Dwarf Phenotype Research Articles

Tissue-Specific Regulation of Vesicular Trafficking Mediated by Rab-GEF Complex MON1/CCZ1 From Solanum chilense Increases Salt Stress Tolerance in Arabidopsis thaliana.

Salt stress constrains the development and growth of plants. To tolerate it, mechanisms of endocytosis and vacuolar compartmentalization of Na+ are induced. In this work, the genes that encode a putative activator of vesicular trafficking called MON1/CCZ1 from Solanum chilense, SchMON1 and SchCCZ1, were co-expressed in roots of Arabidopsis thaliana to determine whether the increase in prevacuolar vesicular trafficking also increases the Na+ compartmentalization capacity and tolerance. Initially, we demonstrated that both SchMON1 and SchCCZ1 genes rescued the dwarf phenotype of both A. thaliana mon1-1 and ccz1a/b mutants associated with the loss of function, and both proteins colocalized with their functional targets, RabF and RabG, in endosomes. Transgenic A. thaliana plants co-expressing these genes improved salt stress tolerance compared to wild type plants, with SchMON1 contributing the most. At the sub-cellular level, co-expression of SchMON1/SchCCZ1 reduced ROS levels and increased endocytic activity, and number of acidic structures associated with autophagosomes. Notably, greater Na+ accumulation in vacuoles of cortex and endodermis was evidenced in the SchMON1 genotype. Molecular analysis of gene expression in each genotype supported these results. Altogether, our analysis shows that root activation of prevacuolar vesicular trafficking mediated by MON1/CCZ1 emerges as a promising physiological molecular mechanism to increase tolerance to salt stress in crops of economic interest.

The inheritance pattern for the dwarf phenotype in hybrids from crosses among sunflower lines differing in alleles of the Rht1 locus

Background. Sunflower seed production is based on utilization of the heterosis effect, manifesting itself in improving both yield and plant height in hybrids. Short-stemmed lines need to be used to develop commercial hybrids with an optimum height. Molecular bases of the trait manifestation in the dwarf lines from VIR’s sunflower genetic collection have not yet been studied. Materials and methods. The material included 27 short-stemmed and 10 tall sunflower lines from VIR’s genetic collection, as well as the F1 and F2 hybrid generations derived from crossing the tall (VIR 340) and dwarf (VIR 171) genotypes. The parental lines and hybrids were phenotyped for plant height, leaf number, and internode length. Genotyping for the Rht1 locus (HaDella1 candidate gene), encoding the negative regulator of the gibberellin response, the DELLA protein, was performed using the developed CAPS marker. Results. The average plant height in the VIR 340 line over a three-year study was 162 cm, the number of leaves 29, and the internode length 6 cm. The VIR 171 line demonstrated the plant height of 66 cm, leaf number of 24, and internode length of 2.8 cm. The F1 hybrids were uniform, with the height of 180–190 cm, that indicated the dominance of the long stem trait. Analyzing the segregation in the F2 hybrid generation led to an assumption admitting the digenic control of the dwarf phenotype in the VIR 171 line. The CAPS marker G-D-1/ Bmt I was developed to identify a missense mutation T>C in the first exon of the HaDella1 gene, which results in the substitution of leucine with proline in the DELLA motif. Using the marker, the mutant Rht1 allele was identified in the VIR 171 and VIR 434 dwarf lines, similar in their origin and phenotype. The results of validation in the F2 hybrid population (VIR 340 × VIR 171) confirmed the efficiency of the G-D-1 / Bmt I marker for selecting dwarf genotypes homozygous for the Rht1 mutant allele.

OsKANADI1 and OsYABBY5 regulate rice plant height by targeting GIBERELLIN 2-OXIDASE6.

