Graph-Based Pangenome of Actinidia chinensis Reveals Structural Variations Mediating Fruit Degreening.

Fruit ripening is associated with the degreening process (loss of chlorophyll) that occurs in most fruit species. Kiwifruit is one of the special species whose fruits may maintain green flesh by accumulating a large amount of chlorophyll even after ripening. However, little is known about the genetic variations related to the fruit degreening process. Here, a graph-based kiwifruit pangenome by analyzing 14 chromosome-scale haplotype-resolved genome assemblies from seven representative cultivars or lines in Actinidia chinensis is built. A total of 49,770 non-redundant gene families are identified, with core genes constituting 46.6%, and dispensable genes constituting 53.4%. A total of 84,591 non-redundant structural variations (SVs) are identified. The pangenome graph integrating both reference genome sequences and variant information facilitates the identification of SVs related to fruit color. The SV in the promoter of the AcBCM gene determines its high expression in the late developmental stage of fruits, which causes chlorophyll accumulation in the green-flesh fruits by post-translationally regulating AcSGR2, a key enzyme of chlorophyll catabolism. Taken together, a high-quality pangenome is constructed, unraveled numerous genetic variations, and identified a novel SV mediating fruit coloration and fruit quality, providing valuable information for further investigating genome evolution and domestication, QTL genes function, and genomics-assisted breeding.

An endogenous promoter LpSUT2 discovered in duckweed: a promising transgenic tool for plants.

Promoters are one of the most critical elements in regulating gene expression. They are considered essential biotechnological tools for heterologous protein production. The one most widely used in plants is the 35S promoter from cauliflower mosaic virus. However, our study for the first time discovered the 35S promoter reduced the expression of exogenous proteins under increased antibiotic stress. We discovered an endogenous strong promoter from duckweed named LpSUT2 that keeps higher initiation activity under antibiotic stress. Stable transformation in duckweed showed that the gene expression of eGFP in the LpSUT2:eGFP was 1.76 times that of the 35S:eGFP at 100 mg.L-1 G418 and 6.18 times at 500 mg.L-1 G418. Notably, with the increase of G418 concentration, the gene expression and the fluorescence signal of eGFP in the 35S:eGFP were weakened, while the LpSUT2:eGFP only changed slightly. This is because, under high antibiotic stress, the 35S promoter was methylated, leading to the gene silencing of the eGFP gene. Meanwhile, the LpSUT2 promoter was not methylated and maintained high activity. This is a previously unknown mechanism that provides us with new insights into screening more stable promoters that are less affected by environmental stress. These outcomes suggest that the LpSUT2 promoter has a high capacity to initiate the expression of exogenous proteins. In conclusion, our study provides a promoter tool with potential application for plant genetic engineering and also provides new insights into screening promoters.

Haplotype-resolved genome assembly provides insights into evolutionary history of the Actinidia arguta tetraploid.

Actinidia arguta, known as hardy kiwifruit, is a widely cultivated species with distinct botanical characteristics such as small and smooth-fruited, rich in beneficial nutrients, rapid softening and tolerant to extremely low temperatures. It contains the most diverse ploidy types, including diploid, tetraploid, hexaploid, octoploid, and decaploid. Here we report a haplotype-resolved tetraploid genome (A. arguta cv. 'Longcheng No.2') containing four haplotypes, each with 40,859, 41,377, 39,833 and 39,222 protein-coding genes. We described the phased genome structure, synteny, and evolutionary analyses to identify and date possible WGD events. Ks calculations for both allelic and paralogous genes pairs throughout the assembled haplotypic individuals showed its tetraploidization is estimated to have formed ~ 1.03 Mya following Ad-α event occurred ~ 18.7 Mya. Detailed annotations of NBS-LRRs or CBFs highlight the importance of genetic variations coming about after polyploidization in underpinning ability of immune responses or environmental adaptability. WGCNA analysis of postharvest quality indicators in combination with transcriptome revealed several transcription factors were involved in regulating ripening kiwi berry texture. Taking together, the assembly of an A. arguta tetraploid genome provides valuable resources in deciphering complex genome structure and facilitating functional genomics studies and genetic improvement for kiwifruit and other crops.

Agrobacterium rhizogenes-mediated marker-free transformation and gene editing system revealed that AeCBL3 mediates the formation of calcium oxalate crystal in kiwifruit.

