HSFA1a modulates plant heat stress responses and alters the 3D chromatin organization of enhancer-promoter interactions

The complex and dynamic three-dimensional organization of chromatin within the nucleus makes understanding the control of gene expression challenging, but also opens up possible ways to epigenetically modulate gene expression. Because plants are sessile, they evolved sophisticated ways to rapidly modulate gene expression in response to environmental stress, that are thought to be coordinated by changes in chromatin conformation to mediate specific cellular and physiological responses. However, to what extent and how stress induces dynamic changes in chromatin reorganization remains poorly understood. Here, we comprehensively investigated genome-wide chromatin changes associated with transcriptional reprogramming response to heat stress in tomato. Our data show that heat stress induces rapid changes in chromatin architecture, leading to the transient formation of promoter-enhancer contacts, likely driving the expression of heat-stress responsive genes. Furthermore, we demonstrate that chromatin spatial reorganization requires HSFA1a, a transcription factor (TF) essential for heat stress tolerance in tomato. In light of our findings, we propose that TFs play a key role in controlling dynamic transcriptional responses through 3D reconfiguration of promoter-enhancer contacts

GENOMIC SEQUENCING OF SELECTED RACES AND A SEXUAL POPULATION FOR IDENTIFICATION AND MOLECULAR MAPPING OF AVIRULENCE GENES AND UNDERSTANDING THE EVOLUTION OF PUCCINIA STRIIFORMIS

Stripe rust fungus Puccinia striiformis (Ps) is a great threat to agriculture in the world. However, the molecular mechanisms underlying the Ps pathogenicity to different cereals and virulence to different cultivars have not been well understood. Taking the advancement of whole-genome sequencing technologies, the objectives of this project were to i) identify and characterize avirulence genes (Avr) in Ps, and ii) decipher the genomic basis of host adaptation of Ps at the forma specialis level.Firstly, we revealed putative pathogenicity mechanisms in P. striiformis f. sp. tritici (Pst, the causal agent of wheat stripe rust) by characterization of secretome and comparison with other rust fungi. We detected a large portion of species-specific secreted proteins that may have specific roles when the fungus interacts with the wheat host. Candidate avirulence genes were identified by incorporating virulence phenotypes into a correlation analysis by whole-genome sequencing of fourteen Pst isolates with balanced virulence profiles.Secondly, we generated genomes with high continuity for Pst and P. striiformis f. sp. hordei (Psh, the causual agent of barley stripe rust). These genomes provide high-quality resources for deciphering the genomic basis of rapid evolution and host adaptation.Thirdly, we compared the genomes of Pst and Psh to study the genomic basis of host adaptation. We found that host adaptation of Ps at the forma specialis level is a complex pathogenic trait, involving not only virulence-related genes but also other genes. Gene loss, which might be adaptive and driven by transposable element activities, provides genomic basis for host adaptation of different Ps formae speciales.Fourthly, we generated a segregating population by self-fertilizing Pst isolate 12-368 to genetically map Avr genes. A genetic map was constructed through whole-genome sequencing of the parental isolate and 117 progeny isolates. Quantitative trait loci analysis mapped six Avr genes to the genetic map. Through referring the molecular markers to the high quality Pst genome, we identified secreted protein genes that could be candidates for further cloning the avirulence genes. Our studies significantly advanced the understanding of the genomic basis of rapid evolution of virulence in the Pst-wheat pathosystem

Development of SP-SNP markers and use them in studying population genetics and identifying markers associated with avirulence genes of Puccinia striiformis F. Sp. Tritici

Stripe rust, caused by Puccinia striiformis Westend. f. sp. tritici Erikss. (Pst), is one of the most destructive diseases of wheat worldwide. Various molecular marker techniques have been used to study the genetic variation of different populations of the pathogen in the world, but none of the previous studies were to develop markers for tagging avirulence genes. The objectives of this study were intended to develop secreted protein - single nucleotide polymorphism (SP-SNP) markers identified in previous studies and use them for studying population genetics and avirulence genes of the pathogen. Firstly, we developed over a hundred SP- SNP markers in the Pst fungus. The development method of the potential non-neutral SP-SNP markers can be applied in other non- v model microorganisms and these markers should be useful to identify genomic regions or genes that are under different selections from both crop and pathogen populations. Secondly, a total of 352 Pst isolates and 23 P. striiformis f. sp. hordei (causing stripe rust on barley) isolates, collected over all stripe rust epidemiological regions in the US to represent a diverse virulence races, were genotyped at 97 SP-SNP loci to study the population diversity and differentiation. Significant differences were detected between the eastern and western US populations and some of the epidemiological regions. Isolates from western US were more diverse than isolates from eastern US. The population diversity and differentiation information from this study would be useful for wheat breeding programs and stripe rust management. Thirdly, 352 Pst isolates were genotyped at 97 SP-SNP loci to identify candidate avirulence (Avr) genes corresponding to various resistance genes using association analyses. Among 19 avirulence genes, significantly associated SP-SNP markers were detected for AvYr1, AvYr2, AvYr6, AvYr7, AvYr8, AvYr44, AvYrExp2, AvYrSP, and AvYrTye. These preliminary results indicated that association analysis is a powerful approach for linking virulence traits of Pst with genetic markers. More SP-SNPs from the whole genome sequences of multiple Pst isolates will be used to identify markers highly associated with avirulence genes using both natural and artificially made sexual populations

