WeiTsing, a pericycle-expressed ion channel, safeguards the stele to confer clubroot resistance.

Plant roots encounter numerous pathogenic microbes that often cause devastating diseases. One such pathogen, Plasmodiophora brassicae (Pb), causes clubroot disease and severe yield losses on cruciferous crops worldwide. Here, we report the isolation and characterization of WeiTsing (WTS), a broad-spectrum clubroot resistance gene from Arabidopsis. WTS is transcriptionally activated in the pericycle upon Pb infection to prevent pathogen colonization in the stele. Brassica napus carrying the WTS transgene displayed strong resistance to Pb. WTS encodes a small protein localized in the endoplasmic reticulum (ER), and its expression in plants induces immune responses. The cryoelectron microscopy (cryo-EM) structure of WTS revealed a previously unknown pentameric architecture with a central pore. Electrophysiology analyses demonstrated that WTS is a calcium-permeable cation-selective channel. Structure-guided mutagenesis indicated that channel activity is strictly required for triggering defenses. The findings uncover an ion channel analogous to resistosomes that triggers immune signaling in the pericycle.

Alternating between even and odd ploidy levels switches on and off the recombination control, even near the centromeres.

Meiotic recombination is a key biological process in plant evolution and breeding, as it generates genetic diversity in each generation through the formation of crossovers (COs). However, due to their importance in genome stability, COs are highly regulated in frequency and distribution. We previously demonstrated that this strict regulation of COs can be modified, both in terms of CO frequency and distribution, in allotriploid Brassica hybrids (2n = 3x = 29; AAC) resulting from a cross between Brassica napus (2n = 4x = 38; AACC) and Brassica rapa (2n = 2x = 20; AA). Using the recently updated B. napus genome now including pericentromeres, we demonstrated that COs occur in these cold regions in allotriploids, as close as 375 kb from the centromere. Reverse transcription quantitative PCR (RT-qPCR) of various meiotic genes indicated that Class I COs are likely involved in the increased recombination frequency observed in allotriploids. We also demonstrated that this modified recombination landscape can be maintained via successive generations of allotriploidy (odd ploidy level). This deregulated meiotic behavior reverts to strict regulation in allotetraploid (even ploidy level) progeny in the second generation. Overall, we provide an easy way to manipulate tight recombination control in a polyploid crop.

ASYNAPSIS3 has diverse dosage-dependent effects on meiotic crossover formation in Brassica napus.

Crossovers create genetic diversity and are required for equal chromosome segregation during meiosis. Crossover number and distribution are highly regulated by different mechanisms that are not yet fully understood, including crossover interference. The chromosome axis is crucial for crossover formation. Here, we explore the function of the axis protein ASYNAPSIS3. To this end, we use the allotetraploid species Brassica napus; due to its polyploid nature, this system allows a fine-grained dissection of the dosage of meiotic regulators. The simultaneous mutation of all 4 ASY3 alleles results in defective synapsis and drastic reduction of crossovers, which is largely rescued by the presence of only one functional ASY3 allele. Crucially, while the number of class I crossovers in mutants with 2 functional ASY3 alleles is comparable to that in wild type, this number is significantly increased in mutants with only one functional ASY3 allele, indicating that reducing ASY3 dosage increases crossover formation. Moreover, the class I crossovers on each bivalent in mutants with 1 functional ASY3 allele follow a random distribution, indicating compromised crossover interference. These results reveal the distinct dosage-dependent effects of ASY3 on crossover formation and provide insights into the role of the chromosome axis in patterning recombination.

Genome-scale targeted mutagenesis in Brassica napus using a pooled CRISPR library.

