Tuesday, December 18, 2012

Genome-wide analysis of a long-term evolution experiment with Drosophila

ResearchBlogging.orgFor decades, most researchers have provided some general insights into the nature of adaptation in asexually reproducing populations with small genome, such as bacteria and yeast. They assumed that sexual species evolve the same way these populations do, i.e. their adaptation is driven by the so-called selective sweeps or newly arising beneficial genetic mutation quickly becomes "fixated" on a particular portion of DNA, with the genome-wide haplotype associated with it. When we relate to obligate sexually reproducing systems, this is much more complicated by the fact that selection can act on standing variation, that means that weak selection can act on many pre-existing genetic variants involved in fitness traits. The idea is that short-term evolution have occurred through a so-called “soft sweep” model, which contrasts the hypothesis of the “hard sweep”, where strong selective sweep originates from a single mutation, while all its linked neutral variants are eliminated. Burke et al. compared outbred, sexually reproducing, replicated populations of D. melanogaster selected for accelerated development and their matched control populations on a genome-wide basis, and this is the first time that such a study of a sexually reproducing species has been done. 


As shown in figure 1, they used the Illumina platform to get short-read sequences from three genomic DNA libraries, obtained from sets of replicated populations experiencing different selection treatments, maintained since 1980 under the specific conditions of large population size (N > 1,000) and discrete generations: 
1) a pooled sample of five replicate populations that have undergone sustained selection for accelerated development and early fertility for over 600 generations (ACO); 
2) a pooled sample of five replicate ancestral control populations, which experience no direct selection on development time (CO); 
3) a single ACO replicate population (ACO1). 
Phenotype was assayed by using longevity assay, starvation resistance assay, development time assay, dry weight assay.

Figure 1. Grey bars represent values measured in each of the five replicate populations in the ACO and CO treatments. Measures from the five baseline (B) replicate populations represent phenotypes typical of populations kept on two-week generation maintenance schedules. Only data for females are shown. Longevity and starvation resistance data were collected after at least 619 generations of ACO treatment, and both development time and dry weight data (dry weight values are mean masses of groups of ten females) were collected after 640 generations of ACO treatment. Error bars, s.e.m. for each replicate population. 

As represented in Figure 2, a 100-kb genome-wide sliding-window analysis was carried out to identify regions diverged in allele frequency, with a large number of genomic regions showing significant difference between the ACO population and their matched controls, while no significant divergence was displayed by the comparison of the single replicate population (ACO1) and the pooled sample consisting of all five ACO populations. The presence of an apparent excess of diverged regions on the X chromosome was explained as a result of selection on initially rare recessive or partially recessive alleles. Another important consideration to do is that the adaptive response was highly multigenic, as not only one or few region were identified to be affected by selection on developmental time, but most likely a larger portion of the genome was involved. 


Figure 2. Sliding-window analysis (100 kb) of differentiation in allele frequency between the ACO and CO populations: the solid black line depicts L10FET5%Q scores at 2-kb steps (Methods). The dotted line is the threshold that any given window has a 0.1% chance of exceeding relative to the genome-wide level of noise. The grey line depicts L10FET5%Q scores for a difference in allele frequency between ACO1 and the ACO pooled sample. The five panels show the five major D. melanogaster chromosome arms (as indicated). 

Looking instead at the heterozigosity throughout the genome, they found a relevant and expected concordance with these results. Regions of reduced heterozygosity are in fact expected to be strongly associated with regions of differentiated allele frequency. Accordingly, if we compare figure 2 with figure 3, we can observe that also in this case the regions identified for divergence in allele frequency were the ones associated with reduced heterozigosity. 

Figure 3. Sliding-window analysis (100 kb) of heterozygosity in the CO pool (blue), the ACO pool (red) and ACO1(grey), with a 2-kb step size. The panels show the five major chromosome arms of D. melanogaster. 

They were also able to exclude that the observed similarity in allele frequencies and patterns of heterozigosity between the ACO1 and ACO libraries was an artifact due to sample preparation or data analysis, by individually genotyping 35 females from the five replicate populations of each selection treatment at 30 loci identified for a divergence in allele frequency (Fig. 4). 

Figure 4. a, Allele frequency estimates of the most common allele at 30 SNPs genotyped in 35 females per replicate population. Red circles represent ACO estimates and grey squares represent CO estimates. Open symbols are allele frequencies for ACO1–ACO5 and CO1–CO5, and filled symbols represent treatment means. Alternating black and grey bars designate the X, 2L, 2R, 3L, and 3R arms, respectively, with grey lines indicating SNP location. b, Scatter plot comparing allele frequency estimates at the same 30 SNPs obtained from the Illumina resequencing versus individual genotyping. Red circles represent ACO, black squares represent CO and the straight line represents a slope of unity. 

Burke found evidence of evolution in more than 500 genes that could be linked to a variety of traits, including size, sexual maturation and life span, indicating a gradual, widespread network of selective adaptation. There are two possible hypothesis to explain the results reported in the paper: either “parallel evolution”, with selection acting on the same intermediate-frequency variants in each population, or unwanted migration between replicate populations. In any case, these results clearly show that the signature of “classic hard sweep” is absent in this population, despite evidence of strong selection, while “soft sweep” model is more consistent with the observations.