The transformation and gene editing of the woody species kiwifruit are difficult and time-consuming. The fast and marker-free genetic modification system for kiwifruit has not been developed yet. Here, we establish a rapid and efficient marker-free transformation and gene editing system mediated by Agrobacterium rhizogenes for kiwifruit. Moreover, a removing-root-tip method was developed to significantly increase the regeneration efficiency of transgenic hairy roots. Through A. rhizogenes-mediated CRISPR/Cas9 gene editing, the editing efficiencies of CEN4 and AeCBL3 achieved 55 and 50%, respectively. And several homozygous knockout lines for both genes were obtained. Our method has been successfully applied in the transformation of two different species of kiwifruit (Actinidia chinensis 'Hongyang' and A.eriantha 'White'). Next, we used the method to study the formation of calcium oxalate (CaOx) crystals in kiwifruit. To date, little is known about how CaOx crystal is formed in plants. Our results indicated that AeCBL3 overexpression enhanced CaOx crystal formation, but its knockout via CRISPR/Cas9 significantly impaired crystal formation in kiwifruit. Together, we developed a fast maker-free transformation and highly efficient CRISPR-Cas9 gene editing system for kiwifruit. Moreover, our work revealed a novel gene mediating CaOx crystal formation and provided a clue to elaborate the underlying mechanisms.

Genome-Wide Identification of HSF Gene Family in Kiwifruit and the Function of AeHSFA2b in Salt Tolerance.

Heat shock transcription factors (HSFs) play a crucial role in regulating plant growth and response to various abiotic stresses. In this study, we conducted a comprehensive analysis of the AeHSF gene family at genome-wide level in kiwifruit (Actinidia eriantha), focusing on their functions in the response to abiotic stresses. A total of 41 AeHSF genes were identified and categorized into three primary groups, namely, HSFA, HSFB, and HSFC. Further transcriptome analysis revealed that the expression of AeHSFA2b/2c and AeHSFB1c/1d/2c/3b was strongly induced by salt, which was confirmed by qRT-PCR assays. The overexpression of AeHSFA2b in Arabidopsis significantly improved the tolerance to salt stress by increasing AtRS5, AtGolS1 and AtGolS2 expression. Furthermore, yeast one-hybrid, dual-luciferase, and electrophoretic mobility shift assays demonstrated that AeHSFA2b could bind to the AeRFS4 promoter directly. Therefore, we speculated that AeHSFA2b may activate AeRFS4 expression by directly binding its promoter to enhance the kiwifruit's tolerance to salt stress. These results will provide a new insight into the evolutionary and functional mechanisms of AeHSF genes in kiwifruit.

Stress-induced endocytosis from chloroplast inner envelope membrane is mediated by CHLOROPLAST VESICULATION but inhibited by GAPC.

Clathrin-mediated vesicular formation and trafficking are responsible for molecular cargo transport and signal transduction among organelles. Our previous study shows that CHLOROPLAST VESICULATION (CV)-containing vesicles (CVVs) are generated from chloroplasts for chloroplast degradation under abiotic stress. Here, we show that CV interacts with the clathrin heavy chain (CHC) and induces vesicle budding toward the cytosol from the chloroplast inner envelope membrane. In the defective mutants of CHC2 and the dynamin-encoding DRP1A, CVV budding and releasing from chloroplast are impeded. The mutations of CHC2 inhibit CV-induced chloroplast degradation and hypersensitivity to water stress. Moreover, CV-CHC2 interaction is impaired by the oxidized GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPC). GAPC1 overexpression suppresses CV-mediated chloroplast degradation and hypersensitivity to water stress, while CV silencing alleviates the hypersensitivity of the gapc1gapc2 plant to water stress. Together, our work identifies a pathway of clathrin-assisted CVV budding outward from chloroplast, which is involved in chloroplast degradation and stress response.

Comparative Transcriptomic Analysis Revealed the Suppression and Alternative Splicing of Kiwifruit (Actinidia latifolia) NAP1 Gene Mediating Trichome Development.

