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Phylogenetic context and inference of homoeologous expression evolution in Gossypium. (A) Phylogentic relationships among the cotton accessions used in this study. An allopolyploidy event between A- and D-genome diploid species (red star) created modern allopolyploid Gossypium hirsutum (AD1). Using models of the ancenstral genome donors (A2 and D5), an interspecific diploid hybrid (F1) was created (blue star). Although not a perfect match, the model A- and D-genome donors are the best modern representatives of the diploids that underwent allopolyploidization to form AD1 and, as such, provide the best available reconstruction this ancient event. (B) Possible expression phenotypes and associated evolutionary inference. The far left pie represents equal expression among model diploid progenitor species (denoted by A2 and D5). Given this starting condition, several expression states are possible following allopolyploidy or hybridization. Some potential outcomes are indicated by the five pies on the right (At and Dt denote co-resident genomes, either in the hybrid or allopolyploid). (C) Detection of conserved homoeolog-specific single nucleotide polymorphism (SNPs). Given an alignment of expressed sequence tag (EST) sequences from orthologous genes from both diploid and allopolyploid genomes, species- and genome-specific SNPs (all SNPs highlighted in gray) can be detected. The middle SNP is an example of a genome-specific SNP. With this conserved SNP, homoeolog- and allele-specific microarray probes can be generated (potential microarray probe region highlighted in blue), and used to assay expression in allopolyploid and hybrid species.
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Polyploidy has played a prominent role in shaping the genomic architecture of the angiosperms. Through allopolyploidization, several modern Gossypium (cotton) species contain two divergent, although largely redundant genomes. Owing to this redundancy, these genomes can play host to an array of evolutionary processes that act on duplicate genes.
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Citations
... To increase our understanding of the transcriptome-level effects of hybridization, methods for comparing, analyzing, and visualizing gene expression patterns in hybrids have been developed and standardized. These methods include normalization techniques to account for differences in biomass (for polyploids) and library size (Visger et al., 2019;Coate, 2023), and the classification of genes in classes and categories based on their expression patterns (Hegarty et al., 2006;Flagel et al., 2008;Hovav et al., 2008;Rapp et al., 2009). Notably, Rapp et al. (2009) were, to our knowledge, pioneers in introducing a 12-category classification for gene expression, which has since become a standard approach in comparative transcriptomics of hybrid triplets (see, for examples, Chagu e et al., 2010;Chelaifa et al., 2010;Bardil et al., 2011;Yoo et al., 2013;Wu et al., 2018). ...
Hybridization, the process of crossing individuals from diverse genetic backgrounds, plays a pivotal role in evolution, biological invasiveness, and crop breeding.
At the transcriptional level, hybridization often leads to complex nonadditive effects, presenting challenges for understanding its consequences. Although standard transcriptomic analyses exist to compare hybrids to their progenitors, such analyses have not been implemented in a software package, hindering reproducibility.
We introduce hybridexpress, an R/Bioconductor package designed to facilitate the analysis, visualization, and comparison of gene expression patterns in hybrid triplets (hybrids and their progenitors). hybridexpress provides users with a user‐friendly and comprehensive workflow that includes all standard comparative analyses steps, including data normalization, calculation of midparent expression values, sample clustering, expression‐based gene classification into categories and classes, and overrepresentation analysis for functional terms.
We illustrate the utility of hybridexpress through comparative transcriptomic analyses of cotton allopolyploidization and rice root trait heterosis. hybridexpress is designed to streamline comparative transcriptomic studies of hybrid triplets, advancing our understanding of evolutionary dynamics in allopolyploids, and enhancing plant breeding strategies. hybridexpress is freely accessible from Bioconductor (https://bioconductor.org/packages/HybridExpress) and its source code is available on GitHub (https://github.com/almeidasilvaf/HybridExpress).