Plant height is an important agronomic characteristic of rice (Oryza sativa L.). Map-based cloning analyses of a natural semi-dwarf rice mutant with inwardly curled leaves found in the field revealed that the defects were due to a mutation of a SHAQKYF-class MYB family transcription factor, OsKANADI1 (OsKAN1). OsKAN1 directly bound to the OsYABBY5 (OsYAB5) promoter to repress its expression and interacted with OsYAB5 to form a functional OsKAN1-OsYAB5 complex. GIBERELLIN 2-OXIDASE6 (OsGA2ox6), encoding an enzyme in the gibberellin (GA) catabolic pathway, was activated by OsYAB5. Furthermore, the OsKAN1-OsYAB5 complex suppressed the inhibitory effect of OsKAN1 toward OsYAB5 and inhibited OsYAB5-induced OsGA2ox6 expression. The proOsKAN1:OsYAB5 transgenic plants were taller than wild-type plants, whereas oskan1 proOsKAN1:OsYAB5 plants exhibited a severe dwarf phenotype due to the absence of the OsKAN1-OsYAB5 complex. The OsKAN1-OsYAB5 complex modulated OsGA2ox6 expression, thereby regulating the levels of bioactive gibberellins and, consequently, plant height. This study elucidated the mechanism underlying the effect of the OsKAN1-OsYAB5-OsGA2ox6 regulatory pathway on plant height at different positions in rice stems and provided insights on stem development and candidate genes for the aerial architecture improvement of crop plants.

The brassinosteroid receptor gene BRI1 safeguards cell-autonomous brassinosteroid signaling across tissues.

Brassinosteroid signaling is essential for plant growth as exemplified by the dwarf phenotype of loss-of-function mutants in BRASSINOSTEROID INSENSITIVE 1 (BRI1), a ubiquitously expressed Arabidopsis brassinosteroid receptor gene. Complementation of brassinosteroid-blind receptor mutants by BRI1 expression with various tissue-specific promoters implied that local brassinosteroid signaling may instruct growth non-cell autonomously. Here, we performed such rescues with a panel of receptor variants and promoters, in combination with tissue-specific transgene knockouts. Our experiments demonstrate that brassinosteroid receptor expression in several tissues is necessary but not sufficient for rescue. Moreover, complementation with tissue-specific promoters requires the genuine BRI1 gene body sequence, which confers ubiquitous expression of trace receptor amounts that are sufficient to promote brassinosteroid-dependent root growth. Our data, therefore, argue for a largely cell-autonomous action of brassinosteroid receptors.

Overexpression of the Poa pratensis GA2ox gene family significantly reduced the plant height of transgenic Arabidopsis thaliana and Poa pratensis

Gibberellin (GAs) is an important plant hormone that plays a key role in plant growth and development. Gibberellin 2-oxidase (GA2ox) catalyzes the inactivation of biologically active GA or their direct precursors. In this study, five GA2ox genes were isolated from the wild type Poa pratensis ‘Baron’, named PpGA2ox3, PpGA2ox4, PpGA2ox5, PpGA2ox8, and PpGA2ox9. Phylogenetic tree analysis showed that PpGA2ox3, PpGA2ox4, PpGA2ox5, and PpGA2ox8 belong to class I GA2ox genes, while PpGA2ox9 belongs to class III GA2ox genes. They expressed in all tissues of Poa pratensis, in each plant tissue and growth stage, the expression patterns were different. After GA3 spraying treatment, the expression of each gene showed different patterns. Subcellular localization showed that PpGA2ox3 was located in chloroplasts, while PpGA2ox5 and PpGA2ox9 were located in the cytoplasm. When PpGA2ox3 and PpGA2ox9 were overexpressed in Arabidopsis thaliana, they all led to a typical dwarf phenotype, as well as low plant height, small leaves and late flowering. Similarly, when they overexpressed in P. pratensis, the transgenic plants also exhibited a dwarf phenotype with a lower leaf length/width ratio. Hormone analysis suggested that these dwarfing traits might be caused by a decrease in GA4 content. These studies indicated that the PpGA2ox gene family played an important role in studying the mechanism of plant dwarfism and also had the potential to become important genes for the breeding of P. pratensis.

A DUF21 domain-containing protein regulates plant dwarfing in watermelon.

