ABSTRACT
Ribosomal RNAs (rRNAs) are the most abundant cellular RNAs, and their synthesis from rDNA repeats by RNA polymerase I accounts for the bulk of all transcription. Despite substantial variation in rRNA transcription rates across cell types, little is known about cell-type-specific factors that bind rDNA and regulate rRNA transcription to meet tissue-specific needs. Using hematopoiesis as a model system, we mapped about 2,200 ChIP-seq datasets for 250 transcription factors (TFs) and chromatin proteins to human and mouse rDNA and identified robust binding of multiple TF families to canonical TF motifs on rDNA. Using a 47S-FISH-Flow assay developed for nascent rRNA quantification, we demonstrated that targeted degradation of C/EBP alpha (CEBPA), a critical hematopoietic TF with conserved rDNA binding, caused rapid reduction in rRNA transcription due to reduced RNA Pol I occupancy. Our work identifies numerous potential rRNA regulators and provides a template for dissection of TF roles in rRNA transcription.
Subject(s)
RNA Polymerase I , Transcription Factors , Humans , Mice , Animals , RNA Polymerase I/genetics , RNA Polymerase I/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism , RNA, Ribosomal/genetics , Transcription, Genetic , DNA, Ribosomal/genetics , RNA , ChromatinABSTRACT
Ribosomal DNA genes (rDNA) encode the major ribosomal RNAs and in eukaryotes typically form tandem repeat arrays. Species have characteristic rDNA copy numbers, but there is substantial intra-species variation in copy number that results from frequent rDNA recombination. Copy number differences can have phenotypic consequences, however difficulties in quantifying copy number mean we lack a comprehensive understanding of how copy number evolves and the consequences. Here we present a genomic sequence read approach to estimate rDNA copy number based on modal coverage to help overcome limitations with existing mean coverage-based approaches. We validated our method using Saccharomyces cerevisiae strains with known rDNA copy numbers. Application of our pipeline to a global sample of S. cerevisiae isolates showed that different populations have different rDNA copy numbers. Our results demonstrate the utility of the modal coverage method, and highlight the high level of rDNA copy number variation within and between populations.
Subject(s)
Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae , DNA Copy Number Variations , DNA, Ribosomal/genetics , RNA, Ribosomal/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/geneticsABSTRACT
Antimicrobial volatile organic compounds (VOCs) may provide fungi an advantage over other competing microorganisms. As these defensive metabolites are often produced in response to microbial competitors, they are easily overlooked in axenic cultures. We used media supplemented with spent medium from Candida albicans to induce the expression of a broad-spectrum antimicrobial response in a previously uncharacterised white-rot fungus, Scytinostroma sp. Crude extractions of Scytinostroma sp. metabolites were found to be cytotoxic to fibroblast cells and antimicrobial to filamentous fungi, yeasts and Gram-positive bacteria. Volatile antimicrobial activity was observed for Scytinostroma sp. cultures and metabolite extracts using antimicrobial assays in bi-compartmentalised plates. Culture headspace analysis using solid-phase microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS) revealed a pronounced shift in Scytinostroma sp. VOCs when cultured on media supplemented with C. albicans spent medium. We observed a significant increase in the levels of 45 identified VOCs, including 7 metabolites with reported antimicrobial activity. Using preparative HPLC combined with GC-MS, we determined that isovelleral is likely to be the main broad-spectrum antimicrobial metabolite produced by Scytinostroma sp. Isovelleral is a sesquiterpene dialdehyde with both antibiotic and antifeedant properties, previously detected in fruit bodies of other Basidiomycetes.
Subject(s)
Basidiomycota , Volatile Organic Compounds , Fruit , Gas Chromatography-Mass Spectrometry , Solid Phase MicroextractionABSTRACT
Hybridization is a route to speciation that occurs widely across the eukaryote tree of life. The success of allopolyploids (hybrid species with increased ploidy) and homoploid hybrids (with unchanged ploidy) is well documented. However, their formation and establishment is not straightforward, with a suite of near-instantaneous and longer term biological repercussions faced by the new species. Central to these challenges is the rewiring of gene regulatory networks following the merger of distinct genomes inherited from both parental species. Research on the evolution of hybrid gene expression has largely involved studies on a single hybrid species or a few gene families. Here, we present the first standardized transcriptome-wide study exploring the fates of genes following hybridization across three kingdoms: animals, plants and fungi. Within each kingdom, we pair an allopolyploid system with a closely related homoploid hybrid to decouple the influence of increased ploidy from genome merger. Genome merger, not changes in ploidy, has the greatest effect on posthybridization expression patterns across all study systems. Strikingly, we find that differentially expressed genes in parent species preferentially switch to more similar expression in hybrids across all kingdoms, likely as a consequence of regulatory trans-acting cross-talk within the hybrid nucleus. We also highlight the prevalence of gene loss or silencing among extremely differentially expressed genes in hybrid species across all kingdoms. These shared patterns suggest that the evolutionary process of hybridization leads to common high-level expression outcomes, regardless of the particular species or kingdom.
