Grain yield improvement by genome editing of TaARF12 that decoupled peduncle and rachis development trajectories via differential regulation of gibberellin signalling in wheat.

Plant breeding is constrained by trade-offs among different agronomic traits by the pleiotropic nature of many genes. Genes that contribute to two or more favourable traits with no penalty on yield are rarely reported, especially in wheat. Here, we describe the editing of a wheat auxin response factor TaARF12 by using CRISPR/Cas9 that rendered shorter plant height with larger spikes. Changes in plant architecture enhanced grain number per spike up to 14.7% with significantly higher thousand-grain weight and up to 11.1% of yield increase under field trials. Weighted Gene Co-Expression Network Analysis (WGCNA) of spatial-temporal transcriptome profiles revealed two hub genes: RhtL1, a DELLA domain-free Rht-1 paralog, which was up-regulated in peduncle, and TaNGR5, an organ size regulator that was up-regulated in rachis, in taarf12 plants. The up-regulation of RhtL1 in peduncle suggested the repression of GA signalling, whereas up-regulation of TaNGR5 in spike may promote GA response, a working model supported by differential expression patterns of GA biogenesis genes in the two tissues. Thus, TaARF12 complemented plant height reduction with larger spikes that gave higher grain yield. Manipulation of TaARF12 may represent a new strategy in trait pyramiding for yield improvement in wheat.

Wheat breeding history reveals synergistic selection of pleiotropic genomic sites for plant architecture and grain yield.

Diversity surveys of crop germplasm are important for gaining insights into the genomic basis for plant architecture and grain yield improvement, which is still poorly understood in wheat. In this study, we exome sequenced 287 wheat accessions that were collected in the past 100 years. Population genetics analysis identified that 6.7% of the wheat genome falls within the selective sweeps between landraces and cultivars, which harbors the genes known for yield improvement. These regions were asymmetrically distributed on the A and B subgenomes with regulatory genes being favorably selected. Genome-wide association study (GWAS) identified genomic loci associated with traits for yield potential, and two underlying genes, TaARF12 encoding an auxin response factor and TaDEP1 encoding the G-protein γ-subunit, were located and characterized to pleiotropically regulate both plant height and grain weight. Elite single-nucleotide haplotypes with increased allele frequency in cultivars relative to the landraces were identified and found to have accumulated over the course of breeding. Interestingly, we found that TaARF12 and TaDEP1 function in epistasis with the classical plant height Rht-1 locus, leading to propose a "Green Revolution"-based working model for historical wheat breeding. Collectively, our study identifies selection signatures that fine-tune the gibberellin pathway during modern wheat breeding and provides a wealth of genomic diversity resources for the wheat research community.

mRNA and Small RNA Transcriptomes Reveal Insights into Dynamic Homoeolog Regulation of Allopolyploid Heterosis in Nascent Hexaploid Wheat.

Nascent allohexaploid wheat may represent the initial genetic state of common wheat (Triticum aestivum), which arose as a hybrid between Triticum turgidum (AABB) and Aegilops tauschii (DD) and by chromosome doubling and outcompeted its parents in growth vigor and adaptability. To better understand the molecular basis for this success, we performed mRNA and small RNA transcriptome analyses in nascent allohexaploid wheat and its following generations, their progenitors, and the natural allohexaploid cultivar Chinese Spring, with the assistance of recently published A and D genome sequences. We found that nonadditively expressed protein-coding genes were rare but relevant to growth vigor. Moreover, a high proportion of protein-coding genes exhibited parental expression level dominance, with genes for which the total homoeolog expression level in the progeny was similar to that in T. turgidum potentially participating in development and those with similar expression to that in Ae. tauschii involved in adaptation. In addition, a high proportion of microRNAs showed nonadditive expression upon polyploidization, potentially leading to differential expression of important target genes. Furthermore, increased small interfering RNA density was observed for transposable element-associated D homoeologs in the allohexaploid progeny, which may account for biased repression of D homoeologs. Together, our data provide insights into small RNA-mediated dynamic homoeolog regulation mechanisms that may contribute to heterosis in nascent hexaploid wheat.

The SEPALLATA MADS-box protein SLMBP21 forms protein complexes with JOINTLESS and MACROCALYX as a transcription activator for development of the tomato flower abscission zone.