The recently constructed mutant libraries of diploid crops by the CRISPR-Cas9 system have provided abundant resources for functional genomics and crop breeding. However, because of the genome complexity, it is a big challenge to accomplish large-scale targeted mutagenesis in polyploid plants. Here, we demonstrate the feasibility of using a pooled CRISPR library to achieve genome-scale targeted editing in an allotetraploid crop of Brassica napus A total of 18,414 sgRNAs were designed to target 10,480 genes of interest, and afterward, 1104 regenerated transgenic plants harboring 1088 sgRNAs were obtained. Editing interrogation results revealed that 93 of the 178 genes were identified as mutated, thus representing an editing efficiency of 52.2%. Furthermore, we have discovered that Cas9-mediated DNA cleavages tend to occur at all the target sites guided by the same individual sgRNA, a novel finding in polyploid plants. Finally, we show the strong capability of reverse genetic screening for various traits with the postgenotyped plants. Several genes, which might dominate the fatty acid profile and seed oil content and have yet to be reported, were unveiled from the forward genetic studies. Our research provides valuable resources for functional genomics, elite crop breeding, and a good reference for high-throughput targeted mutagenesis in other polyploid plants.

Genome balance and dosage effect drive allopolyploid formation in Brassica.

Polyploidy is a major evolutionary force that has shaped plant diversity. However, the various pathways toward polyploid formation and interploidy gene flow remain poorly understood. Here, we demonstrated that the immediate progeny of allotriploid AAC Brassica (obtained by crossing allotetraploid Brassica napus and diploid Brassica rapa) was predominantly aneuploids with ploidal levels ranging from near-triploidy to near-hexaploidy, and their chromosome numbers deviated from the theoretical distribution toward increasing chromosome numbers, suggesting that they underwent selection. Karyotype and phenotype analyses showed that aneuploid individuals containing fewer imbalanced chromosomes had higher viability and fertility. Within three generations of self-fertilization, allotriploids mainly developed into near or complete allotetraploids similar to B. napus via gradually increasing chromosome numbers and fertility, suggesting that allotriploids could act as a bridge in polyploid formation, with aneuploids as intermediates. Self-fertilized interploidy hybrids ultimately generated new allopolyploids carrying different chromosome combinations, which may create a reproductive barrier preventing allotetraploidy back to diploidy and promote gene flow from diploids to allotetraploids. These results suggest that the maintenance of a proper genome balance and dosage drove the recurrent conversion of allotriploids to allotetraploids, which may contribute to the formation and evolution of polyploids.

The lack of negative association between TE load and subgenome dominance in synthesized Brassica allotetraploids.

Polyploidization is important to the evolution of plants. Subgenome dominance is a distinct phenomenon associated with most allopolyploids. A gene on the dominant subgenome tends to express to higher RNA levels in all organs as compared to the expression of its syntenic paralogue (homoeolog). The mechanism that underlies the formation of subgenome dominance remains unknown, but there is evidence for the involvement of transposon/DNA methylation density differences nearby the genes of parents as being causal. The subgenome with lower density of transposon and methylation near genes is positively associated with subgenome dominance. Here, we generated eight generations of allotetraploid progenies from the merging of parental genomes Brassica rapa and Brassica oleracea. We found that transposon/methylation density differ near genes between the parental (rapa:oleracea) existed in the wide hybrid, persisted in the neotetraploids (the synthetic Brassica napus), but these neotetraploids expressed no expected subgenome dominance. This absence of B. rapa vs. B. oleracea subgenome dominance is particularly significant because, while there is no negative relationship between transposon/methylation level and subgenome dominance in the neotetraploids, the more ancient parental subgenomes for all Brassica did show differences in transposon/methylation densities near genes and did express, in the same samples of cells, biased gene expression diagnostic of subgenome dominance. We conclude that subgenome differences in methylated transposon near genes are not sufficient to initiate the biased gene expressions defining subgenome dominance. Our result was unexpected, and we suggest a "nuclear chimera" model to explain our data.

Identification and characterization of transition metal-binding proteins and metabolites in the phloem sap of Brassica napus.