... Recent studies have utilized multiple biological strategies to elucidate the expression bias and dominance in homologous duplicates in synthetic and natural polyploids (Flagel and Wendel, 2010;Rapp et al., 2009;Wu et al., 2018;Yang et al., 2016;Yoo et al., 2013). Over the past decade, the Wendel laboratory has employed microarray, transcriptome and proteome sequencing to characterize the gene expression patterns of diploid, wild and cultivated allopolyploid cotton, with a particular focus on homoeologous gene expression bias or dominance (Bao et al., 2019;Chaudhary et al., 2009;Flagel et al., 2008;Hu et al., 2021;Hu et al., 2016;Hu et al., 2015;Hu et al., 2014;Hu and Wendel, 2018;Udall et al., 2006). These studies revealed biases towards the A-subgenome (At) or D-subgenome (Dt) for specific genes and processes. ...
... Given the specificity observed among the different varieties within each cotton species, we extracted the intersection of the four varieties to eliminate this variation. Based on differences in gene expression between diploid and tetraploid (sub)genomes, we classified them into five distinct categories (Table 1), as previously described (Chaudhary et al., 2009;Flagel et al., 2008;Yoo et al., 2013). ...
... The ELD phenomenon has been observed in a variety of allopolyploids, including common wheat (Powell et al., 2017), oilseed rape , and coffee (Bardil et al., 2011). In recent years, research on cotton has also gained momentum, with researchers investigating the gene expression patterns of wild and cultivated species in tetraploid cotton by examining the differential expression of orthologous genes across different organs, developmental stages, and evolutionary phases (Chaudhary et al., 2009;Flagel et al., 2008;Hu et al., 2016;Manivannan and Cheeran Amal, 2023;Peng et al., 2022;Peng et al., 2020;Rapp et al., 2009). ...
Polyploidization can lead to the emergence of new species. Allotetraploid cotton arose through hybridization and whole genome duplication of its diploid A and D genome progenitors. As a prominent global fiber crop, cotton serves as an excellent model organism for exploring plant evolutionary biology and fiber development, particularly in polyploidization. This study utilized transcriptomics to analyze expression difference between diploid and allotetraploid cotton, aiming to further the understanding of transcriptomic dynamics associated at different fiber developmental stages during polyploidization process. In this study, we identified additive and non-additive genes and clarified the contributions of non-additive genes during the tetraploid formation process. Furthermore, we constructed a gene co-expression network, providing new insights into the transcriptional regulation of polyploid cotton fiber development. Among non-additive genes, more A genome-biased expression level dominance (ELD-A) than D genome (ELD-D) occurred in the tetraploids. In Gossypium hirsutum, ELD-A genes played major roles during initial elongation, exerting greater dominance in fiber development. Transgressive genes impacted early fiber development in Gossypium barbadense. While the At subgenome profoundly influences tetraploid fiber development, Dt sub-genome activation, inheritance and sub/neo-functionalization promote superior high-yield traits. By constructing co-expression networks, we identified key fiber developmental candidate genes, providing valuable resources for functional research and breeding perspectives to develop superior cotton varieties.
... Rapid and extensive genetic and epigenetic changes often occurred during the establishment and stabilization of newly established allopolyploids 5 . In some allopolyploids, genes from one subgenome were preferentially retained or achieved a higher level of expression than those from other subgenomes 6,7 , which is referred to as "subgenome dominance" and has been documented in an increasing number of allopolyploids [6][7][8][9][10][11][12][13][14][15][16][17] . A classic example of subgenome dominance has been established in maize. ...
Subgenome dominance has been reported in diverse allopolyploid species, where genes from one subgenome are preferentially retained and are more highly expressed than those from other subgenome(s). However, the molecular mechanisms responsible for subgenome dominance remain poorly understood. Here, we develop genome-wide map of accessible chromatin regions (ACRs) in cultivated strawberry (2n = 8x = 56, with A, B, C, D subgenomes). Each ACR is identified as an MNase hypersensitive site (MHS). We discover that the dominant subgenome A contains a greater number of total MHSs and MHS per gene than the submissive B/C/D subgenomes. Subgenome A suffers fewer losses of MHS-related DNA sequences and fewer MHS fragmentations caused by insertions of transposable elements. We also discover that genes and MHSs related to stress response have been preferentially retained in subgenome A. We conclude that preservation of genes and their cognate ACRs, especially those related to stress responses, play a major role in the establishment of subgenome dominance in octoploid strawberry.