Subject(s)
Hybridization, Genetic , Transcriptome , Animals , Eukaryota/genetics , Genome , PloidiesABSTRACT
Saccharomyces cerevisiae is an exceptional genetic system, with genetic crosses facilitated by its ability to be maintained in haploid and diploid forms. Such crosses are straightforward if the mating type/ploidy of the strains is known. Several techniques can determine mating type (or ploidy), but all have limitations. Here, we validate a simple, cheap and robust method to identify S. cerevisiae mating types. When cells of opposite mating type are mixed in liquid media, they 'creep' up the culture vessel sides, a phenotype that can be easily detected visually. In contrast, mixtures of the same mating type or with a diploid simply settle out. The phenotype is observable for several days under a range of routine growth conditions and with different media/strains. Microscopy suggests that cell aggregation during mating is responsible for the phenotype. Yeast knockout collection analysis identified 107 genes required for the creeping phenotype, with these being enriched for mating-specific genes. Surprisingly, the RIM101 signaling pathway was strongly represented. We propose that RIM101 signaling regulates aggregation as part of a wider, previously unrecognized role in mating. The simplicity and robustness of this method make it ideal for routine verification of S. cerevisiae mating type, with future studies required to verify its molecular basis.
Subject(s)
Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae , Haploidy , Phenotype , Saccharomyces cerevisiae/physiology , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolismABSTRACT
The beta-actin gene (ACTB) encodes a ubiquitous cytoskeletal protein, essential for embryonic development in humans. De novo heterozygous missense variants in the ACTB are implicated in causing Baraitser-Winter cerebrofrontofacial syndrome (BWCFFS; MIM#243310). ACTB pathogenic variants are rarely associated with intestinal malformations. We report on a rare case of monozygotic twins presenting with proximal small bowel atresia and hydrops in one, and apple-peel bowel atresia and laryngeal dysgenesis in the other. The twin with hydrops could not be resuscitated. Intensive and surgical care was provided to the surviving twin. Rapid trio genome sequencing identified a de novo missense variant in ACTB (NM_00101.3:c.1043C>T; p.(Ser348Leu)) that guided the care plan. The identical variant subsequently was identified in the demised twin. To characterize the functional effect, the variant was recreated as a pseudoheterozygote in a haploid wild-type S. cerevisiae strain. There was an obvious growth defect of the yACT1S348L/WT pseudoheterozygote compared to a yACT1WT/WT strain when grown at 22°C but not when grown at 30°C, consistent with the yACT1 S348L variant having a functional defect that is dominant over the wild-type allele. The functional results provide supporting evidence that the Ser348Leu variant is likely to be a pathogenic variant, including being associated with intestinal malformations in BWCFFS, and can demonstrate variable expressivity within monozygotic twins.
Subject(s)
Intestinal Atresia , Twins, Monozygotic , Actins/genetics , Actins/metabolism , Biological Variation, Population , Craniofacial Abnormalities , Edema , Epilepsy , Facies , Humans , Intellectual Disability , Intestinal Atresia/diagnosis , Intestinal Atresia/genetics , Lissencephaly , Saccharomyces cerevisiae/metabolism , Twins, Monozygotic/geneticsABSTRACT
Structural features of genomes, including the three-dimensional arrangement of DNA in the nucleus, are increasingly seen as key contributors to the regulation of gene expression. However, studies on how genome structure and nuclear organisation influence transcription have so far been limited to a handful of model species. This narrow focus limits our ability to draw general conclusions about the ways in which three-dimensional structures are encoded, and to integrate information from three-dimensional data to address a broader gamut of biological questions. Here, we generate a complete and gapless genome sequence for the filamentous fungus, Epichloë festucae. We use Hi-C data to examine the three-dimensional organisation of the genome, and RNA-seq data to investigate how Epichloë genome structure contributes to the suite of transcriptional changes needed to maintain symbiotic relationships with the grass host. Our results reveal a genome in which very repeat-rich blocks of DNA with discrete boundaries are interspersed by gene-rich sequences that are almost repeat-free. In contrast to other species reported to date, the three-dimensional structure of the genome is anchored by these repeat blocks, which act to isolate transcription in neighbouring gene-rich regions. Genes that are differentially expressed in planta are enriched near the boundaries of these repeat-rich blocks, suggesting that their three-dimensional orientation partly encodes and regulates the symbiotic relationship formed by this organism.
