J. Chris Pires

Understanding mechanisms of novel gene expression in polyploids

Polyploidy has long been recognized as a prominent force shaping the evolution of eukaryotes, especially flowering plants. New phenotypes often arise with polyploid formation and can contribute to the success of polyploids in nature or their selection for use in agriculture. Although the causes of novel variation in polyploids are not well understood, they could involve changes in gene expression through increased variation in dosage-regulated gene expression, altered regulatory interactions, and rapid genetic and epigenetic changes. New research approaches are being used to study these mechanisms and the results should provide a more complete understanding of polyploidy.

Early allopolyploid evolution in the post-neolithic Brassica napus oilseed genome

The genomic origins of rape oilseed Many domesticated plants arose through the meeting of multiple genomes through hybridization and genome doubling, known as polyploidy. Chalhoub et al. sequenced the polyploid genome of Brassica napus, which originated from a recent combination of two distinct genomes approximately 7500 years ago and gave rise to the crops of rape oilseed (canola), kale, and rutabaga. B. napus has undergone multiple events affecting differently sized genetic regions where a gene from one progenitor species has been converted to the copy from a second progenitor species. Some of these gene conversion events appear to have been selected by humans as part of the process of domestication and crop improvement. Science, this issue p. 950 The polyploid genome of oilseed rape exhibits evolution through homologous gene conversion. Oilseed rape (Brassica napus L.) was formed ~7500 years ago by hybridization between B. rapa and B. oleracea, followed by chromosome doubling, a process known as allopolyploidy. Together with more ancient polyploidizations, this conferred an aggregate 72× genome multiplication since the origin of angiosperms and high gene content. We examined the B. napus genome and the consequences of its recent duplication. The constituent An and Cn subgenomes are engaged in subtle structural, functional, and epigenetic cross-talk, with abundant homeologous exchanges. Incipient gene loss and expression divergence have begun. Selection in B. napus oilseed types has accelerated the loss of glucosinolate genes, while preserving expansion of oil biosynthesis genes. These processes provide insights into allopolyploid evolution and its relationship with crop domestication and improvement.

Genomic changes in resynthesized Brassica napus and their effect on gene expression and phenotype.

Many previous studies have provided evidence for genome changes in polyploids, but there are little data on the overall population dynamics of genome change and whether it causes phenotypic variability. We analyzed genetic, epigenetic, gene expression, and phenotypic changes in ∼50 resynthesized Brassica napus lines independently derived by hybridizing double haploids of Brassica oleracea and Brassica rapa. A previous analysis of the first generation (S0) found that genetic changes were rare, and cytosine methylation changes were frequent. Our analysis of a later generation found that most S0 methylation changes remained fixed in their S5 progeny, although there were some reversions and new methylation changes. Genetic changes were much more frequent in the S5 generation, occurring in every line with lines normally distributed for number of changes. Genetic changes were detected on 36 of the 38 chromosomes of the S5 allopolyploids and were not random across the genome. DNA fragment losses within lines often occurred at linked marker loci, and most fragment losses co-occurred with intensification of signal from homoeologous markers, indicating that the changes were due to homoeologous nonreciprocal transpositions (HNRTs). HNRTs between chromosomes A1 and C1 initiated in early generations, occurred in successive generations, and segregated, consistent with a recombination mechanism. HNRTs and deletions were correlated with qualitative changes in the expression of specific homoeologous genes and anonymous cDNA amplified fragment length polymorphisms and with phenotypic variation among S5 polyploids. Our data indicate that exchanges among homoeologous chromosomes are a major mechanism creating novel allele combinations and phenotypic variation in newly formed B. napus polyploids.

The Brassica oleracea genome reveals the asymmetrical evolution of polyploid genomes

Polyploidization has provided much genetic variation for plant adaptive evolution, but the mechanisms by which the molecular evolution of polyploid genomes establishes genetic architecture underlying species differentiation are unclear. Brassica is an ideal model to increase knowledge of polyploid evolution. Here we describe a draft genome sequence of Brassica oleracea, comparing it with that of its sister species B. rapa to reveal numerous chromosome rearrangements and asymmetrical gene loss in duplicated genomic blocks, asymmetrical amplification of transposable elements, differential gene co-retention for specific pathways and variation in gene expression, including alternative splicing, among a large number of paralogous and orthologous genes. Genes related to the production of anticancer phytochemicals and morphological variations illustrate consequences of genome duplication and gene divergence, imparting biochemical and morphological variation to B. oleracea. This study provides insights into Brassica genome evolution and will underpin research into the many important crops in this genus.

