain

• Tested for uniform rate of gene acquisition

• Assume a molecular clock

Rate of Gene Gain

• Tested for uniform rate of gene acquisition

• Assume a molecular clock

• Are gene acquisition events distributed randomly throughout the tree?

Rate of Gene Gain

• Tested for uniform rate of gene acquisition

• Assume a molecular clock

• Are gene acquisition events distributed randomly throughout the tree?

• Simulations

Significant excess

Significant deficit

Increased Gene Gain in the Orthopox Lineage

• Slower rate of amino acid substitution within this clade (leading to abberantly short branch lengths)– Takezaki relative rate test– Branch lengths from synonymous distances

Increased Gene Gain in the Orthopox Lineage

• Slower rate of amino acid substitution within this clade (leading to abberantly short branch lengths)– Takezaki relative rate test– Branch lengths from synonymous distances

• Increased rate of gene gain

• Increased selection for the retention of gained genes

Sources of Gene Acquisition

• Extensive sequence divergence

• Recombination

• Horizontal transfer

Horizontal Transfer

• AMV-EPB_034 – inhibitor of apoptosis from Amsacta moorei entomopoxvirus (AMV-EPB)

• GenBank sequence – inhibitor of apoptosis from Bombyx mori (silkworm) BLAST e-value 9e-81

• Amsacta moorei entomopoxvirus infects Amsacta moorei (Red Hairy Caterpillar)

• Bombyx and Amsacta both Order Lepidoptera

Horizontal Transfer

• AMV-EPB_034 – inhibitor of apoptosis from Amsacta moorei entomopoxvirus (AMV-EPB)

• GenBank sequence – inhibitor of apoptosis from Bombyx mori (silkworm) BLAST e-value 9e-81

• Amsacta moorei entomopoxvirus infects Amsacta moorei (Red Hairy Caterpillar)

• Bombyx and Amsacta both Order Lepidoptera • 62% of best non-viral GenBank hits are from

same taxonomic Class as viral host

Gene loss modelling

• Events are not independent

• Depend on previous (in time) gain and loss events of the gene family

• Requires a probabilistic model?

Gene loss events

Adaptive Evolution

• Selection for diversification– Positive selection

• Characteristic of host-parasite co-evolution

Standard Genetic CodePhe UUU Ser UCU Tyr UAU Cys UGU

UUC UCC UAC UGC

Leu UUA UCA ter UAA ter UGA

UUG UCG ter UAG Trp UGG

Leu CUU Pro CCU His CAU Arg CGU

CUC CCC CAC CGC

CUA CCA Gln CAA CGA

CUG CCG CAG CGG

Ile AUU Thr ACU Asn AAU Ser AGU

AUC ACC AAC AGC

AUA ACA Lys AAA Arg AGA

Met AUG ACG AAG AGG

Val GUU Ala GCU Asp GAU Gly GGU

GUC GCC GAC GGC

GUA GCA Glu GAA GGA

GUG GCG GAG GGG

Detection of Positive Selection

• Two classes of DNA substitutions– Synonymous (DNA change without amino acid

change)– Nonsynonymous (DNA change causing amino acid

change)• Neutral – equal frequencies• Conservative selection – fewer

nonsynonymous substitutions• Positive selection – more nonsynonymous

substitutions

Detection of Positive Selection

• Two classes of DNA substitutions– Synonymous (DNA change without amino acid

change)– Nonsynonymous (DNA change causing amino acid

change)• Neutral – equal frequencies• Conservative selection – fewer

nonsynonymous substitutions• Positive selection – more nonsynonymous

substitutions

Detection of Positive Selection

• Two classes of DNA substitutions– Synonymous (DNA change without amino acid

change)– Nonsynonymous (DNA change causing amino acid

change)• Neutral – equal frequencies• Conservative selection – fewer

nonsynonymous substitutions• Positive selection – more nonsynonymous

substitutions

Detection of Positive Selection

• Two classes of DNA substitutions– Synonymous (DNA change without amino acid

change)– Nonsynonymous (DNA change causing amino acid

change)• Neutral – equal frequencies• Conservative selection – fewer

nonsynonymous substitutions• Positive selection – more nonsynonymous

substitutions

Detection of Positive Selection

• 204 groups of orthologues

• Maximum liklihood test for positive selection (PAML)

• Significantly higher frequency of nonsynonymous substitutions

Positive Selection on Pox Genes

• Detected positive selection on 26 genes

• Examples:– Membrane glycoprotein– Haemagluttinin– Immunoglobulin domain protein

Positive Selection on Pox Genes

• 13 genes are unique to orthopox clade– Significantly more than expected (P < 0.05)

• Disproportionate frequency of positive selection on genes gained within the orthopox lineage

Adaptive Genome Evolution?

• Association of positive selection on protein sequences and increased rate of gene acquisition

Adaptive Genome Evolution?

• Association of positive selection on protein sequences and increased rate of gene acquisition

• Adaptive significance of gene acquisition?– Mimic host defences– Avoid host recognition– Block cell death

Conclusions

• The rate of genome evolution is not constant

• The rate of gene acquisition has increased in the orthopox lineage

• Orthopox lineage is also has an increased frequency of positive selection

• Possible adaptive significance of genome evolution

Acknowledgements

• University of California, Irvine– Brandon Gaut– Pierre Baldi