Transition metal (TM) distribution through the phloem is an essential part of plant metabolism and is required for systemic signaling and balancing source-to-sink relationships. Due to their reactivity, TMs are expected to occur in complexes within the phloem sap; however, metal speciation in the phloem sap remains largely unexplored. Here, we isolated phloem sap from Brassica napus and analyzed it via size exclusion chromatography coupled online to sector-field ICP-MS. Our data identified known TM-binding proteins and molecules including metallothioneins (MT), glutathione, and nicotianamine. While the main peak of all metals was low MW (∼1.5 kD), additional peaks ∼10 to 15 kD containing Cu, Fe, S, and Zn were also found. Further physicochemical analyses of MTs with and without affinity tags corroborated that MTs can form complexes of diverse molecular weights. We also identified and characterized potential artifacts in the TM-biding ability of B. napus MTs between tagged and non-tagged MTs. That is, the native BnMT2 binds Zn, Cu, and Fe, while MT3a and MT3b only bind Cu and Zn. In contrast, his-tagged MTs bind less Cu and were found to bind Co and Mn and aggregated to oligomeric forms to a greater extent compared to the phloem sap. Our data indicates that TM chemistry in the phloem sap is more complex than previously anticipated and that more systematic analyses are needed to establish the precise speciation of TM and TM-ligand complexes within the phloem sap.

Genome-wide association study reveals the genetic basis for petal-size formation in rapeseed (Brassica napus) and CRISPR-Cas9-mediated mutagenesis of BnFHY3 for petal-size reduction.

Petals in rapeseed (Brassica napus) serve multiple functions, including protection of reproductive organs, nutrient acquisition, and attraction of pollinators. However, they also cluster densely at the top, forming a thick layer that absorbs and reflects a considerable amount of photosynthetically active radiation. Breeding genotypes with large, small, or even petal-less varieties, requires knowledge of primary genes for allelic selection and manipulation. However, our current understanding of petal-size regulation is limited, and the lack of markers and pre-breeding materials hinders targeted petal-size breeding. Here, we conducted a genome-wide association study on petal size using 295 diverse accessions. We identified 20 significant single nucleotide polymorphisms and 236 genes associated with petal-size variation. Through a cross-analysis of genomic and transcriptomic data, we focused on 14 specific genes, from which molecular markers for diverging petal-size features can be developed. Leveraging CRISPR-Cas9 technology, we successfully generated a quadruple mutant of Far-Red Elongated Hypocotyl 3 (q-bnfhy3), which exhibited smaller petals compared to the wild type. Our study provides insights into the genetic basis of petal-size regulation in rapeseed and offers abundant potential molecular markers for breeding. The q-bnfhy3 mutant unveiled a novel role of FHY3 orthologues in regulating petal size in addition to previously reported functions.

Genomic asymmetric epigenetic modification of transposable elements is involved in gene expression regulation of allopolyploid Brassica napus.

Polyploids are common and have a wide geographical distribution and environmental adaptability. Allopolyploidy may lead to the activation of transposable elements (TE). However, the mechanism of epigenetic modification of TEs in the establishment and evolution of allopolyploids remains to be explored. We focused on the TEs of model allopolyploid Brassica napus (An An Cn Cn ), exploring the TE characteristics of the genome, epigenetic modifications of TEs during allopolyploidization, and regulation of gene expression by TE methylation. In B. napus, approximately 50% of the genome was composed of TEs. TEs increased with proximity to genes, especially DNA transposons. TE methylation levels were negatively correlated with gene expression, and changes in TE methylation levels were able to regulate the expression of neighboring genes related to responses to light intensity and stress, which promoted powerful adaptation of allopolyploids to new environments. TEs can be synergistically regulated by RNA-directed DNA methylation pathways and histone modifications. The epigenetic modification levels of TEs tended to be similar to those of the diploid parents during the genome evolution of B. napus. The TEs of the An subgenome were more likely to be modified, and the imbalance in TE number and epigenetic modification level in the An and Cn subgenomes may lead to the establishment of subgenome dominance. Our study analyzed the characteristics of TE location, DNA methylation, siRNA, and histone modification in B. napus and highlighted the importance of TE epigenetic modifications during the allopolyploidy process, providing support for revealing the mechanism of allopolyploid formation and evolution.

Trehalose-6-phosphate synthase 8 increases photosynthesis and seed yield in Brassica napus.