Kiwifruit (Actinidia chinensis) is commonly covered by fruit hairs (trichomes) that affect kiwifruit popularity in the commercial market. However, it remains largely unknown which gene mediates trichome development in kiwifruit. In this study, we analyzed two kiwifruit species, A. eriantha (Ae) with long, straight, and bushy trichomes and A. latifolia (Al) with short, distorted, and spare trichomes, by second- and third-generation RNA sequencing. Transcriptomic analysis indicated that the expression of the NAP1 gene, a positive regulator of trichome development, was suppressed in Al compared with that in Ae. Additionally, the alternative splicing of AlNAP1 produced two short transcripts (AlNAP1-AS1 and AlNAP1-AS2) lacking multiple exons, in addition to a full-length transcript of AlNAP1-FL. The defects of trichome development (short and distorted trichome) in Arabidopsis nap1 mutant were rescued by AlNAP1-FL but not by AlNAP1-AS1. AlNAP1-FL gene does not affect trichome density in nap1 mutant. The qRT-PCR analysis indicated that the alternative splicing further reduces the level of functional transcripts. These results indicated that the short and distorted trichomes in Al might be caused by the suppression and alternative splicing of AlNAP1. Together, we revealed that AlNAP1 mediates trichome development and is a good candidate target for genetic modification of trichome length in kiwifruit.

Telomere-to-telomere and gap-free reference genome assembly of the kiwifruit Actinidia chinensis.

Kiwifruit is an economically and nutritionally important fruit crop with extremely high contents of vitamin C. However, the previously released versions of kiwifruit genomes all have a mass of unanchored or missing regions. Here, we report a highly continuous and completely gap-free reference genome of Actinidia chinensis cv. 'Hongyang', named Hongyang v4.0, which is the first to achieve two de novo haploid-resolved haplotypes, HY4P and HY4A. HY4P and HY4A have a total length of 606.1 and 599.6 Mb, respectively, with almost the entire telomeres and centromeres assembled in each haplotype. In comparison with Hongyang v3.0, the integrity and contiguity of Hongyang v4.0 is markedly improved by filling all unclosed gaps and correcting some misoriented regions, resulting in ~38.6-39.5 Mb extra sequences, which might affect 4263 and 4244 protein-coding genes in HY4P and HY4A, respectively. Furthermore, our gap-free genome assembly provides the first clue for inspecting the structure and function of centromeres. Globally, centromeric regions are characterized by higher-order repeats that mainly consist of a 153-bp conserved centromere-specific monomer (Ach-CEN153) with different copy numbers among chromosomes. Functional enrichment analysis of the genes located within centromeric regions demonstrates that chromosome centromeres may not only play physical roles for linking a pair of sister chromatids, but also have genetic features for participation in the regulation of cell division. The availability of the telomere-to-telomere and gap-free Hongyang v4.0 reference genome lays a solid foundation not only for illustrating genome structure and functional genomics studies but also for facilitating kiwifruit breeding and improvement.

Cell Wall Matrix Polysaccharides Contribute to Salt-Alkali Tolerance in Rice.

Salt-alkali stress threatens the resilience to variable environments and thus the grain yield of rice. However, how rice responds to salt-alkali stress at the molecular level is poorly understood. Here, we report isolation of a novel salt-alkali-tolerant rice (SATR) by screening more than 700 germplasm accessions. Using 93-11, a widely grown cultivar, as a control, we characterized SATR in response to strong salt-alkali stress (SSAS). SATR exhibited SSAS tolerance higher than 93-11, as indicated by a higher survival rate, associated with higher peroxidase activity and total soluble sugar content but lower malonaldehyde accumulation. A transcriptome study showed that cell wall biogenesis-related pathways were most significantly enriched in SATR relative to 93-11 upon SSAS. Furthermore, higher induction of gene expression in the cell wall matrix polysaccharide biosynthesis pathway, coupled with higher accumulations of hemicellulose and pectin as well as measurable physio-biochemical adaptive responses, may explain the strong SSAS tolerance in SATR. We mapped SSAS tolerance to five genomic regions in which 35 genes were candidates potentially governing SSAS tolerance. The 1,4-ß-D-xylan synthase gene OsCSLD4 in hemicellulose biosynthesis pathway was investigated in details. The OsCSLD4 function-disrupted mutant displayed reduced SSAS tolerance, biomass and grain yield, whereas the OsCSLD4 overexpression lines exhibited increased SSAS tolerance. Collectively, this study not only reveals the potential role of cell wall matrix polysaccharides in mediating SSAS tolerance, but also highlights applicable value of OsCSLD4 and the large-scale screening system in developing SSAS-tolerant rice.