Gene and genome duplications: the impact of dosage-sensitivity on the fate of nuclear genes

Whole genome duplications (WGDs) followed by diploidization, which includes gene loss, have been an important recurrent process in the evolution of higher eukaryotes. Gene retention is biased to specific functional gene categories during diploidization. Dosage-sensitive genes, which include transcription factors, are significantly over-retained following WGDs. By contrast, these same functional gene categories exhibit lower retention rates following smaller scale duplications (e.g., local and tandem duplicates, segmental duplicates, aneuploidy). In light of these recent observations, we review current theories that address the fate of nuclear genes following duplication events (i.e., Gain of Function Hypothesis, Subfunctionalization Hypothesis, Increased Gene Dosage Hypothesis, Functional Buffering Model, and the Gene Balance Hypothesis). We broadly review different mechanisms of dosage-compensation that have evolved to alleviate harmful dosage-imbalances. In addition, we examine a recently proposed extension of the Gene Balance Hypothesis to explain the shared single copy status for a specific functional class of genes across the flowering plants. We speculate that the preferential retention of dosage-sensitive genes (e.g., regulatory genes such as transcription factors) and gene loss following WGDs has played a significant role in the development of morphological complexity in eukaryotes and facilitating speciation, respectively. Lastly, we will review recent findings that suggest polyploid lineages had increased rates of survival and speciation following mass extinction events, including the Cretaceous-Tertiary (KT) extinction.

Homoeologous shuffling and chromosome compensation maintain genome balance in resynthesized allopolyploid Brassica napus

Polyploidy has contributed to the evolution of eukaryotes, particularly flowering plants. The genomic consequences of polyploidy have been extensively studied, but the mechanisms for chromosome stability and diploidization in polyploids remain largely unknown. By using new cytogenetic tools to identify all of the homoeologous chromosomes, we conducted a cytological investigation of 50 resynthesized Brassica napus allopolyploids across generations S0:1 to S5:6 and in the S10:11 generation. Changes in copy number of individual chromosomes were detected in the S0:1 generation and increased in subsequent generations, despite the fact that the mean chromosome number among lines was approximately 38. The chromosome complement of individual plants (segregants) ranged from 36 to 42, with a bias toward the accumulation of extra chromosomes. Karyotype analysis of the S10:11 generation detected aneuploidy and inter- and intragenomic rearrangements, chromosome breakage and fusion, rDNA changes, and loss of repeat sequences. Chromosome sets with extensive homoeology showed the greatest instability. Dosage balance requirements maintained chromosome numbers at or near the tetraploid level, and the loss and gain of chromosomes frequently involved homoeologous chromosome replacement and compensation. These data indicate that early generations of resynthesized B. napus involved aneuploidy and gross chromosomal rearrangements, and that dosage balance mechanisms enforced chromosome number stability. Seed yield and pollen viability were inversely correlated with increasing aneuploidy, and the greatest fertility was observed in two lines that were additive for parental chromosomes. These data on resynthesized B. napus and the correlation of fertility with additive karyotypes cast light on the origins and establishment of natural B. napus.

Patterns of sequence loss and cytosine methylation within a population of newly resynthesized Brassica napus allopolyploids.

