Biology subjects

Dubcovsky, J.

Publications and source records attributed to Dubcovsky, J..

At least 19 recordsLinked to original sources

bioRxiv2026

A genetic toolkit to reduce wheat immunogenicity and incidence of celiac disease

Celiac disease (CeD) is an immune-mediated condition triggered by wheat gluten in genetically predisposed individuals. The immune reaction in people with CeD is driven by particular gluten amino acid sequences, or immunogenic epitopes. Some of these epitopes elicit strong immune responses in the majority of CeD patients and are designated as immunodominant epitopes. Previous research has shown correlations between the amount of immunogenic wheat epitopes consumed and the onset of CeD, suggesting that reducing wheat immunogenic epitopes may reduce CeD incidence at the population level. Gluten consists of gliadins and glutenins, with gliadins having the majority of the immunodominant epitopes and glutenins playing a major role in dough strength and breadmaking quality (BMQ). This study used radiation-induced deletions, chemical mutagenesis, and natural variation in wheat (Triticum aestivum) to generate genetic stocks with reduced immunogenic epitope content. Most lines were developed in the wheat cultivar Summit, for which we produced a full genome assembly and annotation. We used exome capture to characterize these deletions and identify prolamins located within and outside the deletions. We combined different deletions and developed molecular markers to facilitate their deployment. For chromosome arms 1BS and 1DS, we generated two alternative lines: one lacking immunogenic epitopes for the development of CeD-safe genetic stocks for research purposes, and another retaining selected glutenins for breeding commercial lines with reduced immunogenicity and adequate BMQ. By making these non-transgenic genetic stocks publicly available, we aim to accelerate the development of wheat varieties with reduced immunogenicity and, eventually, a fully CeD-safe wheat.

plant biology↗

bioRxiv2026

Regulation of spikelet number during wheat spike development

Wheat produces unbranched inflorescences (spikes) composed of smaller inflorescences (spikelets) as their fundamental building units. The spikelet number per spike (SNS) is a major determinant of grain yield and the gene networks that regulate this trait are the focus of this review. Spikelet development starts with the transition of the shoot apical meristem into an inflorescence meristem (IM) that produces lateral spikelet meristems (SMs). The rate at which SMs are produced and the timing of the IM transition into a terminal spikelet (IM[->]TS) determine the final SNS. These two traits are regulated by genes expressed in the IM (e.g. meristem identity genes), as well as by the amount of FLOWERING LOCUS T1 (florigen) transported from leaves to developing spikes. Spikelet number can also be increased by the production of spikes with supernumerary spikelets (SS) or branch-like structures that resemble small spikes. Mutations that promote a reversion from SM to IM identity can induce the formation of SS or branches. Initial efforts to incorporate these mutations into commercial wheat varieties have faced trade-offs in fertility and grain weight, which will require additional research and breeding efforts. Meanwhile, genes and allele combinations that increase SNS without affecting the number of spikelets per node have been identified and are being deployed in wheat breeding programs. Recent spatial transcriptomics, single-cell analyses, and multi-omics studies of wheat spike development are accelerating the discovery of new genes affecting SNS and enhancing our ability to engineer more productive wheat spikes.

plant biology↗

bioRxiv2025

MicroRNA156 and its targeted SPL genes interact with the photoperiod, vernalization, and gibberellin pathways to regulate wheat heading time

Heading time has a large impact on adaptation to different environments and crop productivity. In this study, we characterized the effect of the endogenous age pathway on heading time and its interactions with the photoperiod and vernalization pathways in the leaves of tetraploid wheat (Triticum turgidum ssp. durum). Plants with reduced levels of microRNA156 or increased expression of its downstream targets, the SQUAMOSA PROMOTER BINDING PROTEIN-LIKE genes SPL3, SPL4, and SPL13 exhibited accelerated heading time, with stronger effects under suboptimal inductive conditions. Earlier heading was associated with the upregulation of miR172 and flowering-promoting genes VRN1, FUL2, and FT1 and the downregulation of flowering-repressing genes AP2L1 and VRN2. Additionally, we uncovered complex interactions among SPL, SQUAMOSA (VRN1 and FUL2) and DELLA proteins that modulate wheat heading time. We showed that DELLA proteins, which are negative regulators in the gibberellic acid pathway, can interact with SPL proteins reducing their ability to induce flowering. We also discovered previously unknown interactions between SQUAMOSA and DELLA proteins in wheat that compete with the DELLA-SPL interactions, likely reducing DELLAs ability to repress SPL3 and SPL4 activity. Since SPL3 and SPL4 directly promote VRN1 and FUL2 transcription, these interactions generate a positive regulatory feedback loop that accelerates wheat heading time. Finally, we developed dominant miR156-resistant alleles rSPL3, rSPL4, and rSPL13 that accelerate wheat heading time under both optimal and suboptimal inductive conditions. These publicly available genetic resources can be used to fine tune heading time and improve wheat adaptation to changing environments. Significance statementWheat heading time is critical for adaptation to diverse environments. We generated dominant mutations for the SPL3, SPL4 and SPL13 genes that accelerate heading time. Different combinations of these mutations can be used to modulate heading time and improve wheat adaptation to changing environments.