Trehalose-6-phosphate (T6P) functions as a vital proxy for assessing carbohydrate status in plants. While class II T6P synthases (TPS) do not exhibit TPS activity, they are believed to play pivotal regulatory roles in trehalose metabolism. However, their precise functions in carbon metabolism and crop yield have remained largely unknown. Here, BnaC02.TPS8, a class II TPS gene, is shown to be specifically expressed in mature leaves and the developing pod walls of Brassica napus. Overexpression of BnaC02.TPS8 increased photosynthesis and the accumulation of sugars, starch, and biomass compared to wild type. Metabolomic analysis of BnaC02.TPS8 overexpressing lines and CRISPR/Cas9 mutants indicated that BnaC02.TPS8 enhanced the partitioning of photoassimilate into starch and sucrose, as opposed to glycolytic intermediates and organic acids, which might be associated with TPS activity. Furthermore, the overexpression of BnaC02.TPS8 not only increased seed yield but also enhanced seed oil accumulation and improved the oil fatty acid composition in B. napus under both high nitrogen (N) and low N conditions in the field. These results highlight the role of class II TPS in impacting photosynthesis and seed yield of B. napus, and BnaC02.TPS8 emerges as a promising target for improving B. napus seed yield.

Nitrate alleviates ammonium toxicity in Brassica napus by coordinating rhizosphere and cell pH and ammonium assimilation.

In natural and agricultural situations, ammonium ( NH 4 + ) is a preferred nitrogen (N) source for plants, but excessive amounts can be hazardous to them, known as NH 4 + toxicity. Nitrate ( NO 3 - ) has long been recognized to reduce NH 4 + toxicity. However, little is known about Brassica napus, a major oil crop that is sensitive to high NH 4 + . Here, we found that NO 3 - can mitigate NH 4 + toxicity by balancing rhizosphere and intracellular pH and accelerating ammonium assimilation in B. napus. NO 3 - increased the uptake of NO 3 - and NH 4 + under high NH 4 + circumstances by triggering the expression of NO 3 - and NH 4 + transporters, while NO 3 - and H+ efflux from the cytoplasm to the apoplast was enhanced by promoting the expression of NO 3 - efflux transporters and genes encoding plasma membrane H+ -ATPase. In addition, NO 3 - increased pH in the cytosol, vacuole, and rhizosphere, and down-regulated genes induced by acid stress. Root glutamine synthetase (GS) activity was elevated by NO 3 - under high NH 4 + conditions to enhance the assimilation of NH 4 + into amino acids, thereby reducing NH 4 + accumulation and translocation to shoot in rapeseed. In addition, root GS activity was highly dependent on the environmental pH. NO 3 - might induce metabolites involved in amino acid biosynthesis and malate metabolism in the tricarboxylic acid cycle, and inhibit phenylpropanoid metabolism to mitigate NH 4 + toxicity. Collectively, our results indicate that NO 3 - balances both rhizosphere and intracellular pH via effective NO 3 - transmembrane cycling, accelerates NH 4 + assimilation, and up-regulates malate metabolism to mitigate NH 4 + toxicity in oilseed rape.

Photosynthetic plasticity aggravates the susceptibility of magnesium-deficient leaf to high light in rapeseed plants: the importance of Rubisco and mesophyll conductance.

Plants grown under low magnesium (Mg) soils are highly susceptible to encountering light intensities that exceed the capacity of photosynthesis (A), leading to a depression of photosynthetic efficiency and eventually to photooxidation (i.e., leaf chlorosis). Yet, it remains unclear which processes play a key role in limiting the photosynthetic energy utilization of Mg-deficient leaves, and whether the plasticity of A in acclimation to irradiance could have cross-talk with Mg, hence accelerating or mitigating the photodamage. We investigated the light acclimation responses of rapeseed (Brassica napus) grown under low- and adequate-Mg conditions. Magnesium deficiency considerably decreased rapeseed growth and leaf A, to a greater extent under high than under low light, which is associated with higher level of superoxide anion radical and more severe leaf chlorosis. This difference was mainly attributable to a greater depression in dark reaction under high light, with a higher Rubisco fallover and a more limited mesophyll conductance to CO2 (gm ). Plants grown under high irradiance enhanced the content and activity of Rubisco and gm to optimally utilize more light energy absorbed. However, Mg deficiency could not fulfill the need to activate the higher level of Rubisco and Rubisco activase in leaves of high-light-grown plants, leading to lower Rubisco activation and carboxylation rate. Additionally, Mg-deficient leaves under high light invested more carbon per leaf area to construct a compact leaf structure with smaller intercellular airspaces, lower surface area of chloroplast exposed to intercellular airspaces, and CO2 diffusion conductance through cytosol. These caused a more severe decrease in within-leaf CO2 diffusion rate and substrate availability. Taken together, plant plasticity helps to improve photosynthetic energy utilization under high light but aggravates the photooxidative damage once the Mg nutrition becomes insufficient.

