IInversions as barriers to recombination and

facilitators of speciation: A play in 4 actsMohamed A. F. Noor @mafnoor

Katharine KorunesDuke University

SSpeciation, then what?

• Speciation generates biodiversity on our planet

• … but what maintains species once they form?• MANY “good species” still hybridize• MANY “good species” have incomplete RI• MANY “good species” exhibit genetic evidence of

interspecies gene flow • “Leaky” rather than absolute

SSpecies are genotypic clusters in LD

• Reproductive isolation maintains linkage disequilibrium by stopping exchange between types

• Cause/ effect

• If there is no LD anywhere in the genome, unlikely would consider two types to be “distinct species”

SSpecies are genotypic clusters in LD

• Reproductive isolation maintains linkage disequilibrium by stopping exchange between types

• Cause/ effect

• If there is no LD anywhere in the genome, unlikely would consider two types to be “distinct species”

IIf having LD important, then stopping recombination helps

Crossing over is the exchange of genetic material between

homologous chromosomes

(Wikipedia 29 July 2015)

T. H. Morgan (1916)

IIf having LD important, then stopping recombination helps

Crossing over is the exchange of genetic material between

homologous chromosomes

(Wikipedia 29 July 2015)

RECOMBINATION

LD lost!

IIf LD is important, then stopping recombination helps

Crossing over is the exchange of genetic material between

homologous chromosomes

(Wikipedia 29 July 2015)

NO RECOMBINATION

LD (and genotypic clusters) retained!

UUltimate question motivating research in

this talk:

Does creating LD in ways besides RI facilitate

species persistence?

AAct 1:Genetics of speciation

and origin of a hypothesisNoor, Machado, Hey, Kliman, Markert, Grams,

Bertucci, Reiland, Williams

IIntegrated study of speciation in Drosophila pseudoobscura/ D. persimilis

Noor lab• Genetically map all

barriers to gene flow between species

Jody Hey/ Carlos Machado• Survey marker regions

for interspecies introgression

Assumed association, as suggested by decadesof hybrid zone research

• Phenotypically identical sibling species• Estimated divergence ~500,000 – 1M yrs• Hybridize in nature (1/10,000)• Strong prezygotic isolation

• Hybrid male sterility & some inviability• Native to North America and co-occur in

some areas

DDrosophila pseudoobscuraD. persimilis

GGeographic range and overlap

Both speciesD. pseudoobscura

Outgroup speciesD. miranda also in range of sympatry

CChromosomal arrangements• These species differ by

many chromosome “inversions”.

• *Inversions prevent single crossovers in those regions in F1 hybrid (heterozygote).*

X

2 3 4 5

GGenetic mapping studiesoof potential barrier traits

• Hybrid male sterility : N = 2500• Hybrid inviability : N = 2500• Hybrid courtship deficiency: N = 2500• Mating discrimination by females : N = 1100• Courtship song differences : N = 500• Cuticular hydrocarbon differences : N = 234

(Noor & Coyne 1996 Gen Res) Noor et al 2001 PNASNoor et al 2001 Evolution Williams et al 2001 Heredity & others

GGenetic basis of aall ddifferences:MMap exclusively to inversion differences

• STRONG effects detected of inversions on XL and 2nd

chromosome• Some effect detected of

inversion on XR chromosome

• No detectable effect with markers outside inversions• Inversions fully explained

variation in traits

XL

2 3 4 5

******

*

Noor et al 2001 PNAS

XR

CContrast to other studies...

• Allopatric species D. simulans / D. mauritiana• Sterility maps to many regions

• Geographic races of D. melanogaster• Mate preference maps to many regions

• Why traits map just to few (inverted) regions in D. pseudoobscura/ D. persimilis?• Sympatry/ hybridization homogenizing?

CConsistent (fixed) differences:IIn & near inverted regions

• Regions inside or near inversions tended to be ones with fixed differences• Only one fixed difference site found

>3Mb from inversion (chromosome 4)

• Extensive allele sharing in regions far away from inversions

• “Genes … located in genomic regions not associated with isolation phenotypes show more evidence of introgression…”

XL

2 3 4 5

***

***

*

XR

Machado et al 2002 MBE

***

***

***

*

HHypothesis:

D. pseudoobscura and D. persimilis persist as distinct gene pools (species) because of differences maintained in inverted regions

Noor et al 2001 PNAS

Recombinationhomogenized

uninvertedareas

Ancestors

MMost closely related Drosophila species NOT ddiffering by inversions were allopatric…

Noor et al 2001 PNAS

AAct 2:Genome-enabled data &

antirecombinational role of inversions maintaining

speciesNoor, Machado, McGaugh,

Stevison (JMS prize winner!), Garfield, Haselkorn, Schaeffer, Chang, Kulathinal

TTargeted sequencing approach: chromosome 2

Machado, Haselkorn, and Noor 2007 Genetics

PPublic genome sequences

• More sequence difference on average within inverted region in both tests

.01

.02

.03

.04

.05

.06

MedianKs

0 5 10 15 20 25 30Position

CHROMOSOME 4

.01

.02

.03

.04

.05

.06

MedianKs

0 5 10 15 20 25 30 35Position

CHROMOSOME 2

Noor, Garfield, Schaeffer, & Machado 2007 Genetics

inversion no inversion

RRecombinational role???Does it have to do with recombination

& homogenization, or are the inverted regions “special” otherwise?

CCrossovers not recovered within inversion, or just outside

• Crossovers suppressed just outside likely because of chromosome pairing difficulties

RRecombination in Hybrids

• Complete suppression of recombination products until ~2.5 megabases outside inversion

•Within species, 2.5 megabases would experience >10% recombination

Stevison et al 2011 GBE

TTargeted sequencing approach: chromosome 2

Effect associated w/ suppressed crossover

BBetter controls…

• Does the low divergence far outside the inversions result from interbreeding?