Dwarf or semi-dwarf plant structures are well-suited for intensive farming, maximizing yield, and minimizing labor costs. Watermelon (Citrullus lanatus) is classified as an annual vine plant with elongated internodes, yet the mechanism governing watermelon dwarfing remains unclear. In this study, a compact watermelon mutant dwarf, induced by the insertion of T-DNA, was discovered. Through re-sequencing, a gene named domain of unknown function 21 (ClDUF21), located downstream of the T-DNA insertion site, was identified as the candidate gene for the dwarf mutant, and its functionality was subsequently confirmed. Watermelon mutants generated through CRISPR/Cas9-mediated knockout of ClDUF21 revealed that homozygous mutants displayed a pronounced dwarfing phenotype, and protein-protein interaction analysis confirmed the direct interaction between ClDUF21 and ClDWF1. Subsequently, we employed CRISPR/Cas9 technology to precisely modify the homologous gene CsDUF21 in cucumber (Cucumis sativus) and performed protein interaction validation between CsDUF21 and CsDWF1, thereby demonstrating that the CsDUF21 gene also exhibits analogous functionality in plant dwarfing. These findings demonstrate that ClDUF21 governs plant dwarfism by modulating the brassinosteroid synthesis pathway via ClDWF1.

The SINA1-BSD1 Module Regulates Vegetative Growth Involving Gibberellin Biosynthesis in Tomato.

In plants, vegetative growth is controlled by synergistic and/or antagonistic effects of many regulatory factors. Here, the authors demonstrate that the ubiquitin ligase seven in absentia1 (SINA1) mammalian BTF2-like transcription factors, Drosophila synapse-associated proteins, and yeast DOS2-like proteins (BSD1) function as a regulatory module to control vegetative growth in tomato via regulation of the production of plant growth hormone gibberellin (GA). SINA1 negatively regulates the protein level of BSD1 through ubiquitin-proteasome-mediated degradation, and the transgenic tomato over-expressing SINA1 (SINA1-OX) resembles the dwarfism phenotype of the BSD1-knockout (BSD1-KO) tomato plant. BSD1 directly activates expression of the BSD1-regulated gene 1 (BRG1) via binding to a novel core BBS (standing for BSD1 binding site) binding motif in the BRG1 promoter. Knockout of BRG1 (BRG1-KO) in tomato also results in a dwarfism phenotype, suggesting BRG1 plays a positive role in vegetative growth as BSD1 does. Significantly, GA contents are attenuated in transgenic SINA1-OX, BSD1-KO, and BRG1-KO plants exhibiting dwarfism phenotype and exogenous application of bioactive GA3 restores their vegetative growth. Moreover, BRG1 is required for the expression of multiple GA biosynthesis genes and BSD1 activates three GA biosynthesis genes promoting GA production. Thus, this study suggests that the SINA1-BSD1 module controls vegetative growth via direct and indirect regulation of GA biosynthesis in tomato.

NADP-malic Enzyme OsNADP-ME2 Modulates Plant Height Involving in Gibberellin Signaling in Rice

Plants NADP-malic enzymes (NADP-MEs) act as a class of oxidative decarboxylase to mediate malic acid metabolism in organisms. Despite NADP-MEs have been demonstrated to play pivotal roles in regulating diverse biological processes, the role of NADP-MEs involving in plant growth and development remains rarely known. Here, we characterized the function of rice cytosolic OsNADP-ME2 in regulating plant height. The results showed that RNAi silencing and knock-out of OsNADP-ME2 in rice results in a dwarf plant structure, associating with significant expression inhibition of genes involving in phytohormone Gibberellin (GA) biosynthesis and signaling transduction, but with up-regulation for the expression of GA signaling suppressor SLR1. The accumulation of major bioactive GA1, GA4 and GA7 are evidently altered in RNAi lines, and exogenous GA treatment compromises the dwarf phenotype of OsNADP-ME2 RNAi lines. RNAi silencing of OsNADP-ME2 also causes the reduction of NADP-ME activity associating with decreased production of pyruvate. Thus, our data revealed a novel function of plant NADP-MEs in modulation of rice plant height through regulating bioactive GAs accumulation and GA signaling, and provided a valuable gene resource for rice plant architecture improvement.

Identification and analysis of gibberellin 2-oxidase (GA2ox) members in Cunninghamia lanceolate and the negative regulatory character of ClGA2ox12 in tree stature and xylem lignin deposits