Multi-omics-driven advances in the understanding of triacylglycerol biosynthesis in oil seeds.

Vegetable oils are rich sources of polyunsaturated fatty acids and energy as well as valuable sources of human food, animal feed, and bioenergy. Triacylglycerols, which are comprised of three fatty acids attached to a glycerol backbone, are the main component of vegetable oils. Here, we review the development and application of multiple-level omics in major oilseeds and emphasize the progress in the analysis of the biological roles of key genes underlying seed oil content and quality in major oilseeds. Finally, we discuss future research directions in functional genomics research based on current omics and oil metabolic engineering strategies that aim to enhance seed oil content and quality, and specific fatty acids components according to either human health needs or industrial requirements.

A complex receptor locus confers responsiveness to necrosis and ethylene-inducing like peptides in Brassica napus.

Brassica crops are susceptible to diseases which can be mitigated by breeding for resistance. MAMPs (microbe-associated molecular patterns) are conserved molecules of pathogens that elicit host defences known as pattern-triggered immunity (PTI). Necrosis and Ethylene-inducing peptide 1-like proteins (NLPs) are MAMPs found in a wide range of phytopathogens. We studied the response to BcNEP2, a representative NLP from Botrytis cinerea, and showed that it contributes to disease resistance in Brassica napus. To map regions conferring NLP response, we used the production of reactive oxygen species (ROS) induced during PTI across a population of diverse B. napus accessions for associative transcriptomics (AT), and bulk segregant analysis (BSA) on DNA pools created from a cross of NLP-responsive and non-responsive lines. In silico mapping with AT identified two peaks for NLP responsiveness on chromosomes A04 and C05 whereas the BSA identified one peak on A04. BSA delimited the region for NLP-responsiveness to 3 Mbp, containing ~245 genes on the Darmor-bzh reference genome and four co-segregating KASP markers were identified. The same pipeline with the ZS11 genome confirmed the highest-associated region on chromosome A04. Comparative BLAST analysis revealed unannotated clusters of receptor-like protein (RLP) homologues on ZS11 chromosome A04. However, no specific RLP homologue conferring NLP response could be identified. Our results also suggest that BR-SIGNALLING KINASE1 may be involved with modulating the NLP response. Overall, we demonstrate that responsiveness to NLP contributes to disease resistance in B. napus and define the associated genomic location. These results can have practical application in crop improvement.

Defects in Glabrous 3 (GL3) functionality underlie the absence of trichomes in Brassica napus.

Previously, expression of the Arabidopsis thaliana GLABRA3 (GL3) induced trichome formation in Brassica napus. GL3 orthologues were examined from glabrous (B. oleracea), semi-glabrous (B. napus), moderately hirsute (B. rapa), and very hirsute (B. villosa) Brassica species. Ectopic expression of BnGL3, BrGL3 alleles, or BvGL3 induced trichome formation in glabrous B. napus with the effect on trichome number commensurate with density in the original accessions. Chimeric GL3 proteins in which the B. napus amino terminal region, which interacts with MYB proteins, or the middle region, which interacts with the WD40 protein TTG1, was exchanged with corresponding regions from A. thaliana were as stimulatory to trichome production as AtGL3. Exchange of the carboxy-terminal region containing a bHLH domain and an ACT domain did not alter the trichome stimulatory activity, although modeling of the ACT domain identified differences that could affect GL3 dimerization. B. napus A- and C-genomes orthologues differed in their abilities to form homo- and heterodimers. Modeling of the amino-terminal region revealed a conserved domain that may represent the MYB factor binding pocket. This region interacted with the MYB factors GL1, CPC, and TRY, as well as with JAZ8, which is involved in jasmonic acid-mediated regulation of MYC-like transcription factors. Protein interaction studies indicated that GL1 interaction with GL3 from B. napus and A. thaliana may underlie the difference in their respective abilities to induce trichome formation.