• Want to look at patterns if not hybridizing• D. pseudoobscura – D.miranda

(expect uniformly high divergence)

• Want to look at patterns if freely interbreeding• D. pseudoobscura – D. pseudoobscura• D. persimilis – D. persimilis

(expect uniformly low divergence)

mir ps perSuzanne McGaugh

EExperimental pair (ps-per)

Suzanne McGaugh

NNot hybridizing pair (ps-mir)

Suzanne McGaugh

FFreely hybridizing pair 1 (ps)

Suzanne McGaugh

FFreely hybridizing pair 2 (per)

Suzanne McGaugh

AAll together mir ps per

Suzanne McGaugh McGaugh & Noor 2012 PRSLB

SSympatry/ allopatry comparisons• Hypothesis: Gene flow

between D. pseudoobscura and D. persimilis homogenized regions outside the inversions, but effect should be strongest in sympatric populations• Genetic mapping data• DNA sequence data

HHybrid sterility mappingCChromosomes 2 and 4

Sympatric pair Allopatric pair

* *

**

*

Differencestatisticallysignificant

Chang & Noor 2007 GeneticsChang & Noor 2010 Evolution

2 4 2 4

DDNA sequence comparisonCChromosome 2

• Polarized “derived” bases and looked whether more sharing between D. persimilis and sympatric vs. allopatric D. pseudoobscurain single genome sequences

Outside inversions• 219 shared sym ps – per• 185 shared allo ps – per• Binomial sign test, p = 0.05

Kulathinal, Stevison, & Noor 2009 PLoS Genetics

SSummary of Act 2

• High divergence in inverted regions, lots of allele sharing in collinear regions• High divergence corresponds specifically with areas

having no/ low recombination in hybrids• Evidence for greater allele sharing and

homogenization in sympatric populations than in allopatric population in collinear regions

• Inversions facilitate species persistence in this system through their recombinational effects!

AAct 3:Lies, damn lies, and

antirecombinational effects of chromosomal inversions

Feder, Navarro, Schaeffer, Anderson, Nosil, Guerrero, Kirkpatrick

(no Noor!)

MModes of gene “flux”between inverted regions…

Double crossover (DCO) Gene conversion (GC)

Typically <2000bp(Drosophila chromosome is ~30,000,000bp)

Can happen in inversion heterozygotes

MModes of gene “flux”between inverted regions…

Double crossover (DCO) Gene conversion (GC)

From Singh 2012 doi: 10.1371/journal.pgen.1003024

GGene flux expectations

• Constant rate of flux of ~10-5 /site/gen throughout inverted region from gene conversion• Schaeffer & Anderson

(2005) report 3 x 10-6

• Higher rate of flux in center resulting from double-crossovers Navarro et al 1997 Genetics

SSimulation studies qquestion effectiveness

of inversions…

• Over long time, low exchange often erodes sequence differences within inverted regions

• Figure is oneof the “best”case-scenarios

… but we don’t see any erosion betweenD. pseudoobscura and D. persimilis withininverted region???

Feder & Nosil 2009 Evolution; Feder et al 2011 Evolution

CCoalescent model showsmany more considerations

• Old vs. new inversion?• Locally adapted breakpoints vs.

locally adapted alleles contained within?

• Not so unlikely to see differentiation maintained for a very long time, even with gene flux

• Specific predictions depend on rate of flux, number and distribution of locally adapted loci, etc.

Guerrero et al 2012 Phil Trans B

AAct 4:Reconciling predictions with

data: rates of gene flux in inverted regions

(***in progress***)

Noor, Stevison (JMS prize winner!), Korunes

EExamined DCO rate empirically

• Crossing scheme:

Double crossover

EExamined DCO rate empirically

• Crossing scheme:• Looked at largest

inversion (~12.5Mb)• Screened 9739 progeny

individually using markers in middle & edges of chromosomal inversion• One confirmed double-

crossover ~10-4

• ~12% expected for same stretch without inversion Double crossover

Stevison et al 2011 GBE

D. persimilis

X

X

D. pseudoobscura

F1 D. pseudoobscura

Whole-genome sequencing

D. persimilis X D. pseudoobscura

10 offspring

gene c0nversion

Examined GC rate empirically

IN PROGRESS

Pure species 1

Pure species 2

F2 offspring

How gene conversion looks

Gene conversion: results thus far

Katharine Korunes, unpublished

PPutting it all together

• Detectable rates of flux in inversion heterozygotes• ~10-4 from GC, another ~10-4 in middle from DCO

• BUT hybridization rate is also ~10-4

• Cumulative effect is on the order of what was used in theoretical models• YET observe consistently high divergence in

inverted regions

Suggests “locally adapted” alleles also contribute to differentiation within inverted regions, not JUST recombinational inhibition

PPUNCHLINES

• Reducing recombination via inversions DOES facilitate species persistence• Clear evidence in Drosophila pseudoobscura/

D. persimilis• Varying evidence in other systems too

• BUT, inversions imperfect barriers• Leakage happens• Neutral differences preserved despite some gene flux• Likely locally adapted alleles inside make inversions more

effective barriers

• Creating LD in ways besides RI facilitates speciation

AcknowledgementsNoor lab (past & present)Duke University

Carlos Machado

ESEB, Rui Faria, Anja Westram, Juan Galindo, Mark Ravinet& all of you for listening!

Yourpicture

here

NNoor laboratory

2009:

2015:

Slides to bedeposited in

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