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Summary: Grain chalkiness is an undesirable trait that negatively regulates grain yield and quality in rice. However, the regulatory mechanism underlying chalkiness is complex and remains unclear. We identified a positive regulator of white‐belly rate (WBR). The WBR7 gene encodes sucrose synthase 3 (SUS3). A weak functional allele of WBR7 is beneficial in increasing grain yield and quality. During the domestication of indica rice, a functional G/A variation in the coding region of WBR7 resulted in an E541K amino acid substitution in the GT‐4 glycosyltransferase domain, leading to a significant decrease in decomposition activity of WBR7A (allele in cultivar Jin23B) compared with WBR7G (allele in cultivar Beilu130). The NIL(J23B) and knockout line NIL(BL130)KO exhibited lower WBR7 decomposition activity than that of NIL(BL130) and NIL(J23B)COM, resulting in less sucrose decomposition and metabolism in the conducting organs. This caused more sucrose transportation to the endosperm, enhancing the synthesis of storage components in the endosperm and leading to decreased WBR. More sucrose was also transported to the anthers, providing sufficient substrate and energy supply for pollen maturation and germination, ultimately leading to an increase rate of seed setting and increased grain yield. Our findings elucidate a mechanism for enhancing rice yield and quality by modulating sucrose metabolism and allocation, and provides a valuable allele for improved rice quality. [ABSTRACT FROM AUTHOR]

Rapeseed (Brassica napus) is an important oilseed crop worldwide. Plant vascular tissues are responsible for long-distance transport of water and nutrients and for providing mechanical support. The lateral roots absorb water and nutrients. The genetic basis of vascular tissue and lateral root development in rapeseed remains unknown. This study characterized an ethyl methanesulfonate-mutagenized rapeseed mutant, T16 , which showed dwarf stature, reduced lateral roots, and leaf wilting. SEM observations showed that the internode cells were shortened. Observations of tissue sections revealed defects in vascular bundle development in the stems and petioles. Genetic analysis revealed that the phenotypes of T16 were controlled by a single semi-dominant nuclear gene. Map-based cloning and genetic complementarity identified BnaA03.IAA13 as the functional gene; a G-to-A mutation in the second exon changed glycine at position 79 to glutamic acid, disrupting the conserved degron motif VGWPP. Transcriptome analysis in roots and stems showed that auxin and cytokinin signaling pathways were disordered in T16. Evolutionary analysis showed that AUXIN/INDOLE-3-ACETIC ACID is conserved during plant evolution. The heterozygote of T16 showed significantly reduced plant height while maintaining other agronomic traits. Our findings provide novel insights into the regulatory mechanisms of vascular tissue and lateral root development, and offer a new germplasm resource for rapeseed breeding. [ABSTRACT FROM AUTHOR]

Abstract The purple leaves of Brassica napus are abundant in anthocyanins, which are renowned for their role in conferring distinct colors, stress tolerance, and health benefits, however the genetic basis of this trait in B. napus remains largely unelucidated. Herein, the purple leaf B. napus (PL) exhibited purple pigments in the upper epidermis and a substantial increase in anthocyanin accumulation, particularly of cyanidin, compared to green leaf B. napus (GL). The genetic control of the purple leaf trait was attributed to a semi-dominant gene, pl, which was mapped to the end of chromosome A03. However, sequencing of the fragments amplified by the markers linked to pl indicated that they were all mapped to chromosome B05 from B. juncea. Within this B05 chromosomal segment, the BjMYB113 gene-specific marker showed perfect co-segregation with the purple leaf trait in the F2 population, suggesting that the BjMYB113 introgression from B. juncea was the candidate gene for the purple leaf trait in B. napus. To further verify the function of candidate gene, CRISPR/Cas9 was performed to knock out the BjMYB113 gene in PL. The three myb113 mutants exhibited evident green leaf phenotype, absence of purple pigments in the adaxial epidermis, and a significantly reduced accumulation of anthocyanin compared to PL. Additionally, the genes involved in positive regulatory (TT8), late anthocyanin biosynthesis (DFR, ANS, UFGT), as well as transport genes (TT19) were significantly suppressed in the myb113 mutants, further confirming that BjMYB113 was response for the anthocyanin accumulation in purple leaf B. napus. This study contributes to an advanced understanding of the regulation mechanism of anthocyanin accumulation in B. napus.