Knockout of stigmatic ascorbate peroxidase 1 (APX1) delays pollen rehydration and germination by mediating ROS homeostasis in Brassica napus L.

The successful interaction between pollen and stigma is a critical process for plant sexual reproduction, involving a series of intricate molecular and physiological events. After self-compatible pollination, a significant reduction in reactive oxygen species (ROS) production has been observed in stigmas, which is essential for pollen grain rehydration and subsequent pollen tube growth. Several scavenging enzymes tightly regulate ROS homeostasis. However, the potential role of these ROS-scavenging enzymes in the pollen-stigma interaction in Brassica napus remains unclear. Here, we showed that the activity of ascorbate peroxidase (APX), an enzyme that plays a crucial role in the detoxification of hydrogen peroxide (H2O2), was modulated depending on the compatibility of pollination in B. napus. We then identified stigma-expressed APX1s and generated pentuple mutants of APX1s using CRISPR/Cas9 technology. After compatible pollination, the BnaAPX1 pentuple mutants accumulated higher levels of H2O2 in the stigma, while the overexpression of BnaA09.APX1 resulted in lower levels of H2O2. Furthermore, the knockout of BnaAPX1 delayed the compatible response-mediated pollen rehydration and germination, which was consistent with the effects of a specific APX inhibitor, ρ-Aminophenol, on compatible pollination. In contrast, the overexpression of BnaA09.APX1 accelerated pollen rehydration and germination after both compatible and incompatible pollinations. However, delaying and promoting pollen rehydration and germination did not affect the seed set after compatible and incompatible pollination in APX1 pentuple mutants and overexpression lines, respectively. Our results demonstrate the fundamental role of BnaAPX1 in pollen rehydration and germination by regulating ROS homeostasis during the pollen-stigma interaction in B. napus.

BnaABF3 and BnaMYB44 regulate the transcription of zeaxanthin epoxidase genes in carotenoid and abscisic acid biosynthesis.

Zeaxanthin epoxidase (ZEP) is a key enzyme that catalyzes the conversion of zeaxanthin to violaxanthin in the carotenoid and abscisic acid (ABA) biosynthesis pathways. The rapeseed (Brassica napus) genome has 4 ZEP (BnaZEP) copies that are suspected to have undergone subfunctionalization, yet the 4 genes' underlying regulatory mechanisms remain unknown. Here, we genetically confirmed the functional divergence of the gene pairs BnaA09.ZEP/BnaC09.ZEP and BnaA07.ZEP/BnaC07.ZEP, which encode enzymes with tissue-specific roles in carotenoid and ABA biosynthesis in flowers and leaves, respectively. Molecular and transgenic experiments demonstrated that each BnaZEP pair is transcriptionally regulated via ABA-responsive element-binding factor 3 s (BnaABF3s) and BnaMYB44s as common and specific regulators, respectively. BnaABF3s directly bound to the promoters of all 4 BnaZEPs and activated their transcription, with overexpression of individual BnaABF3s inducing BnaZEP expression and ABA accumulation under drought stress. Conversely, loss of BnaABF3s function resulted in lower expression of several genes functioning in carotenoid and ABA metabolism and compromised drought tolerance. BnaMYB44s specifically targeted and repressed the expression of BnaA09.ZEP/BnaC09.ZEP but not BnaA07.ZEP/BnaC07.ZEP. Overexpression of BnaA07.MYB44 resulted in increased carotenoid content and an altered carotenoid profile in petals. Additionally, RNA-seq analysis indicated that BnaMYB44s functions as a repressor in phenylpropanoid and flavonoid biosynthesis. These findings provide clear evidence for the subfunctionalization of duplicated genes and contribute to our understanding of the complex regulatory network involved in carotenoid and ABA biosynthesis in B. napus.