Tea GOLDEN2-LIKE genes enhance catechin biosynthesis through activating R2R3-MYB transcription factor.

The biosynthesis of catechins, a major type of flavonoids accumulated in tea, is mediated by developmental cues and environmental stimuli. Light enhances but shading treatment reduces catechin accumulation in tea leaves. However, the transcription factors involved in light-mediated catechin biosynthesis remain to be identified. Two GOLDEN2 LIKE genes from tea plant (CsGLK1 and CsGLK2) were isolated and characterized in both tomato and tea plants. Transcripts of both CsGLK1 and CsGLK2 were affected by light intensity in tea plants. Overexpression of CsGLK1 and CsGLK2 promoted chloroplast development and carotenoid accumulation in tomato fruits. An integrated metabolomic and transcriptomic approach revealed that both catechin content and related biosynthetic genes were upregulated in CsGLK-overexpressing tomato leaves. Our further studies in tea plants indicated that CsGLKs directly regulate the transcription of CsMYB5b, a transcription factor involved in catechin biosynthesis. Suppression of CsGLKs in tea leaves led to the reduction of both CsMYB5b expression and catechin accumulation. Taken together, the results show that CsGLKs are involved in light-regulated catechin accumulation in tea plants by regulating expression of CsMYB5b and have great potential for enhancing the accumulation of both carotenoids and flavonoids in fruits of horticultural crops.

Genome-Wide Expression Profiling Analysis of Kiwifruit GolS and RFS Genes and Identification of AcRFS4 Function in Raffinose Accumulation.

The raffinose synthetase (RFS) and galactinol synthase (GolS) are two critical enzymes for raffinose biosynthesis, which play an important role in modulating plant growth and in response to a variety of biotic or abiotic stresses. Here, we comprehensively analyzed the RFS and GolS gene families and their involvement in abiotic and biotic stresses responses at the genome-wide scale in kiwifruit. A total of 22 GolS and 24 RFS genes were identified in Actinidia chinensis and Actinidia eriantha genomes. Phylogenetic analysis showed that the GolS and RFS genes were clustered into four and six groups, respectively. Transcriptomic analysis revealed that abiotic stresses strongly induced some crucial genes members including AcGolS1/2/4/8 and AcRFS2/4/8/11 and their expression levels were further confirmed by qRT-PCR. The GUS staining of AcRFS4Pro::GUS transgenic plants revealed that the transcriptionlevel of AcRFS4 was significantly increased by salt stress. Overexpression of AcRFS4 in Arabidopsis demonstrated that this gene enhanced the raffinose accumulation and the tolerance to salt stress. The co-expression networks analysis of hub transcription factors targeting key AcRFS4 genes indicated that there was a strong correlation between AcNAC30 and AcRFS4 expression under salt stress. Furthermore, the yeast one-hybrid assays showed that AcNAC30 could bind the AcRFS4 promoter directly. These results may provide insights into the evolutionary and functional mechanisms of GolS and RFS genes in kiwifruit.

Application of developmental regulators to improve in planta or in vitro transformation in plants.

Plant genetic transformation is a crucial step for applying biotechnology such as genome editing to basic and applied plant science research. Its success primarily relies on the efficiency of gene delivery into plant cells and the ability to regenerate transgenic plants. In this study, we have examined the effect of several developmental regulators (DRs), including PLETHORA (PLT5), WOUND INDUCED DEDIFFERENTIATION 1 (WIND1), ENHANCED SHOOT REGENERATION (ESR1), WUSHEL (WUS) and a fusion of WUS and BABY-BOOM (WUS-P2A-BBM), on in planta transformation through injection of Agrobacterium tumefaciens in snapdragons (Antirrhinum majus). The results showed that PLT5, WIND1 and WUS promoted in planta transformation of snapdragons. An additional test of these three DRs on tomato (Solanum lycopersicum) further demonstrated that the highest in planta transformation efficiency was observed from PLT5. PLT5 promoted calli formation and regeneration of transformed shoots at the wound positions of aerial stems, and the transgene was stably inherited to the next generation in snapdragons. Additionally, PLT5 significantly improved the shoot regeneration and transformation in two Brassica cabbage varieties (Brassica rapa) and promoted the formation of transgenic calli and somatic embryos in sweet pepper (Capsicum annum) through in vitro tissue culture. Despite some morphological alternations, viable seeds were produced from the transgenic Bok choy and snapdragons. Our results have demonstrated that manipulation of PLT5 could be an effective approach for improving in planta and in vitro transformation efficiency, and such a transformation system could be used to facilitate the application of genome editing or other plant biotechnology application in modern agriculture.