plant biology↗

bioRxiv2025

Spatial and single-cell expression analyses reveal complex expression domains in early wheat spike development

Wheat is important for global food security and understanding the molecular mechanisms driving spike and spikelet development can inform the development of more productive varieties. In this study, we integrated single-molecule fluorescence in situ hybridization (smFISH) and single-cell RNA sequencing (scRNA-seq) to generate an atlas of cell clusters and expression domains during the early stages of wheat spike development. We characterized spatiotemporal expression of 99 genes by smFISH in 48,225 cells at the early transition, late double ridge, and floret primordia stages. These cells were grouped into 21 different expression domains, including four in the basal region of the developing spikelets and three different meristematic regions, which were consistent across spikelets and sections. Using induced mutants, we revealed functional roles associated with the specific expression patterns of LFY in intercalary meristems, SPL14 in inflorescence meristems, and FZP in glume axillae. Complementary scRNA-seq profiling of 26,009 cells from W2.5 and W3.5 stages identified 23 distinct cell clusters. We used the scRNA-seq information to impute the expression of 74,464 genes into the spatially anchored smFISH-labelled cells and generated a public website to visualize them. We used experimental and imputed expression profiles, together with co-expression studies and correlation matrices, to annotate the scRNA-seq clusters. From co-expression analyses, we uncovered genes associated with boundary genes TCP24 and FZP, as well as the meristematic genes AGL6 and ULT1. The smFISH and scRNA-seq studies provided complementary tools for dissecting gene networks that regulate spike development and identifying new co-expressed genes for functional characterization.

plant biology↗

bioRxiv2024

Natural alleles of LEAFY and WAPO1 interact to regulate spikelet number per spike in wheat

Spikelet number per spike (SNS) is an important yield component in wheat that determines the maximum number of grains that can be formed in a wheat spike. In wheat, loss-of-function mutations in LEAFY (LFY) or its interacting protein WHEAT ORTHOLOG OF APO1 (WAPO1) significantly reduce SNS by reducing the rate of formation of spikelet meristems. In previous studies, we identified a natural amino acid change in WAPO1 (C47F) that significantly increases SNS in hexaploid wheat. In this study, we searched for natural variants in LFY that were associated with differences in SNS, and detected significant effects in the LFY-B region in a nested association mapping population. We generated a large mapping population and confirmed that the LFY-B polymorphism R80S is linked with the differences in SNS, suggesting that LFY-B is the likely causal gene. A haplotype analysis revealed two amino acid changes P34L and R80S, which were both enriched during wheat domestication and breeding suggesting positive selection. We also explored the interactions between the LFY and WAPO1 natural variants using biparental populations and identified significant interaction, in which the positive effect of the 80S and 34L alleles from LFY-B was only detected in the WAPO-A1 47F background but not in the 47C background. Based on these results we propose that the allele combination WAPO-A1-47F / LFY-B 34L 80S can be used in wheat breeding programs to maximize SNS and increase grain yield potential in wheat. Key messageSpecific combinations of LFY and WAPO1 natural alleles maximize spikelet number per spike in wheat.

plant biology↗

bioRxiv2024

Manipulation of the microRNA172 - AP2L2 interaction provides precise control of wheat and triticale plant height