Cunninghamia lanceolata (Lamb.) Hook (C. lanceolate) is a fast-growing evergreen coniferous tree species with high wood yield and fantabulous timber quality. To date, numerous studies have revealed that gibberellins (GAs) regulate the formation of plant wood (secondary xylem). Moreover, GA 2-oxidases (GA2oxs) are known as the major enzymes that deactivate GAs. However, a small or no number of the GA2oxs involved in xylem development and lignin biosynthesis mediated by the GA pathway in woody plants have been identified previously. In this study, 5 GA2oxs members were identified in C. lanceolate, including 4 ClGA2ox12s and one ClGA2ox8-like. Exogenous gibberellin not only promoted lignin deposition in the stem of C. lanceolate but also up-regulated the expression of the lignin biosynthesis gene ClCAD1 by 2–3 folds and one of ClGA2ox12s (designated ClGA2ox12) by 4 folds, suggesting that ClGA2ox12 may be involved in the modulation of lignin deposition by the gibberellin signaling pathway in C. lanceolate. Furthermore, transgenic Populus tomentosa overexpressing ClGA2ox12 exhibited growth retardation and a typical dwarfing phenotype, with a reduction of approximately 40 % in the number of xylem cell layers and a decrease of approximately 16.7 %-26.9 % in lignin content relative to wild-type plants. At the same time, the expression level of lignin biosynthetic genes and lignin content were down-regulated in transgenic plants. Together, these results suggested that ClGA2ox12 functioned as a GA 2oxs catabolic enzyme that plays pivotal roles in plant growth processes, xylem development, and lignin biosynthesis, which gives insights into producing dwarf and low-lignin varieties of C. lanceolate.

Genome-Wide Analysis of the Gibberellin-Oxidases Family Members in Four Prunus Species and a Functional Analysis of PmGA2ox8 in Plant Height.

Gibberellins (GAs), enzymes that play a significant role in plant growth and development, and their levels in plants could be regulated by gibberellin-oxidases (GAoxs). As important fruit trees and ornamental plants, the study of the mechanism of plant architecture formation of the Prunus genus is crucial. Here, 85 GAox genes were identified from P. mume, P. armeniaca, P. salicina, and P. persica, and they were classified into six subgroups. Conserved motif and gene structure analysis showed that GAoxs were conserved in the four Prunus species. Collinearity analysis revealed two fragment replication events of PmGAoxs in the P. mume genome. Promoter cis-elements analysis revealed 24 PmGAoxs contained hormone-responsive elements and development regulatory elements. The expression profile indicated that PmGAoxs have tissue expression specificity, and GA levels during the dormancy stage of flower buds were controlled by certain PmGAoxs. After being treated with IAA or GA3, the transcription level of PmGA2ox8 in stems was significantly increased and showed a differential expression level between upright and weeping stems. GUS activity driven by PmGA2ox8 promoter was detected in roots, stems, leaves, and flower organs of Arabidopsis. PmGA2ox8 overexpression in Arabidopsis leads to dwarfing phenotype, increased number of rosette leaves but decreased leaf area, and delayed flowering. Our results showed that GAoxs were conserved in Prunus species, and PmGA2ox8 played an essential role in regulating plant height.

Convergent dwarfism consequences of minipigs under independent artificial selections

BackgroundCurrently, diverse minipigs have acquired a common dwarfism phenotype through independent artificial selections. Characterizing the population and genetic diversity in minipigs is important to unveil genetic mechanisms regulating their body sizes and effects of independent artificial selections on those genetic mechanisms. However, full understanding for the genetic mechanisms and phenotypic consequences in minipigs still lag behind.ResultsHere, using whole genome sequencing data of 41 pig breeds, including eight minipigs, we identified a large genomic diversity in a minipig population compared to other pig populations in terms of population structure, demographic signatures, and selective signatures. Selective signatures reveal diverse biological mechanisms related to body size in minipigs. We also found evidence for neural development mechanism as a minipig-specific body size regulator. Interestingly, selection signatures within those mechanisms containing neural development are also highly different among minipig breeds. Despite those large genetic variances, PLAG1, CHM, and ESR1 are candidate key genes regulating body size which experience different differentiation directions in different pig populations.ConclusionsThese findings present large variances of genetic structures, demographic signatures, and selective signatures in the minipig population. They also highlight how different artificial selections with large genomic diversity have shaped the convergent dwarfism.

A transcriptional cascade involving BBX22 and HY5 finely regulates both plant height and fruit pigmentation in citrus.