... A burst of TE activity (transposition, expansion, and/or amplification) firstly cause changes in gene expression and following directional elimination of gene duplicates in one of the parental subgenomes making more prominent differences between them, i.e. subgenome dominance (Edger et al. 2018;Alger and Edger 2020). Although subgenome conservation is more usual for autopolyploids (Garsmeur et al. 2014;Wang et al. 2017), some allopolyploid genomes also display genome stability and exhibit no genome fractionation, for instance, the allopolyploid genome of the upland cotton Gossypium hirsutum (Flagel et al. 2008). In contrast to allopolyploids, autopolyploids arise through the doubling of the same or highly similar genomes also coming from individuals of polymorphic populations or subspecies. ...
Whole genome duplication (WGD) is an evolutionary event resulting in a redundancy of genetic material. Different mechanisms of WGD, allo- or autopolyploidization, lead to distinct evolutionary trajectories of newly formed polyploids. Genome studies on such species are important for understanding the early stages of genome evolution. However, assembling neopolyploid is a challenging task due to the presence of two homologous (or homeologous) chromosome sets and therefore the existence of the extended paralogous regions in its genome. Post-WGD evolution of polyploids includes cytogenetic diploidization leading to the formation of species, whose polyploid origin might be hidden by disomic inheritance. Earlier we uncovered the hidden polyploid origin of the free-living flatworms of the genus Macrostomum (Macrostomum lignano, M. janickei, and M. mirumnovem). Cytogenetic diploidization in these species accompanied by intensive chromosomal rearrangements including chromosomes fusions. In this study, we unravel the M. lignano genome organization through generation and sequencing of two sublines of the commonly used inbred line of M. lignano (called DV1) differring only in a copy number of the largest chromosome (MLI1). Using non-trivial assembly-free comparative analysis of their genomes, we deciphered DNA sequences belonging to MLI1 and validated them by sequencing the pool of microdissected MLI1. Here we presented the uncommon mechanism of genome rediplodization of M. lignano, which consists in (1) presence of three subgenomes, emerged via formation of large fused chromosome and its variants, and (2) sustaining their heterozygosity through inter- and intrachromosomal rearrangements.
... Homeologue expression bias, expressing one progenitor copy at a higher level than the other progenitor copy, often occurs in allopolyploids across the green plant tree of life (Combes et al. 2013, Akama et al. 2014, Yoo and Wendel 2014, Yang et al. 2016, Hu and Wendel 2019, Sigel et al. 2019, Lee and Adams 2020, Shan et al. 2020, Chen et al. 2021. These biases can occur immediately following allopolyploidy (Flagel et al. 2008, Flagel and Wendel 2010, Edger et al. 2017, Wu et al. 2018) and the number of genes exhibiting homeologue bias appears to increase in older allopolyploid lineages (Flagel et al. 2008, Flagel and Wendel 2010, Edger et al. 2017. Some polyploids exhibit reciprocal homeologue bias (bias towards one progenitor in some conditions and towards the other progenitor in other conditions) in different tissues (Walsh et al. 2020) or under different stress treatments (Dong and Adams 2011), whereas other polyploids do not (Clevenger et al. 2016). ...
... Homeologue expression bias, expressing one progenitor copy at a higher level than the other progenitor copy, often occurs in allopolyploids across the green plant tree of life (Combes et al. 2013, Akama et al. 2014, Yoo and Wendel 2014, Yang et al. 2016, Hu and Wendel 2019, Sigel et al. 2019, Lee and Adams 2020, Shan et al. 2020, Chen et al. 2021. These biases can occur immediately following allopolyploidy (Flagel et al. 2008, Flagel and Wendel 2010, Edger et al. 2017, Wu et al. 2018) and the number of genes exhibiting homeologue bias appears to increase in older allopolyploid lineages (Flagel et al. 2008, Flagel and Wendel 2010, Edger et al. 2017. Some polyploids exhibit reciprocal homeologue bias (bias towards one progenitor in some conditions and towards the other progenitor in other conditions) in different tissues (Walsh et al. 2020) or under different stress treatments (Dong and Adams 2011), whereas other polyploids do not (Clevenger et al. 2016). ...