The REDUCED HEIGHT (RHT) dwarfing alleles Rht-B1b and Rht-D1b were essential in the "Green Revolution" to optimize wheat plant height and increase grain yield. However, those alleles reduce coleoptile length limiting sowing depth, which triggered the search for alternative dwarfing genes. In this study, we engineered the interaction between miR172 and AP2L2 genes to fine-tune wheat and triticale plant height without affecting coleoptile and first-leaf length.

plant biology↗

bioRxiv2024

Deletion of wheat alpha-gliadins from chromosome 6D improves gluten strength and reduces immunodominant celiac disease epitopes

Wheat gliadins and glutenins confer valuable end-use characteristics but include amino acid sequences (epitopes) that can elicit celiac disease (CeD) in genetically predisposed individuals. The onset of CeD in these individuals is affected by the amount and duration of the exposure to immunogenic epitopes. Therefore, a reduction of epitopes that result in high immune responses in the majority of CeD patients (immunodominant epitopes) can reduce the incidence of CeD at a population level. We identified deletions encompassing the -gliadins at the three wheat genomes, designated hereafter as{Delta} gli-A2 (PI 704906),{Delta} gli-B2 (PI 704907), and{Delta} gli-D2 (PI 704908). The{Delta} gli-D2 deletion, which eliminates major immunodominant epitopes, significantly increases gluten strength, improves breadmaking quality, and has no negative effects on grain yield or grain protein content. By contrast,{Delta} gli-A2 and{Delta} gli-B2 showed limited effects on breadmaking quality. The stronger effect of the{Delta} gli-D2 deletion on gluten strength is associated with the presence of -gliadins with seven cysteines in GLI-D2 that are absent in GLI-A2 and GLI-B2 loci, which all have -gliadins with six cysteines. We show that -gliadins with seven cysteines are incorporated into the gluten polymer, where they likely function as chain-terminators limiting the expansion of the gluten polymer and reducing its strength. In summary, the publicly available{Delta} gli-D2 deletion developed in this study can be used to simultaneously improve wheat gluten strength and reduce immunodominant CeD epitopes.

plant biology↗

bioRxiv2023

LEAFY interacts with WHEAT ORTHOLOG OF APO1 to regulate spikelet number per spike in wheat

In wheat, the transition of the inflorescence meristem to a terminal spikelet (IM[->]TS) determines the spikelet number per spike (SNS), an important yield component. In this study, we demonstrate that the plant-specific transcription factor LEAFY (LFY) physically and genetically interacts with WHEAT ORTHOLOG OF APO1 (WAPO1) to regulate SNS and floret development. Loss-of-function mutations in either or both genes result in significant and similar reductions in SNS, as a result of a reduction in the rate of spikelet meristems formation per day. SNS is also modulated by significant genetic interactions between LFY and SQUAMOSA MADS-box genes VRN1 and FUL2, which promote the IM[->]TS transition. Single-molecule fluorescence in-situ hybridization revealed a down-regulation of LFY and up-regulation of the SQUAMOSA MADS-box genes in the distal part of the developing spike during the IM[->]TS transition, supporting their opposite roles in the regulation of SNS in wheat. Concurrently, the overlap of LFY and WAPO1 transcription domains in the developing spikelets contributes to normal floret development. Understanding the genetic network regulating SNS is a necessary first step to engineer this important agronomic trait. SUMMARY STATEMENTThe plant specific transcription factor LEAFY plays an important role in the regulation of the number of spikelets per spike in wheat.

plant biology↗

bioRxiv2023

Wheat bZIPC1 interacts with FT2 and contributes to the regulation of spikelet number per spike

Loss-of-function mutations and natural variation in the gene FLOWERING LOCUS T2 (FT2) in wheat have previously been shown to affect spikelet number per spike (SNS). However, while other FT-like wheat proteins interact with bZIP-containing transcription factors from the A-group, FT2 does not interact with any of them. In this study, we used a yeast-two-hybrid screen with FT2 as bait and identified a grass-specific bZIP-containing transcription factor from the C-group, designated here as bZIPC1. Within the C-group, we identified four clades including wheat proteins that show Y2H interactions with different sets of FT-like and CEN-like encoded proteins. bZIPC1 and FT2 expression partially overlap in the developing spike, including the inflorescence meristem. Combined loss-of-function mutations in bZIPC-A1 and bZIPC-B1 (bzipc1) in tetraploid wheat resulted in a drastic reduction in SNS with a limited effect on heading date. Analysis of natural variation in the bZIPC-B1 (TraesCS5B02G444100) region revealed three major haplotypes (H1-H3), with the H1 haplotype showing significantly higher SNS, grain number per spike and grain weight per spike than both the H2 and H3 haplotypes. The favorable effect of the H1 haplotype was also supported by its increased frequency from the ancestral cultivated tetraploids to the modern durum and common wheat varieties. We developed markers for the two non-synonymous SNPs that differentiate the bZIPC-B1b allele in the H1 haplotype from the ancestral bZIPC-B1a allele present in all other haplotypes. These diagnostic markers are useful tools to accelerate the deployment of the favorable bZIPC-B1b allele in pasta and bread wheat breeding programs. Key MessageThe wheat transcription factor bZIPC1 interacts with FT2 and affects spikelet and grain number per spike. We identified a natural allele with positive effects on these two economically important traits.