Current Status and Future Perspectives of Genomics Research in the Rust Fungi

Rust fungi in Pucciniales have caused destructive plant epidemics, have become more aggressive with new virulence, rapidly adapt to new environments, and continually threaten global agriculture. With the rapid advancement of genome sequencing technologies and data analysis tools, genomics research on many of the devastating rust fungi has generated unprecedented insights into various aspects of rust biology. In this review, we first present a summary of the main findings in the genomics of rust fungi related to variations in genome size and gene composition between and within species. Then we show how the genomics of rust fungi has promoted our understanding of the pathogen virulence and population dynamics. Even with great progress, many questions still need to be answered. Therefore, we introduce important perspectives with emphasis on the genome evolution and host adaptation of rust fungi. We believe that the comparative genomics and population genomics of rust fungi will provide a further understanding of the rapid evolution of virulence and will contribute to monitoring the population dynamics for disease management

Secreted protein gene derived-single nucleotide polymorphisms (SP-SNPs) reveal population diversity and differentiation of Puccinia striiformis f. sp. tritici in the United States

Single nucleotide polymorphism (SNP) is a powerful molecular marker technique that has been widely used in population genetics and molecular mapping studies for various organisms. However, the technique has not been used for studying Puccinia striiformis f. sp. tritici (Pst), the wheat stripe rust pathogen. In this study, we developed over a hundred secreted protein gene-derived SNP (SP-SNP) markers and used 92 markers to study the population structure of Pst. From 352 isolates collected in the United States, we identified 242 multi-locus genotypes. The SP-SNP genotypes had a moderate, but significant correlation with the virulence phenotype data. Clustering of the multi-locus genotypes was consistent by various analyses, revealing distinct genetic groups. Analysis of molecular variance detected significant differences between the eastern and western US Pst populations. High heterozygosity was found in the US population with significant differences identified among epidemiological regions. Analysis of population differentiation revealed that populations between the eastern and western US were highly differentiated while moderate differentiation was found in populations within the western or eastern US. Isolates from the western US were more diverse than isolates from the eastern US. The information is useful for guiding the disease management in different epidemiological regions. •We developed the first set of SNP markers of Puccinia striiformis f. sp. tritici based on secreted protein genes.•We detected significant differences between the eastern and western US P. striiformis f. sp. tritici populations.•We identified high heterozygosity in the pathogen populations with significant differences among epidemiological regions.•We demonstrated the usefulness of SNPs in studying population genetics of wheat stripe rust fungi

Genomic insights into host adaptation between the wheat stripe rust pathogen (Puccinia striiformis f. sp. tritici) and the barley stripe rust pathogen (Puccinia striiformis f. sp. hordei)

Abstract Background Plant fungal pathogens can rapidly evolve and adapt to new environmental conditions in response to sudden changes of host populations in agro-ecosystems. However, the genomic basis of their host adaptation, especially at the forma specialis level, remains unclear. Results We sequenced two isolates each representing Puccinia striiformis f. sp. tritici (Pst) and P. striiformis f. sp. hordei (Psh), different formae speciales of the stripe rust fungus P. striiformis highly adapted to wheat and barley, respectively. The divergence of Pst and Psh, estimated to start 8.12 million years ago, has been driven by high nucleotide mutation rates. The high genomic variation within dikaryotic urediniospores of P. striiformis has provided raw genetic materials for genome evolution. No specific gene families have enriched in either isolate, but extensive gene loss events have occurred in both Pst and Psh after the divergence from their most recent common ancestor. A large number of isolate-specific genes were identified, with unique genomic features compared to the conserved genes, including 1) significantly shorter in length; 2) significantly less expressed; 3) significantly closer to transposable elements; and 4) redundant in pathways. The presence of specific genes in one isolate (or forma specialis) was resulted from the loss of the homologues in the other isolate (or forma specialis) by the replacements of transposable elements or losses of genomic fragments. In addition, different patterns and numbers of telomeric repeats were observed between the isolates. Conclusions Host adaptation of P. striiformis at the forma specialis level is a complex pathogenic trait, involving not only virulence-related genes but also other genes. Gene loss, which might be adaptive and driven by transposable element activities, provides genomic basis for host adaptation of different formae speciales of P. striiformis