Dwarfing is a pivotal agronomic trait affecting both yield and quality. Citrus species exhibit substantial variation in plant height, among which internode length is a core element. However, the molecular mechanism governing internode elongation remains unclear. Here, we unveiled that the transcriptional cascade consisting of B-BOX DOMAIN PROTEIN 22 (BBX22) and ELONGATED HYPOCOTYL 5 (HY5) finely tunes plant height and internode elongation in citrus. Loss-of-function mutations of BBX22 in an early-flowering citrus (Citrus hindsii "SJG") promoted internode elongation and reduced pigment accumulation, whereas ectopic expression of BBX22 in SJG, sweet orange (C. sinensis), pomelo (C. maxima) or heterologous expression of BBX22 in tomato (Solanum lycopersicum) significantly decreased internode length. Furthermore, exogenous application of gibberellin A3 (GA3) rescued the shortened internode and dwarf phenotype caused by BBX22 overexpression. Additional experiments revealed that BBX22 played a dual role in regulation internode elongation and pigmentation in citrus. On the one hand, it directly bound to and activated the expression of HY5, GA metabolism gene (GA2 OXIDASE 8, GA2ox8), carotenoid biosynthesis gene (PHYTOENE SYNTHASE 1, PSY1) and anthocyanin regulatory gene (Ruby1, a MYB DOMAIN PROTEIN). On the other hand, it acted as a cofactor of HY5, enhancing the ability of HY5 to regulate target genes expression. Together, our results reveal the critical role of the transcriptional cascade consisting of BBX22 and HY5 in controlling internode elongation and pigment accumulation in citrus. Unraveling the crosstalk regulatory mechanism between internode elongation and fruit pigmentation provides key genes for breeding of novel types with both dwarf and health-beneficial fortification in citrus.

Genome-Wide Identification of Height-Related Genes Using Three Maize Dwarfs and RNA-Seq

Plant height is an important grain yield-associated trait in maize. To date, few genes related to plant height have been characterized in maize. To better understand the genetic mechanisms of plant height in maize, we revealed the transcriptional changes of three dwarf mutants compared to the wild type. By ethyl methane sulfonate treatment of the wild-type maize cultivar PH6WC, we obtained three dwarfs—PH6WCdwarf1 (pd1), PH6WCdwarf2 (pd2), and PH6WCdwarf3 (pd3)—and their plant heights were reduced by 42%, 38%, and 24%, respectively. RNA-Seq data suggested that 1641 differentially expressed genes (DEGs) overlapped with each other among the three dwarfs at the seedling stage. Further analysis showed that the DEGs were divided into four groups with different expression patterns. Functional analysis revealed that these DEGs were commonly enriched in 47 GO terms mainly involved in cytokinesis, hormone, and energy metabolism pathways. Among them, An1, involved in the GA biosynthesis pathway, and mutations in An1 result in reduced plant height. EREB182 encodes ethylene-responsive element binding protein 2, which is critical for internode elongation. Microtubule-related genes Zmtub2, Zmtub3, Zmtub5, Zmtub6, and TUBG2 were commonly enriched among the three comparisons. Previous studies have shown that mutations in microtubule-associated genes cause the dwarf phenotype. However, nearly half of the common DEGs had no functional information, such as Zm00001d000107, Zm00001d000279, etc., implying their novel and specific functions in maize. Overall, this study identifies several potential plant height-related genes and contributes to linking genetic resources with maize breeding.

Glycosylation mode of phloretin affects the morphology and stress resistance of apple plant.

Phloretin has different glycosylation modes in plants. Phlorizin (phloretin 2'-O-glucoside) is one of the glycosylation products of phloretin, and accumulates abundantly in apple plants. However, it is still unclear whether phlorizin is more beneficial for apple plants compared with other glycosylation products of phloretin. We created transgenic apple plants with different glycosylation modes of phloretin. In transgenic plants, the accumulation of phlorizin was partly replaced by that of trilobatin (phloretin 4'-O-glucoside) or phloretin 3',5'-di-C-glycoside. Compared with wild type, transgenic plants with less phlorizin showed dwarf phenotype, larger stomatal size, higher stomatal density and less tolerance to drought stress. Transcriptome and phytohormones assay indicate that phlorizin might regulate stomatal development and behaviour via controlling auxin and abscisic acid signalling pathways as well as carbonic anhydrase expressions. Transgenic apple plants with less phlorizin also showed less resistance to spider mites. Apple plants may hydrolyse phlorizin to produce phloretin, but cannot hydrolyse trilobatin or phloretin 3',5'-di-C-glycoside. Compared with its glycosylation products, phloretin is more toxic to spider mites. These results suggest that the glycosylation of phloretin to produce phlorizin is the optimal glycosylation mode in apple plants, and plays an important role in apple resistance to stresses.