Homeologue expression bias occurs when one progenitor copy of a gene is expressed at a higher level than the other in allopolyploids. Morphological variation, including differences in flower colour, exists between natural and synthetic allopolyploids of Nicotiana tabacum and their progenitors. In this study, we use a comparative transcriptomic approach to investigate gene expression differences as well as homeologue bias in the flavonoid biosynthetic pathway (FBP) in these accessions. We do not observe reciprocal homeologue bias between dark and light pink allopolyploids, but the production of light pink flowers is correlated with high FLAVONOL SYNTHASE:DIHYDROFLAVONOL-4-REDUCTASE (FLS:DFR) ratio at 60% of anthesis length due to delayed activation of DFR in these accessions. We do find that natural allopolyploids have stronger homeologue bias than synthetic allopolyploids in both FBP genes and across the transcriptome. While there is no overall subgenome dominance, there is a bias towards expression of N. tomentosiformis homeologues in FBP genes; however, the magnitude of this bias is reduced in allopolyploids compared to the progenitors, suggesting that N. sylvestris homeologues play an active role in the development of flower colour in N. tabacum allopolyploids. In addition, synthetic allopolyploids tend to exhibit trans regulation of homeologues whereas natural allopolyploids often have evolved cis-regulatory differences between homeologues since their origin.
... Transcriptional divergence of duplicated genes in polyploids can be resulted from "subgenome dominance," in which genes from one of the parental genomes (subgenomes) are preferentially retained or gain higher levels of expression than those from other subgenomes (12). Subgenome dominance has been documented in diverse polyploids (12)(13)(14)(15)(16)(17)(18)(19)(20). For example, maize is an ancient tetraploid and experienced a WGD 5 to 12 Mya (21). ...
Transcriptional divergence of duplicated genes after whole genome duplication (WGD) has been described in many plant lineages and is often associated with subgenome dominance, a genome-wide mechanism. However, it is unknown what underlies the transcriptional divergence of duplicated genes in polyploid species that lack subgenome dominance. Soybean is a paleotetraploid with a WGD that occurred 5 to 13 Mya. Approximately 50% of the duplicated genes retained from this WGD exhibit transcriptional divergence. We developed accessible chromatin region (ACR) datasets from leaf, flower, and seed tissues using MNase-hypersensitivity sequencing. We validated enhancer function of several ACRs associated with known genes using CRISPR/Cas9-mediated genome editing. The ACR datasets were used to examine and correlate the transcriptional patterns of 17,111 pairs of duplicated genes in different tissues. We demonstrate that ACR dynamics are correlated with divergence of both expression level and tissue specificity of individual gene pairs. Gain or loss of flanking ACRs and mutation of cis -regulatory elements (CREs) within the ACRs can change the balance of the expression level and/or tissue specificity of the duplicated genes. Analysis of DNA sequences associated with ACRs revealed that the extensive sequence rearrangement after the WGD reshaped the CRE landscape, which appears to play a key role in the transcriptional divergence of duplicated genes in soybean. This may represent a general mechanism for transcriptional divergence of duplicated genes in polyploids that lack subgenome dominance.
... One extensively demonstrated effect is the profound rewiring of transcriptomes in response to genomic merger and doubling during allopolyploidization (Giraud et al., 2021;Grover et al., 2012;Shan et al., 2020;Visger et al., 2019). This genome-wide rewiring encompasses a diversity of phenomena, including unequal expression of homoeologs at the genic level (referred to as "homoeolog expression bias") (Flagel et al., 2008;Grover et al., 2012) or the genomic level ("genome dominance") (Schnable et al., 2011), inconsistency in homoeolog biases across tissues or conditions ("expression subfunctionalization and neofunctionalization") (Adams et al., 2003) even at the single cell level (K. Zhang et al., 2023), apparent trans-control of duplicate expression ("expression level dominance") Rapp et al., 2009;, and altered co-expression gene networks (Gallagher et al., 2016;Hu et al., 2016). ...