Genome Sequence Resources for the Wheat Stripe Rust Pathogen (Puccinia striiformis f. sp. tritici) and the Barley Stripe Rust Pathogen (Puccinia striiformis f. sp. hordei)

Puccinia striiformis f. sp. tritici causes devastating stripe (yellow) rust on wheat and P. striiformis f. sp. hordei causes stripe rust on barley. Several P. striiformis f. sp. tritici genomes are available, but no P. striiformis f. sp. hordei genome is available. More genomes of P. striiformis f. sp. tritici and P. striiformis f. sp. hordei are needed to understand the genome evolution and molecular mechanisms of their pathogenicity. We sequenced P. striiformis f. sp. tritici isolate 93-210 and P. striiformis f. sp. hordei isolate 93TX-2, using PacBio and Illumina technologies and RNA sequencing. Their genomic sequences were assembled to contigs with high continuity and showed significant structural differences. The circular mitochondria genomes of both were complete. These genomes provide high-quality resources for deciphering the genomic basis of rapid evolution and host adaptation, identifying genes for avirulence and other important traits, and studying host-pathogen interactions

Secretome Characterization and Correlation Analysis Reveal Putative Pathogenicity Mechanisms and Identify Candidate Avirulence Genes in the Wheat Stripe Rust Fungus Puccinia striiformis f. sp. tritici

Stripe (yellow) rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat worldwide. Planting resistant cultivars is an effective way to control this disease, but race-specific resistance can be overcome quickly due to the rapid evolving Pst population. Studying the pathogenicity mechanisms is critical for understanding how Pst virulence changes and how to develop wheat cultivars with durable resistance to stripe rust. We re-sequenced 7 Pst isolates and included additional 7 previously sequenced isolates to represent balanced virulence/avirulence profiles for several avirulence loci in seretome analyses. We observed an uneven distribution of heterozygosity among the isolates. Secretome comparison of Pst with other rust fungi identified a large portion of species-specific secreted proteins, suggesting that they may have specific roles when interacting with the wheat host. Thirty-two effectors of Pst were identified from its secretome. We identified candidates for Avr genes corresponding to six Yr genes by correlating polymorphisms for effector genes to the virulence/avirulence profiles of the 14 Pst isolates. The putative AvYr76 was present in the avirulent isolates, but absent in the virulent isolates, suggesting that deleting the coding region of the candidate avirulence gene has produced races virulent to resistance gene Yr76. We conclude that incorporating avirulence/virulence phenotypes into correlation analysis with variations in genomic structure and secretome, particularly presence/absence polymorphisms of effectors, is an efficient way to identify candidate Avr genes in Pst. The candidate effector genes provide a rich resource for further studies to determine the evolutionary history of Pst populations and the co-evolutionary arms race between Pst and wheat. The Avr candidates identified in this study will lead to cloning avirulence genes in Pst, which will enable us to understand molecular mechanisms underlying Pst-wheat interactions, to determine the effectiveness of resistance genes and further to develop durable resistance to stripe rust

Barley FASCIATED EAR genes determine inflorescence meristem size and yield traits

In flowering plants, the inflorescence meristem (IM) provides founder cells to form successive floral meristems, which are precursors of fruits and seeds. The activity and developmental progression of IM are thus critical for yield production in seed crops. In some cereals, such as rice (Oryza sativa) and maize (Zea mays), the size of undifferentiated IM, which is located at the inflorescence apex, is positively associated with yield traits such as spikelet number. However, the relationship between IM size and yield-related spike traits remains unknown in the Triticeae tribe. Here we report that IM size has a negative correlation with yield traits in barley (Hordeum vulgare). Three FASCIATED EAR (FEA) orthologs, HvFEA2, HvFEA3, and HvFEA4, regulate IM size and spike morphogenesis and ultimately affect yield traits. Three HvFEAs genes are highly expressed in developing spikes, and all three loss-of-function mutants exhibit enlarged IM size, shortened spikes, and reduced spikelet number, which may lead to reduced grain yield. Natural variations identified in HvFEAs indicate selection events during barley domestication. We further reveal that HvFEA4, as a transcription factor, potentially targets multiple pathways during reproductive development, including transcriptional control, phytohormone signaling, and redox status. The roles of barley FEA genes in limiting IM size and promoting spikelet formation suggest the potential of increasing yield by manipulating IM activity