Characterization of the ddt1 Mutant in Rice and Its Impact on Plant Height Reduction and Water Use Efficiency.

Rice (Oryza sativa L.), a fundamental global staple, nourishes over half of the world's population. The identification of the ddt1 mutant in rice through EMS mutagenesis of the indica cultivar Shuhui527 revealed a dwarf phenotype, characterized by reduced plant height, smaller grain size, and decreased grain weight. Detailed phenotypic analysis and map-based cloning pinpointed the mutation to a single-base transversion in the LOC_Os03g04680 gene, encoding a cytochrome P450 enzyme, which results in a premature termination of the protein. Functional complementation tests confirmed LOC_Os03g04680 as the DDT1 gene responsible for the observed phenotype. We further demonstrated that the ddt1 mutation leads to significant alterations in gibberellic acid (GA) metabolism and signal transduction, evidenced by the differential expression of key GA-related genes such as OsGA20OX2, OsGA20OX3, and SLR1. The mutant also displayed enhanced drought tolerance, as indicated by higher survival rates, reduced water loss, and rapid stomatal closure under drought conditions. This increased drought resistance was linked to the mutant's improved antioxidant capacity, with elevated activities of antioxidant enzymes and higher expression levels of related genes. Our findings suggest that DDT1 plays a crucial role in regulating both plant height and drought stress responses. The potential for using gene editing of DDT1 to mitigate the dwarf phenotype while retaining improved drought resistance offers promising avenues for rice improvement.

A cargo sorting receptor mediates chloroplast protein trafficking through the secretory pathway.

Nucleus-encoded chloroplast proteins can be transported via the secretory pathway. The molecular mechanisms underlying the trafficking of chloroplast proteins between the intracellular compartments are largely unclear, and a cargo sorting receptor has not previously been identified in the secretory pathway. Here, we report a cargo sorting receptor that is specifically present in Viridiplantae and mediates the transport of cargo proteins to the chloroplast. Using a forward genetic analysis, we identified a gene encoding a transmembrane protein (MtTP930) in barrel medic (Medicago truncatula). Mutation of MtTP930 resulted in impaired chloroplast function and a dwarf phenotype. MtTP930 is highly expressed in the aerial parts of the plant and is localized to the endoplasmic reticulum (ER) exit sites and Golgi. MtTP930 contains typical cargo sorting receptor motifs, interacts with Sar1, Sec12, and Sec24, and participates in coat protein complex II vesicular transport. Importantly, MtTP930 can recognize the cargo proteins plastidial N-glycosylated nucleotide pyrophosphatase/phosphodiesterase (MtNPP) and α-carbonic anhydrase (MtCAH) in the ER and then transport them to the chloroplast via the secretory pathway. Mutation of a homolog of MtTP930 in Arabidopsis (Arabidopsis thaliana) resulted in a similar dwarf phenotype. Furthermore, MtNPP-GFP failed to localize to chloroplasts when transgenically expressed in Attp930 protoplasts, implying that these cargo sorting receptors are conserved in plants. These findings fill a gap in our understanding of the mechanism by which chloroplast proteins are sorted and transported via the secretory pathway.

OsFK1 encodes C-14 sterol reductase, which is involved in sterol biosynthesis and affects premature aging of leaves in rice

The enzyme C-14 sterol reductase is involved in biosynthesis of brassinosteroids (BR) and sterols, as well as plant development. OsFK1, a member of the sterol biosynthesis pathway located in the endoplasmic reticulum (ER), encodes C-14 sterol reductase. However, there is little research on the function of C-14 sterol reductase in rice. Compared with the wild type, an osfk1 mutant showed dwarf phenotype and premature aging in the second leaf during the trefoil stage, and abnormal development of leaf veins during the tillering stage. The osfk1 mutant showed signs of aberrant PCD, as evidenced by TUNEL staining. This suggested that high ROS buildup caused DNA damage and ROS-mediated cell death in the mutant. The osfk1 mutant also showed decreased chlorophyll content and aberrant chloroplast structure. Sequencing of the osfk1 mutant allele revealed a non-synonymous G to A mutation in the final intron, leading to early termination. Here, we identified the OsFK1 allele, cloned it by Mutmap sequencing, and verified it by complementation. HPLC-MS/MS assays demonstrated that the osfk1 mutation caused lower phytosterol levels. These findings showed that the OsFK1 allele encoding C-14 sterol reductase is involved in phytosterol biosynthesis and mediates normal development of rice plants.