Subject(s)
DNA, Fungal/genetics , Epichloe/genetics , Gene Expression Regulation, Fungal , Genome, Fungal/genetics , Repetitive Sequences, Nucleic Acid/genetics , AT Rich Sequence/genetics , DNA, Fungal/chemistry , Fungal Proteins/genetics , GC Rich Sequence/genetics , Gene Expression Profiling/methods , Hyphae/genetics , Sequence Analysis, DNA/methods , Symbiosis/geneticsABSTRACT
This study investigated the impact of varying sound conditions (frequency and intensity) on yeast growth, fermentation performance and production of volatile organic compounds (VOCs) in beer. Fermentations were carried out in plastic bags suspended in large water-filled containers fitted with underwater speakers. Ferments were subjected to either 200-800 or 800-2000 Hz at 124 and 140 dB @ 20 µPa. Headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) was used to identify and measure the relative abundance of the VOCs produced. Sound treatment had significant effects on the number of viable yeast cells in suspension at 10 and 24 h (p < 0.05), with control (silence) samples having the highest cell numbers. For wort gravity, there were significant differences between treatments at 24 and 48 h, with the silence control showing the lowest density before all ferments converged to the same final gravity at 140 h. A total of 33 VOCs were identified in the beer samples, including twelve esters, nine alcohols, three acids, three aldehydes, and six hop-derived compounds. Only the abundance of some alcohols showed any consistent response to the sound treatments. These results show that the application of audible sound via underwater transmission to a beer fermentation elicited limited changes to wort gravity and VOCs during fermentation.
Subject(s)
Beer/analysis , Fermentation , Saccharomyces cerevisiae/growth & development , Sound , Volatile Organic Compounds/analysis , Cell Count , Esters/analysis , Hydrogen-Ion Concentration , Principal Component Analysis , Saccharomyces cerevisiae/cytologyABSTRACT
Mammalian genomes contain a number of duplicated genes, and sequence identity between these duplicates can be maintained by purifying selection. However, between-duplicate recombination can also maintain sequence identity between copies, resulting in a pattern known as concerted evolution where within-genome repeats are more similar to each other than to orthologous repeats in related species. Here we investigated the tandemly-repeated keratin-associated protein 1 (KAP1) gene family, KRTAP1, which encodes proteins that are important components of hair and wool in mammals. Comparison of eutherian mammal KRTAP1 gene repeats within and between species shows a strong pattern of concerted evolution. However, in striking contrast to the coding regions of these genes, we find that the flanking regions have a divergent pattern of evolution. This contrast in evolutionary pattern transitions abruptly near the start and stop codons of the KRTAP1 genes. We reveal that this difference in evolutionary patterns is not explained by conventional purifying selection, nor is it likely a consequence of codon adaptation or reverse transcription of KRTAP1-n mRNA. Instead, the evidence suggests that these contrasting patterns result from short-tract gene conversion events that are biased to the KRTAP1 coding region by selection and/or differential sequence divergence. This work demonstrates the power that gene conversion has to finely shape the evolution of repetitive genes, and provides another distinctive pattern of contrasting evolutionary outcomes that results from gene conversion. A greater emphasis on exploring the evolution of multi-gene eukaryotic families will reveal how common different contrasting evolutionary patterns are in gene duplicates.
Subject(s)
Evolution, Molecular , Keratins/genetics , Mammals/genetics , Open Reading Frames/genetics , Animals , Base Sequence , Codon/genetics , DNA, Intergenic/genetics , Gene Conversion , Keratins/metabolism , Phylogeny , Polymorphism, Genetic , RNA, Messenger/genetics , RNA, Messenger/metabolism , Selection, Genetic , Sheep/genetics , Tandem Repeat Sequences/geneticsABSTRACT
In eukaryotes, the ribosomal DNA (rDNA) consists of long tandem repeat arrays. These repeated genes are unstable because homologous recombination between them results in copy number loss. To maintain high copy numbers, yeast has an amplification system that works through a pathway involving the replication fork barrier site and unequal sister chromatid recombination. In this study, we show that an active replication origin is essential for amplification, and the amplification rate correlates with origin activity. Moreover, origin activity affects the levels of extrachromosomal rDNA circles (ERC) that are thought to promote aging. Surprisingly, we found that reduction in ERC level results in shorter life span. We instead show that life span correlates with rDNA stability, which is preferentially reduced in mother cells, and that episomes can induce rDNA instability. These data support a model in which rDNA instability itself is a cause of aging in yeast.