Burke, M., Dunham, J., Shahrestani, P., Thornton, K., Rose, M., & Long, A. (2010). Genome-wide analysis of a long-term evolution experiment with Drosophila Nature, 467 (7315), 587-590 DOI: 10.1038/nature09352

Monday, December 17, 2012

Research blogging is 5 years old

And they have published an informative article in PLOS One about their first 4 years:
http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0050109

More cited papers also have more views of the blog post
Biology rules

Friday, December 14, 2012

Genome-wide analysis of a long-term evolution experiment with Drosophila

ResearchBlogging.org

In this paper, Molly K. Burke and his collogues did an experimental evolution systems, which allows the genomic study of adaptation. They selected outbred, sexually reproducing, replicated populations of Drosophila melanogaster, which experienced over 600 generations of laboratory selection for accelerated development.

Short-read sequences from three genomic DNA libraries, were obtained using Illumina platform, they are as follows:
a)    A pooled sample of five replicate populations that have undergone sustained selection for accelerated development and early fertility for over 600 generations (ACO);
b)   A pooled sample of five replicate ancestral control populations, which experience no direct selection on development time (CO);
c)    A single ACO replicate population (ACO1);
Figure 1: Phenotypic divergence in the selection treatments    
In the above figure, the grey bar indicates values measured in the ACO and CO treatments for each of the five replicate populations. B indicates replicate populations, which represent phenotypes typical of populations kept on two-week generation maintenance schedules.

This figure shows a comparative analysis between the ACO population and the population with the CO treatment. Every time, ACO featured significantly differentiated phenotypes, including shorter development time and reductions in pre-adult viability, longevity, adult body size and stress resistance. Furthermore, the CO treatment does not show stringent selection, as it entails no more than moderate selection for postponed reproduction, resulting in moderately increased development time and longevity.

Figure 2: Differentiation throughout the genome    

A 100-kb genome-wide sliding-window analysis test was carried out to identify regions diverged in allele frequency between the ACO and CO libraries and between the ACO and ACO1 libraries. This is due to the fact that, linkage disequilibrium in individual ACO and CO replicate populations may extend anywhere from 20 to 100 (kb). The five different panels are basically for the five major D. melanogaster chromosome arms.

It identifies a large number of genomic regions showing significant divergence between the accelerated development populations and their matched controls depicted by black lines in Figure 2, and very little divergence is observed between a single replicate evolved population (ACO1) and the pooled sample consisting of all five ACO populations and this is indicated by the grey lines. The dotted line is the threshold that any given window has a 0.1% chance of exceeding relative to the genome-wide level of noise. Interestingly, an excess of diverged regions on the X chromosome relative to on the autosomes is very evident. This observation is only expected if adaptation were driven by selection on initially rare recessive or partially recessive alleles. Furthermore, the sharpness of the peaks suggests that regions of the genome that have responded to experimental evolution are precisely identified. However, even the sharpest peaks tend to delineate, 50–100-kb regions.

Kaplan et al. stated that, recent research on evolutionary genetics has focused on classic selective sweeps. In a recombining region, a selected sweep is expected to reduce heterozygosity at SNPs flanking the selected site.
Figure 3: Heterozygosity throughout the genome
Figure 3, shows a similar sliding-window analysis (100 kb) of heterozygosity in ACO and CO lines suggesting that there are indeed local losses of heterozygosity, which is depicted by the red and the blue lines, respectively. Heterozygosity in ACO1 depicted by grey line, shows remarkable concordance with the reductions in heterozygosity in the ACO pool. Regions of reduced heterozygosity are strongly associated with regions of differentiated allele frequency. Interestingly, we observed no location in the genome where heterozygosity is reduced to anywhere near zero, and therefore, it lacks the evidence for a classic sweep is a feature of the data regardless of window size. Nevertheless, both the figure 2 and 3 are quite similar.

Figure 4: Analysis of individual genotypes, measured by cleaved amplified
polymorphic sequence (CAPS) techniques. 

Figure 4a shows the allele frequency estimates of the most common allele at 30 SNPs genotyped in 35 females per replicate population. Red circles and grey squares represent ACO and CO estimates. Open symbols are allele frequencies for ACO1–ACO5 and CO1–CO5, and filled symbols represent treatment means. Alternating black and grey bars designate the X, 2L, 2R, 3L, and 3R arms, respectively. The grey lines indicate SNP location. We observe that replicate populations within a selection treatment have very similar allele frequencies.

In Figure 4b, we see a scatter plot comparing allele frequency estimates at the same 30 SNPs obtained from the Illumina resequencing versus individual genotyping. Red circles represent ACO, black squares represent CO and the straight line represents a slope of unity. Here we see that individual genotypes are consistent with allele frequency estimates from the resequenced pooled libraries.
Therefore, it can be concluded that the congruence in allele frequencies and patterns of heterozygosity between the ACO1 and ACO libraries is unlikely to be some sort of artefact of sample preparation or data analysis.

The study shows a convergence of allele frequencies and heterozygosity levels between replicate populations. This convergence might be due to selection, acting on the same intermediate-frequency variants in each population. Under this scenario, convergence in allele frequencies is due to parallel evolution. Another reason could be, unwanted migration between replicate populations, even at very low levels.

Conclusively, it was very interesting to see that despite strong selection, Molly K. Burke and his collogues failure to observe the signature of a classic sweep in these populations.







Burke, M., Dunham, J., Shahrestani, P., Thornton, K., Rose, M., & Long, A. (2010). Genome-wide analysis of a long-term evolution experiment with Drosophila Nature, 467 (7315), 587-590 DOI: 10.1038/nature09352