Allopolyploid formation requires the adaptation of two nuclear genomes within a single cytoplasm, which may involve programmed genetic and epigenetic changes during the initial generations following genome fusion. To study the dynamics of genome change, we synthesized 49 isogenic Brassica napus allopolyploids and surveyed them with 76 restriction fragment length polymorphism (RFLP) probes and 30 simple sequence repeat (SSR) primer pairs. Here, we report on the types and distribution of genetic and epigenetic changes within the S1 genotypes. We found that insertion/deletion (indel) events were rare, but not random. Of the 57,710 (54,383 RFLP and 3,327 SSR) parental fragments expected among the amphidiploids, we observed 56,676 or 99.9%. Three loci derived from Brassica rapa had indels, and one indel occurred repeatedly across 29% (14/49) of the lines. Loss of one parental fragment was due to the 400-bp reduction of a guanine-adenine dinucleotide repeat-rich sequence. In contrast to the 4% (3/76) RFLP probes that detected indels, 48% (35/73) detected changes in the CpG methylation status between parental genomes and the S1 lines. Some loci were far more likely than others to undergo epigenetic change, but the number of methylation changes within each synthetic polyploid was remarkably similar to others. Clear de novo methylation occurred at a much higher frequency than de novo demethylation within allopolyploid sequences derived from B. rapa. Our results suggest that there is little genetic change in the S0 generation of resynthesized B. napus polyploids. In contrast, DNA methylation was altered extensively in a pattern that indicates tight regulation of epigenetic changes.

Current perspectives and the future of domestication studies

It is difficult to overstate the cultural and biological impacts that the domestication of plants and animals has had on our species. Fundamental questions regarding where, when, and how many times domestication took place have been of primary interest within a wide range of academic disciplines. Within the last two decades, the advent of new archaeological and genetic techniques has revolutionized our understanding of the pattern and process of domestication and agricultural origins that led to our modern way of life. In the spring of 2011, 25 scholars with a central interest in domestication representing the fields of genetics, archaeobotany, zooarchaeology, geoarchaeology, and archaeology met at the National Evolutionary Synthesis Center to discuss recent domestication research progress and identify challenges for the future. In this introduction to the resulting Special Feature, we present the state of the art in the field by discussing what is known about the spatial and temporal patterns of domestication, and controversies surrounding the speed, intentionality, and evolutionary aspects of the domestication process. We then highlight three key challenges for future research. We conclude by arguing that although recent progress has been impressive, the next decade will yield even more substantial insights not only into how domestication took place, but also when and where it did, and where and why it did not.

Homoeologous recombination in allopolyploids: the polyploid ratchet

Polyploidization and recombination are two important processes driving evolution through the building and reshaping of genomes. Allopolyploids arise from hybridization and chromosome doubling among distinct, yet related species. Polyploids may display novel variation relative to their progenitors, and the sources of this variation lie not only in the acquisition of extra gene dosages, but also in the genomic changes that occur after divergent genomes unite. Genomic changes (deletions, duplications, and translocations) have been detected in both recently formed natural polyploids and resynthesized polyploids. In resynthesized Brassica napus allopolyploids, there is evidence that many genetic changes are the consequence of homoeologous recombination. Homoeologous recombination can generate novel gene combinations and phenotypes, but may also destabilize the karyotype and lead to aberrant meiotic behavior and reduced fertility. Thus, natural selection plays a role in the establishment and maintenance of fertile natural allopolyploids that have stabilized chromosome inheritance and a few advantageous chromosomal rearrangements. We discuss the evidence for genome rearrangements that result from homoeologous recombination in resynthesized B. napus and how these observations may inform phenomena such as chromosome replacement, aneuploidy, non-reciprocal translocations and gene conversion seen in other polyploids.

An atlas of over 90,000 conserved noncoding sequences provides insight into crucifer regulatory regions

Despite the central importance of noncoding DNA to gene regulation and evolution, understanding of the extent of selection on plant noncoding DNA remains limited compared to that of other organisms. Here we report sequencing of genomes from three Brassicaceae species (Leavenworthia alabamica, Sisymbrium irio and Aethionema arabicum) and their joint analysis with six previously sequenced crucifer genomes. Conservation across orthologous bases suggests that at least 17% of the Arabidopsis thaliana genome is under selection, with nearly one-quarter of the sequence under selection lying outside of coding regions. Much of this sequence can be localized to approximately 90,000 conserved noncoding sequences (CNSs) that show evidence of transcriptional and post-transcriptional regulation. Population genomics analyses of two crucifer species, A. thaliana and Capsella grandiflora, confirm that most of the identified CNSs are evolving under medium to strong purifying selection. Overall, these CNSs highlight both similarities and several key differences between the regulatory DNA of plants and other species.