Subject(s)
Cell Division/genetics , DNA Replication , DNA, Ribosomal/biosynthesis , Gene Amplification , Gene Expression Regulation, Fungal , Genomic Instability , Replication Origin , Saccharomyces cerevisiae/genetics , Aging/genetics , DNA, Circular/metabolism , DNA-Binding Proteins/metabolism , Kinetics , Mutation , Plasmids/metabolism , Recombination, Genetic , Saccharomyces cerevisiae/growth & development , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Sister Chromatid ExchangeABSTRACT
Polyploidy, a state in which the chromosome complement has undergone an increase, is a major force in evolution. Understanding the consequences of polyploidy has received much attention, and allopolyploids, which result from the union of two different parental genomes, are of particular interest because they must overcome a suite of biological responses to this merger, known as "genome shock." A key question is what happens to gene expression of the two gene copies following allopolyploidization, but until recently the tools to answer this question on a genome-wide basis were lacking. Here we utilize high throughput transcriptome sequencing to produce the first genome-wide picture of gene expression response to allopolyploidy in fungi. A novel pipeline for assigning sequence reads to the gene copies was used to quantify their expression in a fungal allopolyploid. We find that the transcriptional response to allopolyploidy is predominantly conservative: both copies of most genes are retained; over half the genes inherit parental gene expression patterns; and parental differential expression is often lost in the allopolyploid. Strikingly, the patterns of gene expression change are highly concordant with the genome-wide expression results of a cotton allopolyploid. The very different nature of these two allopolyploids implies a conserved, eukaryote-wide transcriptional response to genome merger. We provide evidence that the transcriptional responses we observe are mostly driven by intrinsic differences between the regulatory systems in the parent species, and from this propose a mechanistic model in which the cross-kingdom conservation in transcriptional response reflects conservation of the mutational processes underlying eukaryotic gene regulatory evolution. This work provides a platform to develop a universal understanding of gene expression response to allopolyploidy and suggests that allopolyploids are an exceptional system to investigate gene regulatory changes that have evolved in the parental species prior to allopolyploidization.
Subject(s)
Biological Evolution , Hybridization, Genetic/genetics , Polyploidy , Gene Expression Profiling , Gene Expression Regulation, Fungal , Genome, Fungal , High-Throughput Nucleotide SequencingABSTRACT
Polyploidy-the increase in the number of whole chromosome sets-is an important evolutionary force in eukaryotes. Polyploidy is well recognized throughout the evolutionary history of plants and animals, where several ancient events have been hypothesized to be drivers of major evolutionary radiations. However, fungi provide a striking contrast: while numerous recent polyploids have been documented, ancient fungal polyploidy is virtually unknown. We present a survey of known fungal polyploids that confirms the absence of ancient fungal polyploidy events. Three hypotheses may explain this finding. First, ancient fungal polyploids are indeed rare, with unique aspects of fungal biology providing similar benefits without genome duplication. Second, fungal polyploids are not successful in the long term, leading to few extant species derived from ancient polyploidy events. Third, ancient fungal polyploids are difficult to detect, causing the real contribution of polyploidy to fungal evolution to be underappreciated. We consider each of these hypotheses in turn and propose that failure to detect ancient events is the most likely reason for the lack of observed ancient fungal polyploids. We examine whether existing data can provide evidence for previously unrecognized ancient fungal polyploidy events but discover that current resources are too limited. We contend that establishing whether unrecognized ancient fungal polyploidy events exist is important to ascertain whether polyploidy has played a key role in the evolution of the extensive complexity and diversity observed in fungi today and, thus, whether polyploidy is a driver of evolutionary diversifications across eukaryotes. Therefore, we conclude by suggesting ways to test the hypothesis that there are unrecognized polyploidy events in the deep evolutionary history of the fungi.