EGY3 mediates chloroplastic ROS homeostasis and promotes retrograde signaling in response to salt stress in Arabidopsis.

The chloroplast is the main organelle for stress-induced production of reactive oxygen species (ROS). However, how chloroplastic ROS homeostasis is maintained under salt stress is largely unknown. We show that EGY3, a gene encoding a chloroplast-localized protein, is induced by salt and oxidative stresses. The loss of EGY3 function causes stress hypersensitivity while EGY3 overexpression increases the tolerance to both salt and chloroplastic oxidative stresses. EGY3 interacts with chloroplastic Cu/Zn-SOD2 (CSD2) and promotes CSD2 stability under stress conditions. In egy3-1 mutant plants, the stress-induced CSD2 degradation limits H2O2 production in chloroplasts and impairs H2O2-mediated retrograde signaling, as indicated by the decreased expression of retrograde-signal-responsive genes required for stress tolerance. Both exogenous application of H2O2 (or APX inhibitor) and CSD2 overexpression can rescue the salt-stress hypersensitivity of egy3-1 mutants. Our findings reveal that EGY3 enhances the tolerance to salt stress by promoting the CSD2 stability and H2O2-mediated chloroplastic retrograde signaling.

Characterization of the 'Oat-Like Rice' Caused by a Novel Allele OsMADS1Olr Reveals Vital Importance of OsMADS1 in Regulating Grain Shape in Oryza sativa L.

BACKGROUND: Grain shape is a critical agronomic trait affecting grain yield and quality. Exploration and functional characterization of grain shape-related genes will facilitate rice breeding for higher quality and yield. RESULTS: Here, we characterized a recessive mutant named Oat-like rice for its unique grain shape which highly resembles oat grains. The Oat-like rice displayed abnormal floral organs, an open hull formed by remarkably elongated leafy lemmas and paleae, occasionally formed conjugated twin brown rice, an aberrant grain shape and a low seed setting rate. By map-based cloning, we discovered that Oat-like rice harbors a novel allele of OsMADS1 gene (OsMADS1Olr), which has a spontaneous point mutation that causes the substitution of an amino acid that is highly conserved in the MADS-box domain of the MADS-box family. Further linkage analysis indicated that the point mutation in the OsMADS1Olr is associated with Oat-like rice phenotype, and expression analysis of the OsMADS1 by qRT-PCR and GUS staining also indicated that it is highly expressed in flower organs as well as in the early stages of grain development. Furthermore, OsMADS1Olr-overexpressing plants showed similar phenotypes of Oat-like rice in grain shape, possibly due to the dominant negative effect. And OsMADS1-RNAi plants also displayed grain phenotypes like Oat-like rice. These results suggested that OsMADS1Olr is responsible for the Oat-like rice phenotype including aberrant grain shape. Moreover, the expression levels of representative genes related to grain shape regulation were apparently altered in Oat-like rice, OsMADS1Olr-overexpressing and OsMADS1-RNAi transgenic plants. Finally, compared with Oat-like rice, OsMADS1Olr-overexpressing and OsMADS1-RNAi plants, mild phenotype of seed-specific OsMADS1-RNAi transgenic plants indicated that OsMADS1 may has has a direct regulation role in grain development and the grain phenotypes of Oat-like rice, OsMADS1Olr-overexpressing and OsMADS1-RNAi plants are majorly caused by the abnormal lemma and palea development. CONCLUSIONS: Altogether, our results showed that grain shape and a low seed setting rate of the notable 'Oat-like rice' are caused by a spontaneous point mutation in the novel allele OsMADS1Olr. Furthermore, our findings suggested that OsMADS1 mediates grain shape possibly by affecting the expression of representative genes related to grain shape regulation. Thus, this study not only revealed that OsMADS1 plays a vital role in regulating grain shape of rice but also highlighted the importance and value of OsMADS1 to improve the quality and yield of rice by molecular breeding.