plant biology↗

bioRxiv2023

Sequencing 4.3 million mutations in wheat promoters to understand and modify gene expression

Wheat is an important contributor to global food security, and further improvements are required to feed a growing human population. New functional genetics and genomics tools can help us to understand the function of different genes and to engineer beneficial changes. In this study, we used a promoter capture assay to sequence 2-kb regions upstream of all high-confidence annotated genes from 1,513 mutagenized plants from the tetraploid wheat variety Kronos. We identified 4.3 million induced mutations with an accuracy of 99.8%, resulting in a mutation density of 41.9 mutations per kb. We also remapped Kronos exome capture reads to Chinese Spring RefSeq v1.1, identified 4.7 million mutations, and predicted their effects on annotated genes. Using these predictions, we identified 59% more non-synonymous substitutions and 49% more truncation mutations than in the original study. To show the biological value of the new promoter dataset, we selected two mutations within the promoter of the VRN-A1 vernalization gene. Both mutations, located within transcription factor binding sites, significantly altered VRN-A1 expression, and one reduced the number of spikelets per spike. These publicly available sequenced mutant datasets provide rapid and inexpensive access to induced variation in the promoters and coding regions of most wheat genes. These mutations can be used to understand and modulate gene expression and phenotypes for both basic and commercial applications, where limited governmental regulations can facilitate deployment. These mutant collections, together with gene editing, provide valuable tools to accelerate functional genetic studies in this economically important crop. Significance StatementWe sequenced 4.3 million induced mutations in the promoters and 4.7 million in the coding regions of most wheat genes. We also show how this public resource can be used to understand gene function, modulate gene expression, and generate changes in valuable wheat agronomic traits.

genomics↗

bioRxiv2023

GIGANTEA accelerates wheat heading time through gene interactions converging on FLOWERING LOCUS T1

Precise regulation of flowering time is critical for cereal crops to synchronize reproductive development with optimum environmental conditions, thereby maximizing grain yield. The plant specific gene GIGANTEA (GI) plays an important role in the control of flowering time, with additional functions on the circadian clock and plant stress responses. In this study, we show that GI loss-of-function mutants in a photoperiod sensitive tetraploid wheat background exhibit significant delays in heading time under both long-day (LD) and short-day (SD) photoperiods, with stronger effects under LD. However, this interaction between GI and photoperiod is no longer observed in isogenic lines carrying either a photoperiod insensitive allele in the PHOTOPERIOD1 (PPD1) gene or a loss-of-function allele in EARLY FLOWERING 3 (ELF3), a known repressor of PPD1. These results suggest that the normal circadian regulation of PPD1 is required for the differential effect of GI on heading time in different photoperiods. Using crosses between mutants or transgenic of GI and those of critical genes in the flowering regulation pathway, we show that GI accelerates wheat heading time by promoting FLOWERING LOCUS T1 (FT1) expression via interactions with ELF3, VERNALIZATION 2 (VRN2), CONSTANS (CO), and the age-dependent microRNA172-APETALA2 (AP2) pathway, at both transcriptional and protein levels. Our study reveals conserved GI mechanisms between wheat and Arabidopsis, but also identifies specific interactions of GI with the distinctive photoperiod and vernalization pathways of the temperate grasses. These results provide valuable knowledge for modulating wheat heading time and engineering new varieties better adapted to a changing environment.

plant biology↗

bioRxiv2023

Wheat plant height locus RHT25 encodes a PLATZ transcription factor that interacts with DELLA (RHT1)