Subject(s)
Evolution, Molecular , Fungi/genetics , Polyploidy , Biological EvolutionABSTRACT
The short arms of the five acrocentric human chromosomes harbor sequences that direct the assembly and function of the nucleolus, one of the key functional domains of the nucleus, yet they are absent from the current human genome assembly. Here we describe the genomic architecture of these human nucleolar organizers. Sequences distal and proximal to ribosomal gene arrays are conserved among the acrocentric chromosomes, suggesting they are sites of frequent recombination. Although previously believed to be heterochromatic, characterization of these two flanking regions reveals that they share a complex genomic architecture similar to other euchromatic regions of the genome, but they have distinct genomic characteristics. Proximal sequences are almost entirely segmentally duplicated, similar to the regions bordering centromeres. In contrast, the distal sequence is predominantly unique to the acrocentric short arms and is dominated by a very large inverted repeat. We show that the distal element is localized to the periphery of the nucleolus, where it appears to anchor the ribosomal gene repeats. This, combined with its complex chromatin structure and transcriptional activity, suggests that this region is involved in nucleolar organization. Our results provide a platform for investigating the role of NORs in nucleolar formation and function, and open the door for determining the role of these regions in the well-known empirical association of nucleoli with pathology.
Subject(s)
Cell Nucleolus/genetics , Chromatin/genetics , DNA, Ribosomal/genetics , Nucleolus Organizer Region/genetics , Cell Line, Tumor , Centromere , Chromosomes, Human , Genome, Human , HeLa Cells , Humans , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis , Ribosomes/genetics , Segmental Duplications, Genomic , Sequence Analysis, DNA , TransfectionABSTRACT
BACKGROUND: Forming a new species through the merger of two or more divergent parent species is increasingly seen as a key phenomenon in the evolution of many biological systems. However, little is known about how expression of parental gene copies (homeologs) responds following genome merger. High throughput RNA sequencing now makes this analysis technically feasible, but tools to determine homeolog expression are still in their infancy. RESULTS: Here we present HyLiTE - a single-step analysis to obtain tables of homeolog expression in a hybrid or allopolyploid and its parent species directly from raw mRNA sequence files. By implementing on-the-fly detection of diagnostic parental polymorphisms, HyLiTE can perform SNP calling and read classification simultaneously, thus allowing HyLiTE to be run as parallelized code. HyLiTE accommodates any number of parent species, multiple data sources (including genomic DNA reads to improve SNP detection), and implements a statistical framework optimized for genes with low to moderate expression. CONCLUSIONS: HyLiTE is a flexible and easy-to-use program designed for bench biologists to explore patterns of gene expression following genome merger. HyLiTE offers practical advantages over manual methods and existing programs, has been designed to accommodate a wide range of genome merger systems, can identify SNPs that arose following genome merger, and offers accurate performance on non-model organisms.
Subject(s)
Biological Evolution , Gene Expression Regulation , High-Throughput Nucleotide Sequencing/methods , Polyploidy , Sequence Analysis, DNA/methods , Software , Animals , Fungi/genetics , Goldfish/genetics , Gossypium/genetics , Hybridization, Genetic , Polymorphism, Single Nucleotide/geneticsABSTRACT
We sequenced and compared the genomes of the Dothideomycete fungal plant pathogens Cladosporium fulvum (Cfu) (syn. Passalora fulva) and Dothistroma septosporum (Dse) that are closely related phylogenetically, but have different lifestyles and hosts. Although both fungi grow extracellularly in close contact with host mesophyll cells, Cfu is a biotroph infecting tomato, while Dse is a hemibiotroph infecting pine. The genomes of these fungi have a similar set of genes (70% of gene content in both genomes are homologs), but differ significantly in size (Cfu >61.1-Mb; Dse 31.2-Mb), which is mainly due to the difference in repeat content (47.2% in Cfu versus 3.2% in Dse). Recent adaptation to different lifestyles and hosts is suggested by diverged sets of genes. Cfu contains an α-tomatinase gene that we predict might be required for detoxification of tomatine, while this gene is absent in Dse. Many genes encoding secreted proteins are unique to each species and the repeat-rich areas in Cfu are enriched for these species-specific genes. In contrast, conserved genes suggest common host ancestry. Homologs of Cfu effector genes, including Ecp2 and Avr4, are present in Dse and induce a Cf-Ecp2- and Cf-4-mediated hypersensitive response, respectively. Strikingly, genes involved in production of the toxin dothistromin, a likely virulence factor for Dse, are conserved in Cfu, but their expression differs markedly with essentially no expression by Cfu in planta. Likewise, Cfu has a carbohydrate-degrading enzyme catalog that is more similar to that of necrotrophs or hemibiotrophs and a larger pectinolytic gene arsenal than Dse, but many of these genes are not expressed in planta or are pseudogenized. Overall, comparison of their genomes suggests that these closely related plant pathogens had a common ancestral host but since adapted to different hosts and lifestyles by a combination of differentiated gene content, pseudogenization, and gene regulation.