A module involving HIGH LEAF TEMPERATURE1 controls instantaneous water use efficiency.

Drought stress inhibits plant growth and agricultural production. Improving plant instantaneous water use efficiency (iWUE), which is strictly regulated by stomata, is an effective way to cope with drought stress. However, the mechanisms of iWUE regulation are poorly understood. Through genetic screening for suppressors of mpk12-4, an Arabidopsis (Arabidopsis thaliana) mutant with a major iWUE quantitative trait locus gene MITOGEN-ACTIVATED PROTEIN KINASE12 deleted, we identified HIGH LEAF TEMPERATURE1 (HT1). Genetic interaction and physiological analyses showed that MPK12 controls iWUE through multiple modules in a high CO2-induced stomatal closing pathway that regulate SLOW ANION CHANNEL-ASSOCIATED1 (SLAC1) activity. HT1 acts downstream of MPK12, whereas OPEN STOMATA1 (OST1) and GUARD CELL HYDROGEN PEROXIDE-RESISTANT1 (GHR1) function downstream of HT1 by activating SLAC1 in iWUE. Photosynthetic-CO2 response curves and biomass analyses under different water-supply conditions showed that HT1 dysfunction improved iWUE and also increased plant growth capacity, and products of HT1 putative orthologs from Brassica (Brassica napus) and rice (Oryza sativa) exhibited functions similar to that of Arabidopsis HT1 in iWUE and the CO2-signaling pathway. Our study revealed the mechanism of MPK12-mediated iWUE regulation in Arabidopsis and provided insight into the internal relationship between iWUE and CO2 signaling in guard cells and a potential target for improving crop iWUE and drought tolerance.

Winter warming post floral initiation delays flowering via bud dormancy activation and affects yield in a winter annual crop.

Winter annual life history is conferred by the requirement for vernalization to promote the floral transition and control the timing of flowering. Here we show using winter oilseed rape that flowering time is controlled by inflorescence bud dormancy in addition to vernalization. Winter warming treatments given to plants in the laboratory and field increase flower bud abscisic acid levels and delay flowering in spring. We show that the promotive effect of chilling reproductive tissues on flowering time is associated with the activity of two FLC genes specifically silenced in response to winter temperatures in developing inflorescences, coupled with activation of a BRANCHED1-dependent bud dormancy transcriptional module. We show that adequate winter chilling is required for normal inflorescence development and high yields in addition to the control of flowering time. Because warming during winter flower development is associated with yield losses at the landscape scale, our work suggests that bud dormancy activation may be important for effects of climate change on winter arable crop yields.

BABY BOOM regulates early embryo and endosperm development.

The BABY BOOM (BBM) AINTEGUMENTA-LIKE (AIL) AP2/ERF domain transcription factor is a major regulator of plant cell totipotency, as it induces asexual embryo formation when ectopically expressed. Surprisingly, only limited information is available on the role of BBM during zygotic embryogenesis. Here we reexamined BBM expression and function in the model plant Arabidopsis thaliana (Arabidopsis) using reporter analysis and newly developed CRISPR mutants. BBM was expressed in the embryo from the zygote stage and also in the maternal (nucellus) and filial (endosperm) seed tissues. Analysis of CRISPR mutant alleles for BBM (bbm-cr) and the redundantly acting AIL gene PLETHORA2 (PLT2) (plt2-cr) uncovered individual roles for these genes in the timing of embryo progression. We also identified redundant roles for BBM and PLT2 in endosperm proliferation and cellularization and the maintenance of zygotic embryo development. Finally, we show that ectopic BBM expression in the egg cell of Arabidopsis and the dicot crops Brassica napus and Solanum lycopersicon is sufficient to bypass the fertilization requirement for embryo development. Together these results highlight roles for BBM and PLT2 in seed development and demonstrate the utility of BBM genes for engineering asexual embryo development in dicot species.