... In both the F1 and AD1, the total expression of homoeologous genes exhibited more differential expression relative to A2 than to D5 (F1 -13.7% versus 8.0%; AD1 -11.5% versus 7.0%; Figure 6A), and, correspondingly, the ELD analysis revealed more D-dominant than A-dominant expression patterns (F1 -5.8% versus 2.3%; AD1 -4.4% versus 2.2%; Figure 6B). These observations suggest an asymmetric resemblance of the overall transcriptome towards the D-genome diploid parent, as noted previously (Flagel et al., 2008;Rapp et al., 2009;. This trend was consistent across different mapping strategies (Table S16-18). ...
Polyploidy is a prominent mechanism of plant speciation and adaptation, yet the mechanistic understandings of duplicated gene regulation remain elusive. Chromatin structure dynamics are suggested to govern gene regulatory control. Here we characterized genome-wide nucleosome organization and chromatin accessibility in allotetraploid cotton, Gossypium hirsutum (AADD, 2n=4X=52), relative to its two diploid parents (AA or DD genome) and their synthetic diploid hybrid (AD), using DNS-seq. The larger A-genome exhibited wider average nucleosome spacing in diploids, and this inter-genomic difference diminished in the allopolyploid but not hybrid. Allopolyploidization also exhibited increased accessibility at promoters genome-wide and synchronized cis-regulatory motifs between subgenomes. A prominent cis-acting control was inferred for chromatin dynamics and demonstrated by transposable element removal from promoters. Linking accessibility to gene expression patterns, we found distinct regulatory effects for hybridization and later allopolyploid stages, including nuanced establishment of homoeolog expression bias and expression level dominance. Histone gene expression and nucleosome organization are coordinated through chromatin accessibility. Our study demonstrates the capability to track high resolution chromatin structure dynamics and reveals their role in the evolution of cis-regulatory landscapes and duplicate gene expression in polyploids, illuminating regulatory ties to subgenomic asymmetry and dominance.
... We found that both mowed and unmowed plants showed a homoeolog expression bias favoring the B subgenome (Wilcoxon test: p = 0.001 and p = 0.0008, respectively), indicating that P. annua preferentially utilizes B (supina) genes regardless of mowing stress (Fig. 5). Although P. annua's B subgenome expression bias is statistically significant in both treatment comparisons, the bias is not as evident as reported in other neo-allopolyploids [7,[51][52][53], likely reflecting the recent timescale of the hybridization but perhaps also pointing to a more equitable relationship between P. annua's subgenomes where primary metabolic function is partitioned across pairs of homoeologs ( Supplementary Fig. 10). Only chromosomes Homoeolog expression bias tests the polyploid plasticity hypothesis under golf course-style mowing stress. ...
Background
Poa annua (annual bluegrass) is an allotetraploid turfgrass, an agronomically significant weed, and one of the most widely dispersed plant species on earth. Here, we report the chromosome-scale genome assemblies of P. annua’s diploid progenitors, P. infirma and P. supina, and use multi-omic analyses spanning all three species to better understand P. annua’s evolutionary novelty.
Results
We find that the diploids diverged from their common ancestor 5.5 – 6.3 million years ago and hybridized to form P. annua ≤ 50,000 years ago. The diploid genomes are similar in chromosome structure and most notably distinguished by the divergent evolutionary histories of their transposable elements, leading to a 1.7 × difference in genome size. In allotetraploid P. annua, we find biased movement of retrotransposons from the larger (A) subgenome to the smaller (B) subgenome. We show that P. annua’s B subgenome is preferentially accumulating genes and that its genes are more highly expressed. Whole-genome resequencing of several additional P. annua accessions revealed large-scale chromosomal rearrangements characterized by extensive TE-downsizing and evidence to support the Genome Balance Hypothesis.
