PhD defense - Benoit Madec (eq. Espagne - Genome Biology dpt)

Title: Impact of polymorphism on meiotic crossover distribution in Arabidopsis thaliana.

Abstract: During meiosis, crossovers are formed between

homologous chromosomes, which leads to the

reassortment of parental alleles, maintaining and

generating genetic diversity in the offspring.

Meiotic crossover distribution along

chromosomes is neither uniform nor random but

the mechanisms and evolutionary forces that

impose crossover patterning are poorly

understood. While detection of polymorphism is

absolutely essential to prevent illegitimate

recombination and avoid chromosome

rearrangements in hybrids, only limited

information is available on the interplay between

sequence polymorphism and crossover

patterning along chromosomes. Interestingly,

historical levels of polymorphism, present

naturally in different Arabidopsis thaliana

ecotypes, do not impact crossover distribution,

suggesting that other chromosomal features are

driving crossover distribution. However, two

effects of polymorphism on recombination and

crossover formation have been demonstrated in

A. thaliana. A first effect, at the local scale,

showed that recombination and crossovers tend

to avoid polymorphism. The same effect has

been demonstrated in both mitotic and meiotic

cells, in bacteria and the yeast Saccharomyces

cerevisiae. However, at a global scale, studies in

genetic backgrounds where heterozygous

regions are juxtaposed to homozygous regions

suggest that class I crossovers could

preferentially form in heterozygous regions. To

specifically test the effect of polymorphism on

crossover distribution in a genome wide manner,

independent of other chromosomal features, I

used available A. thaliana Recombinant Inbred

Lines (RILs). RILs genomes are a patchwork of

two parental genomes, backcrossing the RIL to

either parent produces F1 individuals where each

chromosome contains heterozygous and

homozygous regions, spread out throughout the

genome. Using the backcrossed RILs, I was able

to probe each locus in a polymorphic or

non-polymorphic state, and test whether

crossovers form preferentially in polymorphic

regions or not.

I established bioinformatic tools to detect

crossovers using two levels of divergence: low

and high. The high level of divergence (1 SNP

every 250bp) is present in heterozygous

regions and comes from the natural divergence

between two ecotypes of A. thaliana:

Columbia-0 and Catania-1. The low level of

divergence (1 SNP every 200,000kb) is present

in homozygous regions and comes from an

EMS treatment. I used F1 hybrids between

Col-0 and Ct-1 EMS treated parents to assess

that both levels of divergence allow for the

precise detection of crossover events. I

showed that polymorphism can shape

crossover distribution extensively throughout

the genome and compete with genomic

features of the recombination landscape,

altering them in both of either sex of A.

thaliana. Polymorphism was able to delocalize

crossovers from very stable crossover rich

regions to very stable crossover poor regions.

My data also supports that, potentially through

the interplay with interference, polymorphism

could even increase recombination in both

heterozygous and homozygous regions of the

same chromosome. I have also demonstrated

an homogeneous effect of polymorphism along

heterozygous, polymorphic intervals. Based on

previous data, this effect could rely on MSH2,

for which I am currently analyzing the data in

CRISPR/Cas9 knock-out mutants. These data

will be presented during my defense. My results

altogether support a major role of

polymorphism as a driver of recombination and

crossover formation in A. thaliana.

Polymorphism is a strong tool to study ancient

questions such as what underlies the

mechanisms of heterochiasmy and

interference. In my manuscrit, I discuss the

implications of my work in the context of the

different rules that regulate the global

recombination landscape in A. thaliana and

beyond.