Organ abscission is a key step in a plant's life cycle and is one of the most important agronomic traits for crops. In tomato, two MADS-box genes, JOINTLESS (J) and MACROCAYLYX (MC), have been shown to be implicated in development of the flower abscission zone (AZ), but the molecular mechanisms underlying this process are not well known. We report here that the SEPALLATA (SEP) MADS-box gene SLMBP21 acts as an additional factor for development of the AZ in tomato. We show that knockdown of SLMBP21 abolishes development of the flower AZ, while overexpression of SLMBP21 produces small cells at the proximal section of the pedicel and the peduncle. Bimolecular fluorescence complementation analysis confirms that SLMBP21 interacts with J and MC, and co-immunoprecipitation assays further demonstrates that these three proteins may form higher-order protein complexes. In situ hybridization shows that SLMBP21, J, and MC transcripts accumulate in distinct regions, but overlap at the AZ vasculature. In addition, transactivation assays in yeast show that, of the three interacting proteins, only SLMBP21 can activate reporter gene transcription. RNA-seq analysis furthermore reveals that loss of function of SLMBP21, J, or MC affects a common subset of meristem activity genes including LeWUS and LATERAL SUPPRESSOR that were specifically expressed in the AZ on the tomato flower pedicel. Since SLMBP21 belongs to the FBP9/23 subclade of the SEP gene family, which is absent in Arabidopsis, the SLMBP21-J-MC complex may represent a distinct mechanism for development of the AZ in plants.

TaCPK2-A, a calcium-dependent protein kinase gene that is required for wheat powdery mildew resistance enhances bacterial blight resistance in transgenic rice.

Calcium-dependent protein kinases (CPKs) are important Ca2+ signalling components involved in complex immune and stress signalling networks; but the knowledge of CPK gene functions in the hexaploid wheat is limited. Previously, TaCPK2 was shown to be inducible by powdery mildew (Blumeria graminis tritici, Bgt) infection in wheat. Here, its functions in disease resistance are characterized further. This study shows the presence of defence-response and cold-response cis-elements on the promoters of the A subgenome homoeologue (TaCPK2-A) and D subgenome homoeologue (TaCPK2-D), respectively. Their expression patterns were then confirmed by quantitative real-time PCR (qRT-PCR) using genome-specific primers, where TaCPK2-A was induced by Bgt treatment while TaCPK2-D mainly responded to cold treatment. Downregulation of TaCPK2-A by virus-induced gene silencing (VIGS) causes loss of resistance to Bgt in resistant wheat lines, indicating that TaCPK2-A is required for powdery mildew resistance. Furthermore, overexpression of TaCPK2-A in rice enhanced bacterial blight (Xanthomonas oryzae pv. oryzae, Xoo) resistance. qRT-PCR analysis showed that overexpression of TaCPK2-A in rice promoted the expression of OsWRKY45-1, a transcription factor involved in both fungal and bacterial resistance by regulating jasmonic acid and salicylic acid signalling genes. The opposite effect was found in wheat TaCPK2-A VIGS plants, where the homologue of OsWRKY45-1 was significantly repressed. These data suggest that modulation of WRKY45-1 and associated defence-response genes by CPK2 genes may be the common mechanism for multiple disease resistance in grass species, which may have undergone subfunctionalization in promoters before the formation of hexaploid wheat.

PMTED: a plant microRNA target expression database.

BACKGROUND: MicroRNAs (miRNAs) are identified in nearly all plants where they play important roles in development and stress responses by target mRNA cleavage or translation repression. MiRNAs exert their functions by sequence complementation with target genes and hence their targets can be predicted using bioinformatics algorithms. In the past two decades, microarray technology has been employed to study genes involved in important biological processes such as biotic response, abiotic response, and specific tissues and developmental stages, many of which are miRNA targets. Despite their value in assisting research work for plant biologists, miRNA target genes are difficult to access without pre-processing and assistance of necessary analytical and visualization tools because they are embedded in a large body of microarray data that are scattered around in public databases. DESCRIPTION: Plant MiRNA Target Expression Database (PMTED) is designed to retrieve and analyze expression profiles of miRNA targets represented in the plethora of existing microarray data that are manually curated. It provides a Basic Information query function for miRNAs and their target sequences, gene ontology, and differential expression profiles. It also provides searching and browsing functions for a global Meta-network among species, bioprocesses, conditions, and miRNAs, meta-terms curated from well annotated microarray experiments. Networks are displayed through a Cytoscape Web-based graphical interface. In addition to conserved miRNAs, PMTED provides a target prediction portal for user-defined novel miRNAs and corresponding target expression profile retrieval. Hypotheses that are suggested by miRNA-target networks should provide starting points for further experimental validation. CONCLUSIONS: PMTED exploits value-added microarray data to study the contextual significance of miRNA target genes and should assist functional investigation for both miRNAs and their targets. PMTED will be updated over time and is freely available for non-commercial use at http://pmted.agrinome.org.