Subject(s)
Adaptation, Physiological/genetics , Cladosporium/genetics , Genome , Host-Pathogen Interactions , Base Sequence , Fungal Proteins/genetics , Gene Expression Regulation, Fungal , Solanum lycopersicum/genetics , Solanum lycopersicum/parasitology , Phylogeny , Pinus/genetics , Pinus/parasitology , Plant Diseases/geneticsABSTRACT
The yeast Saccharomyces cerevisiae is a powerful model system to understand the molecular basis of aging. It has been known for over 50 years that yeast cells have a finite replicative capacity and develop an aging phenotype, and much recent research has focused on the molecular changes that underlie this replicative senescence. A common thread in many yeast replicative aging studies is the involvement of the ribosomal DNA gene repeats (rDNA), beginning with the discovery that the rDNA silencing gene, SIR2, regulates life span. In 2008, a novel aging hypothesis, termed the rDNA theory of aging, was presented where the high level of genomic instability at the rDNA repeats was proposed to dominate global genome stability and determine the life span. Here, we review the rDNA theory of aging and discuss a number of recent studies that provide important new data on the roles of the rDNA in yeast replicative aging. Based on these recent results, we propose an integrative model of the rDNA theory of aging that encompasses genomic instability, chromatin relocalization following DNA repair, and replication stress in a self-reinforcing cyclical pathway that is primarily manifested at the rDNA repeats and results in the aging phenotype.
Subject(s)
Cell Death , DNA, Ribosomal/metabolism , Saccharomyces cerevisiae/physiology , Silent Information Regulator Proteins, Saccharomyces cerevisiae/metabolism , Sirtuin 2/metabolism , Models, Biological , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolismABSTRACT
The genes encoding ribosomal RNA are highly conserved across life and in almost all eukaryotes are present in large tandem repeat arrays called the rDNA. rDNA repeat unit size is conserved across most eukaryotes, but has expanded dramatically in mammals, principally through expansion of the intergenic spacer region that separates adjacent rRNA coding regions. Here we used long-read sequence data from representatives of the major amniote lineages to determine where in amniote evolution rDNA unit size increased. We find that amniote rDNA unit sizes fall into two narrow size classes: 'normal' (â¼11-20 kb) in all amniotes except monotreme, marsupial and eutherian mammals, which have 'large' (â¼35-45 kb) sizes. We confirm that increases in intergenic spacer length explain much of this mammalian size increase but, in stark contrast to the uniformity of mammalian rDNA unit size, mammalian intergenic spacers differ greatly in sequence. These results suggest a large increase in intergenic spacer size occurred in a mammalian ancestor and has been maintained despite substantial sequence changes over the course of mammalian evolution. This points to a previously unrecognized constraint on the length of the intergenic spacer, a region that was thought to be largely neutral. We finish by speculating on possible causes of this constraint.
ABSTRACT
Plant pathogens use a complex arsenal of weapons, such as toxic secondary metabolites, to invade and destroy their hosts. Knowledge of how secondary metabolite pathways evolved is central to understanding the evolution of host specificity. The secondary metabolite dothistromin is structurally similar to aflatoxins and is produced by the fungal pine pathogen Dothistroma septosporum. Our study focused on dothistromin genes, which are widely dispersed across one chromosome, to determine whether this unusual distributed arrangement evolved from an ancestral cluster. We combined comparative genomics and population genetics approaches to elucidate the origins of the dispersed arrangement of dothistromin genes over a broad evolutionary time-scale at the phylum, class and species levels. Orthologs of dothistromin genes were found in two major classes of fungi. Their organization is consistent with clustering of core pathway genes in a common ancestor, but with intermediate cluster fragmentation states in the Dothideomycetes fungi. Recombination hotspots in a D. septosporum population matched sites of gene acquisition and cluster fragmentation at higher evolutionary levels. The results suggest that fragmentation of a larger ancestral cluster gave rise to the arrangement seen in D. septosporum. We propose that cluster fragmentation may facilitate metabolic retooling and subsequent host adaptation of plant pathogens.