Plant height is an important agronomic trait with a significant impact on grain yield, as demonstrated by the positive effect of the REDUCED HEIGHT (RHT) dwarfing alleles (Rht1b) on lodging and harvest index in the "Green Revolution" wheat varieties. However, these gibberellic acid (GA) insensitive alleles also reduce coleoptile length, biomass production, and yield potential in some environments, triggering the search for alternative GA-sensitive dwarfing genes. Here we report the identification, validation and characterization of the gene underlying the GA-sensitive dwarfing locus RHT25 in wheat. This gene, designated as PLATZ-A1 (TraesCS6A02G156600), is expressed mainly in the elongating stem and developing spike and encodes a plant-specific AT-rich sequence- and zinc-binding protein (PLATZ). Natural and induced loss-of-function mutations in PLATZ-A1 reduce plant height and its over-expression increases it, demonstrating that PLATZ-A1 is the causative gene of RHT25. PLATZ-A1 interacts physically and genetically with RHT1 (DELLA), and both genes have stronger effects on plant height in the presence of the wildtype than in the presence of the mutant allele of the other gene. These results suggest that PLATZ1 can modulate the effect of DELLA on wheat plant height. We identified four natural truncation mutations and one promoter insertion in PLATZ-A1 that are more frequent in modern varieties than in landraces, suggesting positive selection during wheat breeding. These mutations can be used to fine-tune wheat plant height and, in combination with other GA-sensitive dwarfing genes, to replace the GA-insensitive Rht1b alleles to search for grain yield improvements beyond those of the Green Revolution varieties. Significance StatementWe have identified and characterized a previously unknown gene controlling plant height in wheat and named it PLATZ1. Mutations in PLATZ1 reduce plant height while its overexpression results in taller plants. PLATZ1 is expressed mainly in elongating stems and developing spikes and interacts physically and genetically with the "Green Revolution" dwarfing gene REDUCED HEIGHT 1 (RHT1). We discovered five natural mutants in the A genome copy of PLATZ1 in common wheat that have been favored during breeding, suggesting an overall positive effect on wheat performance. These mutations can be used to fine-tune wheat plant height and, eventually, to replace the RHT1 dwarfing alleles that impose limitations on planting depth and grain yield potential in some environments.

plant biology↗

bioRxiv2022

PHYTOCHROME C regulation of PHOTOPERIOD1 is mediated by EARLY FLOWERING 3 in Brachypodium distachyon

Daylength sensing in many plants is critical for coinciding the timing of flowering with the appropriate season. Temperate-climate-adapted grasses such as Brachypodium distachyon flower during the spring when days are becoming longer. The photoreceptor PHYTOCHROME C is essential for long-day (LD) flowering in B. distachyon. PHYC is required for the LD activation of a suite of genes in the photoperiod pathway including PHOTOPERIOD1 (PPD1) that, in turn, result in the activation of FLOWERING LOCUS T (FT1)/FLORIGEN, which causes flowering. Thus, phyC mutants are extremely delayed in flowering. Here we show that PHYC-mediated activation of PPD1 occurs via EARLY FLOWERING 3 (ELF3), a component of the evening complex in the circadian clock. The extreme delay of flowering of the phyC mutant disappears when combined with an elf3 loss-of-function mutation. Moreover, the dampened PPD1 expression in phyC mutant plants is elevated in phyC/elf3 mutant plants consistent with the rapid flowering of the double mutant. We show that loss of PPD1 function also results in reduced FT1 expression levels and extremely delayed flowering consistent with reports from wheat and barley. Additionally, elf3 mutant plants have elevated expression levels of PPD1 and we show that overexpression of ELF3 results in delayed flowering, which is associated with a reduction of PPD1 and FT1, demonstrating ELF3 represses PPD1 transcription, consistent with previous studies showing that ELF3 binds to the PPD1 promoter. Indeed, PPD1 is the main target of ELF3-mediated flowering as elf3/ppd1 double mutant plants are delayed flowering. Our results indicate that ELF3 operates downstream from PHYC and acts as a repressor of PPD1 in the photoperiod flowering pathway of B. distachyon. AUTHOR SUMMARYDaylength is an important environmental cue that plants and animals use to coincide important life history events with a proper season. In plants, timing of flowering to a particular season is an essential adaptation to many ecological niches. Perceiving changes in daylength starts with the perception of light via specific photoreceptors such as phytochromes. In temperate grasses, how daylength perception is integrated into downstream pathways to trigger flowering is not fully understood. However, some of the components involved in the translation of daylength perception into the induction of flowering in temperate grasses have been identified from studies of natural variation. For example, specific alleles of two genes called EARLY FLOWERING 3 (ELF3) and PHOTOPERIOD1 (PPD1) have been selected during breeding of different wheat and barley varieties to modulate the photoperiodic response to maximize reproduction in different environments. Here, we show in the temperate grass model Brachypodium distachyon that the translation of the light signal perceived by phytochromes into a flowering response is mediated by ELF3, and that PPD1 is genetically downstream of ELF3 in the photoperiodic flowering pathway. These results provide a genetic framework for understanding the photoperiodic response in temperate grasses that include agronomically important crops such as wheat, oats, barley, and rye.