Characterization of a new dwarf watercress (Nasturtium officinale R Br.) ‘Boldrewood’ in commercial trials reveals a consistent increase in chemopreventive properties in a longer-grown crop

We describe ‘Boldrewood’, a new accession of watercress (Nasturtium officinale R. Br.) that was initially found to be of short stature with high antioxidant capacity (Payne et al. 2015). This was of particular commercial interest because it offered the potential to develop a novel watercress product with fork-friendly size and improved health-benefits. In two commercial trials comparing Boldrewood to a control, we confirmed that Boldrewood exhibits a dwarf phenotype with a significantly shorter stem and consistently produced more leaves per stem area alongside comparable crop biomass. The antioxidant and chemopreventive capacity of Boldrewood were comparable to the commercial crop. For the first time, we observed a novel increase in glucosinolate concentrations and cytotoxicity to cancer cells, characterised as decreased IC50 (half-maximal concentration of an inhibitor), associated with increased crop age at harvest. This suggests that a slower-growing and longer to harvest crop provides a significant improvement in health benefits gained in this leafy crop which is already known to be highly nutrient dense and with considerable chemopreventive ability.

Chromatin profiling identifies chondrocyte-specific Sox9 enhancers important for skeletal development.

The transcription factor SRY-related HMG box 9 (Sox9) is essential for chondrogenesis. Mutations in and around SOX9 cause campomelic dysplasia (CD) characterized by skeletal malformations. Although the function of Sox9 in this context is well studied, the mechanisms that regulate Sox9 expression in chondrocytes remain to be elucidated. Here, we have used genome-wide profiling to identify 2 Sox9 enhancers located in a proximal breakpoint cluster responsible for CD. Enhancer activity of E308 (located 308 kb 5' upstream) and E160 (located 160 kb 5' upstream) correlated with Sox9 expression levels, and both enhancers showed a synergistic effect in vitro. While single deletions in mice had no apparent effect, simultaneous deletion of both E308 and E160 caused a dwarf phenotype, concomitant with a reduction of Sox9 expression in chondrocytes. Moreover, bone morphogenetic protein 2-dependent chondrocyte differentiation of limb bud mesenchymal cells was severely attenuated in E308/E160 deletion mice. Finally, we found that an open chromatin region upstream of the Sox9 gene was reorganized in the E308/E160 deletion mice to partially compensate for the loss of E308 and E160. In conclusion, our findings reveal a mechanism of Sox9 gene regulation in chondrocytes that might aid in our understanding of the pathophysiology of skeletal disorders.

Identification and characterization of CsERECTA, a major gene controlling stem elongation through regulating GA biosynthesis in cucumber.

Dwarfing is an ideal agronomic trait in crop breeding, which can improve lodging resistance and increase crop productivity. In this study, we identified a dwarf mutant cp-3 from an EMS-mutagenized population, which had extremely short internodes, and the cell length and number of internodes were significantly reduced. Meanwhile, exogenous GA3 treatment partially rescued the plant height of the cp-3. Inheritance analysis showed that the cp-3 mutant was regulated via a recessive nuclear locus. A candidate gene, CsERECTA, encoding an LRR receptor-like serine/threonine-protein kinase, was cloned through a map-based cloning strategy. Sequence analysis showed that a nucleotide mutation (C ~ T) in exon 26 of CsERECTA led to premature termination of the protein. Subsequently, two transgenic lines were generated using the CRISPR/Cas9 system, and they showed plant dwarfing. Plant endogenous hormones quantitative and RNA-sequencing analysis revealed that GA3 content and the expression levels of genes related to GA biosynthesis were significantly reduced in Cser knockout mutants. Meanwhile, exogenous GA3 treatment partially rescued the dwarf phenotype of Cser knockout mutants. These findings revealed that CsERECTA controls stem elongation by regulating GA biosynthesis in cucumber.