Proteomic Analysis of Rice Subjected to Low Light Stress and Overexpression of OsGAPB Increases the Stress Tolerance.

BACKGROUND: Light provides the energy for photosynthesis and determines plant morphogenesis and development. Low light compromises photosynthetic efficiency and leads to crop yield loss. It remains unknown how rice responds to low light stress at a proteomic level. RESULTS: In this study, the quantitative proteomic analysis with isobaric tags for relative and absolute quantitation (iTRAQ) was used and 1221 differentially expressed proteins (DEPs) were identified from wild type rice plants grown in control or low light condition (17% light intensity of control), respectively. Bioinformatic analysis of DEPs indicated low light remarkably affects the abundance of chloroplastic proteins. Specifically, the proteins involved in carbon fixation (Calvin cycle), electron transport, and ATPase complex are severely downregulated under low light. Furthermore, overexpression of the downregulated gene encoding rice ß subunit of glyceraldehyde-3-phosphate dehydrogenase (OsGAPB), an enzyme in Calvin cycle, significantly increased the CO2 assimilation rate, chlorophyll content and fresh weight under low light conditions but have no obvious effect on rice growth and development under control light. CONCLUSION: Our results revealed that low light stress on vegetative stage of rice inhibits photosynthesis possibly by decreasing the photosynthetic proteins and OsGAPB gene is a good candidate for manipulating rice tolerance to low light stress.

The cloning and characterization of hypersensitive to salt stress mutant, affected in quinolinate synthase, highlights the involvement of NAD in stress-induced accumulation of ABA and proline.

Nicotinamide adenine dinucleotide (NAD), a ubiquitous coenzyme, is required for many physiological reactions and processes. However, it remains largely unknown how NAD affects plant response to salt stress. We isolated a salt-sensitive mutant named hypersensitive to salt stress (hss) from an ethyl methanesulfonate-induced mutation population. A point mutation was identified by MutMap in the encoding region of Quinolinate Synthase (QS) gene required for the de novo synthesis of NAD. This point mutation caused a substitution of amino acid in the highly-conserved NadA domain of QS, resulting in an impairment of NAD biosynthesis in the mutant. Molecular and chemical complementation have restored the response of the hss mutant to salt stress, indicating that the decreased NAD contents in the mutant were responsible for its hypersensitivity to salt stress. Furthermore, the endogenous levels of abscisic acid (ABA) and proline were also reduced in stress-treated hss mutant. The application of ABA or proline could alleviate stress-induced oxidative damage of the mutant and partially rescue its hypersensitivity to salt stress, but not affect NAD concentration. Taken together, our results demonstrated that the NadA domain of QS is important for NAD biosynthesis, and NAD participates in plant response to salt stress by affecting stress-induced accumulation of ABA and proline.

Unraveling microbial community diversity and succession of Chinese Sichuan sausages during spontaneous fermentation by high-throughput sequencing.

The microbial community diversity and succession of Chinese Sichuan sausages during the spontaneous fermentation were demonstrated using high-throughput sequencing technology. The bacterial diversity was abundant and the succession of bacterial community along the direction of Lactobacillus spp. increased and Weissella spp. decreased. While fungal diversity was single and trace fungal population was detected. The core functional microbiota were lactic acid bacteria, including Lactobacillus spp., Weissella spp. and Pediococcus spp. In initial fermentation, Weissella spp. was the dominant bacteria and its relative abundance was 49.84%, but then its relative abundance decreased to 11.96% during fermentation before recovering to 26.74% at the end of fermentation. Meanwhile, Lactobacillus spp. rose from 24.70 to 55.74% and became the dominant genus. Moreover, Pediococcus spp. increased from 0.06 to 18.05% on day 20 but then decreased to 1.89% on day 30. These results revealed that the primary microorganisms contributing to spontaneous fermentation of Chinese Sichuan sausages were bacteria, while eukaryotic microorganisms such as yeast scarcely contributed to fermentation.

Natural variation in the HAN1 gene confers chilling tolerance in rice and allowed adaptation to a temperate climate.