plant biology↗

bioRxiv2022

EARLY FLOWERING 3 interactions with PHYTOCHROME B and PHOTOPERIOD1 are critical for the photoperiodic regulation of wheat heading time

The photoperiodic response is critical for plants to adjust their reproductive phase to the most favorable season. Wheat heads earlier under long days (LD) than under short days (SD) and this difference is mainly regulated by the PHOTOPERIOD1 (PPD1) gene. Tetraploid wheat plants carrying the Ppd-A1a allele with a large deletion in the promoter head earlier under SD than plants carrying the wildtype Ppd-A1b allele with an intact promoter. Phytochromes PHYB and PHYC are necessary for the light activation of PPD1, and mutations in either of these genes result in the downregulation of PPD1 and very late heading time. We show here that both effects are reverted when the phyB mutant is combined with loss-of-function mutations in EARLY FLOWERING 3 (ELF3), a component of the Evening Complex (EC) in the circadian clock. We also show that the wheat ELF3 protein interacts with PHYB and PHYC, is rapidly modified by light, and binds to the PPD1 promoter in planta (likely as part of the EC). Deletion of the ELF3 binding region in the Ppd-A1a promoter results in its upregulation at dawn, similar to PPD1 alleles with intact promoters in the elf3 mutant background. The upregulation of PPD1 is correlated with the upregulation of the florigen gene FLOWERING LOCUS T1 (FT1) and early heading time. Loss-of-function mutations in ppd1 result in the downregulation of FT1 and delayed heading, even when combined with the elf3 mutation. Taken together, these results indicate that ELF3 operates downstream of PHYB as a direct transcriptional repressor of PPD1, and that this repression is relaxed both by light and by the deletion of the ELF3 binding region in the Ppd-A1a promoter. In summary, the regulation of the light mediated activation of PPD1 by ELF3 is critical for the photoperiodic regulation of wheat heading time. Author SummaryThe coordination of reproductive development with the optimal season for seed production is critical to maximize grain yield in crop species. Plants can perceive the length of the day or night (photoperiod) and use this information to anticipate seasonal changes. In most eudicot plants, CONSTANS plays a central role in the perception of photoperiod, but in wheat the main photoperiod gene is PHOROPERIOD1 (PPD1). In this study, we show that the clock gene EARLY FLOWERING 3 (ELF3) regulates the phytochrome-mediated light activation of PPD1. Loss-of-function mutations in ELF3 result in the upregulation of PPD1 at and dawn, and in early heading under both long and short days, even in the absence of PHYB. A deletion in the PPD1 promoter including an ELF3 binding region also results in earlier heading under short days, indicating that ELF3 acts as a direct transcriptional repressor of PPD1. This study shows that ELF3 plays a critical role in the wheat photoperiod pathway by regulating the light signal between the phytochromes and PPD1. ELF3 provides an additional entry point to engineer heading time in wheat, an important trait for the development of better adapted varieties to a changing environment.

plant biology↗

bioRxiv2022

A second generation capture panel for cost-effective sequencing of genome regulatory regions in wheat and relatives