Conclusions
The divergent evolutions of the diploid progenitors played a central role in conferring onto P. annua its remarkable phenotypic plasticity. We find that plant genes (guided by selection and drift) and transposable elements (mostly guided by host immunity) each respond to polyploidy in unique ways and that P. annua uses whole-genome duplication to purge highly parasitized heterochromatic sequences. The findings and genomic resources presented here will enable the development of homoeolog-specific markers for accelerated weed science and turfgrass breeding.
... Previous work on cotton has suggested a general D-genome bias in QTLs associated with domestication [26,[106][107][108], although surveys of homoeolog expression bias have been less clear [27,109,110]. Notably, these biases appear mostly vertically inherited, although some evidence indicates that homoeolog expression bias also evolves post-polyploidization and during domestication [27,58,[109][110][111]. Here we find few changes in homoeolog expression bias under domestication (Supplementary Table S3), which is expected given analyses of allelic expression in wild x domesticated G. hirsutum hybrids [112]. ...
Cotton has been domesticated independently four times for its fiber, but the genomic targets of selection during each domestication event are mostly unknown. Comparative analysis of the transcriptome during cotton fiber development in wild and cultivated materials holds promise for revealing how independent domestications led to the superficially similar modern cotton fiber phenotype in upland (G. hirsutum) and Pima (G. barbadense) cotton cultivars. Here we examined the fiber transcriptomes of both wild and domesticated G. hirsutum and G. barbadense to compare the effects of speciation versus domestication, performing differential gene expression analysis and coexpression network analysis at four developmental timepoints (5, 10, 15, or 20 days after flowering) spanning primary and secondary wall synthesis. These analyses revealed extensive differential expression between species, timepoints, domestication states, and particularly the intersection of domestication and species. Differential expression was higher when comparing domesticated accessions of the two species than between the wild, indicating that domestication had a greater impact on the transcriptome than speciation. Network analysis showed significant interspecific differences in coexpression network topology, module membership, and connectivity. Despite these differences, some modules or module functions were subject to parallel domestication in both species. Taken together, these results indicate that independent domestication led G. hirsutum and G. barbadense down unique pathways but that it also leveraged similar modules of coexpression to arrive at similar domesticated phenotypes.
... It remains unclear to what extent genome dominance is established instantaneously or gradually. The evidence suggests both: expression bias established in the early generations following WGD may be reinforced over evolutionary time, with the dominant subgenome ultimately contributing more genes to the fully diploidized genome (Flagel et al. 2008;Feldman and Levy 2009;Flagel and Wendel 2010;Woodhouse et al. 2014;Edger et al. 2017). ...
The 'genomic shock' hypothesis posits that unusual challenges to genome integrity such as whole genome duplication (WGD) may induce chaotic genome restructuring. Decades of research on polyploid genomes have revealed that this is often, but not always the case. While some polyploids show major chromosomal rearrangements and de-repression of transposable elements (TEs) in the immediate aftermath of WGD, others do not. Nonetheless, all polyploids show gradual diploidization over evolutionary time. To evaluate these hypotheses, we produced a chromosome-scale reference genome for the natural allotetraploid grass Brachypodium hybridum, accession 'Bhyb26'. We compared two independently-derived accessions of B. hybridum and their deeply diverged diploid progenitor species B. stacei and B. distachyon. The two B. hybridum lineages provide a natural timecourse in genome evolution because one formed 1.4 million years ago, and the other formed 140 thousand years ago. The genome of the older lineage reveals signs of gradual post-WGD genome evolution including minor gene loss and genome rearrangement that are missing from the younger lineage. In neither B. hybridum lineage do we find signs of homeologous recombination or pronounced TE activation, though we find evidence supporting steady post-WGD TE activity in the older lineage. Gene loss in the older lineage was slightly biased toward one subgenome, but genome dominance was not observed at the transcriptomic level. We propose that relaxed selection, rather than an abrupt genomic shock, drives evolutionary novelty in B. hybridum, and that the progenitor species' similarity in TE load may account for the subtlety of the observed genome dominance.