Rice (Oryza sativa L.) is a chilling-sensitive staple crop that originated in subtropical regions of Asia. Introduction of the chilling tolerance trait enables the expansion of rice cultivation to temperate regions. Here we report the cloning and characterization of HAN1, a quantitative trait locus (QTL) that confers chilling tolerance on temperate japonica rice. HAN1 encodes an oxidase that catalyzes the conversion of biologically active jasmonoyl-L-isoleucine (JA-Ile) to the inactive form 12-hydroxy-JA-Ile (12OH-JA-Ile) and fine-tunes the JA-mediated chilling response. Natural variants in HAN1 diverged between indica and japonica rice during domestication. A specific allele from temperate japonica rice, which gained a putative MYB cis-element in the promoter of HAN1 during the divergence of the two japonica ecotypes, enhances the chilling tolerance of temperate japonica rice and allows it to adapt to a temperate climate. The results of this study extend our understanding of the northward expansion of rice cultivation and provide a target gene for the improvement of chilling tolerance in rice.

Tomato SlAN11 regulates flavonoid biosynthesis and seed dormancy by interaction with bHLH proteins but not with MYB proteins.

The flavonoid compounds are important secondary metabolites with versatile human nutritive benefits and fulfill a multitude of functions during plant growth and development. The abundance of different flavonoid compounds are finely tuned with species-specific pattern by a ternary MBW complex, which consists of a MYB, a bHLH, and a WD40 protein, but the essential role of SlAN11, which is a WD40 protein, is not fully understood in tomato until now. In this study, a tomato WD40 protein named as SlAN11 was characterized as an effective transcription regulator to promote plant anthocyanin and seed proanthocyanidin (PA) contents, with late flavonoid biosynthetic genes activated in 35S::SlAN11 transgenic lines, while the dihydroflavonol flow to the accumulation of flavonols or their glycosylated derivatives was reduced by repressing the expression of SlFLS in this SlAN11-overexpressed lines. The above changes were reversed in 35S::SlAN11-RNAi transgenic lines except remained levels of flavonol compounds and SlFLS expression. Interestingly, our data revealed that SlAN11 gene could affect seed dormancy by regulating the expressions of abscisic acid (ABA) signaling-related genes SlABI3 and SlABI5, and the sensitivity to ABA treatment in seed germination is conversely changed by SlAN11-overexpressed or -downregulated lines. Yeast two-hybrid assays demonstrated that SlAN11 interacted with bHLH but not with MYB proteins in the ternary MBW complex, whereas bHLH interacted with MYB in tomato. Our results indicated that low level of anthocyanins in tomato fruits, with low expression of bHLH (SlTT8) and MYB (SlANT1 and SlAN2) genes, remain unchanged upon modification of SlAN11 gene alone in the transgenic lines. These results suggest that the tomato WD40 protein SlAN11, coordinating with bHLH and MYB proteins, plays a crucial role in the fine adjustment of the flavonoid biosynthesis and seed dormancy in tomato.

Tomato UV-B receptor SlUVR8 mediates plant acclimation to UV-B radiation and enhances fruit chloroplast development via regulating SlGLK2.

Plants utilize energy from sunlight to perform photosynthesis in chloroplast, an organelle that could be damaged by solar UV radiation. The ultraviolet-B (UV-B) photoreceptor UVR8 is required for UV-B perception and signal transduction. However, little is known about how UVR8 influence chloroplast development under UV-B radiation. Here, we characterized tomato UVR8 gene (SlUVR8) and our results indicated that SlUVR8 facilitate plant acclimation to UV-B stress by orchestrating expression of the UVB-responsive genes (HY5 and CHS) and accumulating UV-absorptive compounds. In addition, we also discovered that SlUVR8 promotes fruit chloroplast development through enhancing accumulation of transcription factor GOLDEN2-LIKE2 (SlGLK2) which determines chloroplast and chlorophyll levels. Furthermore, UV-B radiation could increase expression of SlGLK2 and its target genes in fruits and leaves. SlUVR8 is required for UVB-induced SlGLK2 expression. Together, our work not only identified the conserved functions of SlUVR8 gene in response to UV-B stress, but also uncovered a novel role that SlUVR8 could boost chloroplast development by accumulating SlGLK2 proteins.