As genome resources for wheat expand at a rapid pace, it is important to update targeted sequencing tools to incorporate improved sequence assemblies and regions of previously unknown significance. Here, we developed an updated regulatory region enrichment capture for wheat (Triticum L.) and other Triticeae species. The core target space includes sequences from two Kbp upstream of each gene predicted in the Chinese Spring wheat genome (IWGSC RefSeq Annotation v1.1) and regions of open chromatin identified with ATAC-seq from wheat leaf and root samples. To improve specificity, we aggressively filtered candidate repetitive sequences using a combination of BLASTN searches to the Triticeae Repetitive Sequence Database (TREP), identification of regions with read over-coverage from previous target enrichment experiments, and k-mer frequency analyses. The final design comprises 216 Mbp of predicted hybridization space in hexaploid wheat and showed increased specificity and coverage of targeted sequences relative to previous protocols. Test captures on hexaploid and tetraploid wheat and other diploid cereals show that the assay has broad potential utility for cost-effective promoter and open chromatin resequencing and general-purpose genotyping of various Triticeae species.

genomics↗

bioRxiv2022

The combination of morphogenic regulators BABY BOOM and GRF-GIF improves maize transformation efficiency

Transformation is an indispensable tool for plant genetics and functional genomic studies. Although stable transformation no longer represents a major technology bottleneck in maize, there is still need for easily accessible and efficient transformation methods in most academic labs. Here we present the GGB transformation system, a rapid and highly efficient transformation system optimized for the immature embryo transformation of two maize genetic backgrounds, including the inbred line B104. The combination of distinct morphogenetic factors, the maize BABY BOOM transcriptional regulator (ZmBBM/EREB53) and the wheat GRF4-GIF1 (GROWTH REGULATING FACTOR4 - GRF-INTERACTING FACTOR1) chimera, together with a modified QuickCorn protocol, regenerated transformed maize seedlings in approximately two months with an efficiency of 26 to 37%; notably, the efficiency was 7-fold higher than with using either component in isolation. Additionally, ectopic expression of both morphogenetic factors did not show obvious effects on B104 development, and in particular fertility was not affected, obviating the need to remove the morphogenetic regulators post Agrobacterium infections. The GGB transformation system is designed for CRISPR-Cas9 editing but can be adapted for other purposes and should be easy to implement in most academic labs with little transformation experience.

plant biology↗

bioRxiv2022

Dosage differences in 12-OXOPHYTODIENOATE REDUCTASE genes modulate wheat primary root growth

Wheat is an essential crop for global food security and is well adapted to a wide variety of soils1. However, the gene networks regulating different root architectures remain poorly understood. We report here the identification of a cluster of a monocot-specific 12-OXOPHYTODIENOATE REDUCTASE genes from subfamily III (OPRIII) that modulate key differences in wheat root architecture associated with grain yield under water-limited conditions. Wheat plants with a loss-of-function mutation in OPRIII showed longer seminal roots, whereas plants with increased OPRIII dosage or transgenic over-expression showed reduced seminal root growth, precocious development of lateral roots and increased jasmonic acid (JA). A JA-biosynthesis inhibitor eliminated the root differences, confirming a JA-mediated mechanism. Multiple transcriptome analysis of transgenic and wild-type lines revealed significant enriched JA-biosynthetic and reactive oxygen species (ROS) pathways that paralleled changes in ROS distribution. The OPRIII genes provide a useful entry point to engineer root architecture in wheat and other cereals.

plant biology↗

bioRxiv2022

Transcriptional signatures of wheat inflorescence development

In order to maintain global food security, it will be necessary to increase yields of the cereal crops that provide most of the calories and protein for the worlds population, which includes common wheat (Triticum aestivum L.). An important factor contributing to wheat yield is the number of grain-holding spikelets which form on the spike during inflorescence development. Characterizing the gene regulatory networks controlling the timing and rate of inflorescence development will facilitate the selection of natural and induced gene variants that contribute to increased spikelet number and yield. In the current study, co-expression and gene regulatory networks were assembled from a temporal wheat spike transcriptome dataset, revealing the dynamic expression profiles associated with the progression from vegetative meristem to terminal spikelet formation. Consensus co-expression networks revealed enrichment of several transcription factor families at specific developmental stages including the sequential activation of different classes of MIKC-MADS box genes. This gene regulatory network highlighted interactions among a small number of regulatory hub genes active during terminal spikelet formation. Finally, the CLAVATA and WUSCHEL gene families were investigated, revealing potential roles for TaCLE13, TaWOX2, and TaWOX7 in wheat meristem development. The hypotheses generated from these datasets and networks further our understanding of wheat inflorescence development.

plant biology↗