Plant height, spike, leaf, stem and grain morphologies are key components of plant architecture and related to wheat yield. A wheat (Triticum aestivum L.) mutant, wpa1, displaying temperature-dependent pleiotropic developmental anomalies, was isolated. The WPA1 gene, encoding a von Willebrand factor type A (vWA) domain protein, was located on chromosome arm 7DS and isolated by map-based cloning. The functionality of WPA1 was validated by multiple independent EMS-induced mutants and gene editing. Phylogenetic analysis revealed that WPA1 is monocotyledon-specific in higher plants. The identification of WPA1 provides opportunity to study the temperature regulated wheat development and grain yield. [ABSTRACT FROM AUTHOR]

Simple Summary: CRM domain proteins play key roles in plant growth and chloroplast development. To explore the role of CRM domain protein in the chloroplast development of rice, we cloned a rice CRM domain protein gene, YL4, and then conducted a tissue-specific expression analysis, subcellular localization, and transcription analysis. The experimental results showed that YL4 was highly expressed in leaves and localized to the chloroplast, and its mutation affected chloroplast-related gene transcription levels and some agronomic traits. These results indicate that the YL4 gene is a pivotal regulatory factor in chloroplast development and rice growth. Chloroplast RNA splicing and ribosome maturation (CRM) domain proteins are a family of plant-specific proteins associated with RNA binding. In this study, we have conducted a detailed characterization of a novel rice CRM gene (LOC_Os04g39060) mutant, yl4, which showed yellow-green leaves at all the stages, had fewer tillers, and had a decreased plant height. Map-based cloning and CRISPR/Cas9 editing techniques all showed that YL4 encoded a CRM domain protein in rice. In addition, subcellular localization revealed that YL4 was in chloroplasts. YL4 transcripts were highly expressed in all leaves and undetectable in roots and stems, and the mutation of YL4 affected the transcription of chloroplast-development-related genes. This study indicated that YL4 is essential for chloroplast development and affects some agronomic traits. [ABSTRACT FROM AUTHOR]

Leaf is an important organ of photosynthesis in plants, and leaf color has an important influence on photosynthesis. However, the molecular mechanisms of leaf color regulation are still unclear. In this study, we identified a rice (Oryza sativa) temperature sensitive albino trans-green (tag) mutant, which showed albino leaves at the seedling stage, and gradually turned green. Map-based cloning showed that TAG is a Novel allele gene of OsBT3, encoding an adenine nucleotide transport protein. Compared with the allele mutants of sla and wsl2, tag displayed some new phenotypes, such as it was a low-temperature-sensitive albino trans-green mutant with serious DNA damage in the albino leaves, and the PSI photosynthetic efficiency and photosynthesis of it were enhanced significantly in mature stage. Besides, we also found TAG was also localized in chloroplasts and played an important role on chloroplast development. Moreover, we newly discovered that TAG is highly expressed in L3U, which is induced by low temperature, and in situ hybridization results showed TAG expressed in the shoot apical meristem, leaf primordium and vascular bundle. Taken together, the above new results provide deeper clues to explain the molecular mechanisms by which TAG regulates chloroplast development. [ABSTRACT FROM AUTHOR]

Leaf shape is considered to be one of the most significant agronomic traits in crop breeding. However, the molecular basis underlying leaf morphogenesis in cotton is still largely unknown. In this study, through genetic mapping and molecular investigation using a natural cotton mutant cu with leaves curling upward, the causal gene GHCU is successfully identified as the key regulator of leaf flattening. Knockout of GHCU or its homolog in cotton and tobacco using CRISPR results in abnormal leaf shape. It is further discovered that GHCU facilitates the transport of the HD protein KNOTTED1‐like (KNGH1) from the adaxial to the abaxial domain. Loss of GHCU function restricts KNGH1 to the adaxial epidermal region, leading to lower auxin response levels in the adaxial boundary compared to the abaxial. This spatial asymmetry in auxin distribution produces the upward‐curled leaf phenotype of the cu mutant. By analysis of single‐cell RNA sequencing and spatiotemporal transcriptomic data, auxin biosynthesis genes are confirmed to be expressed asymmetrically in the adaxial‐abaxial epidermal cells. Overall, these findings suggest that GHCU plays a crucial role in the regulation of leaf flattening through facilitating cell‐to‐cell trafficking of KNGH1 and hence influencing the auxin response level. [ABSTRACT FROM AUTHOR]

Our study investigates the genetic mechanisms underlying the spotted leaf phenotype in rice, focusing on the spl43 mutant. This mutant is characterized by persistent reddish-brown leaf spots from the seedling stage to maturity, leading to extensive leaf necrosis. Using map-based cloning, we localized the responsible locus to a 330 Kb region on chromosome 2. We identified LOC_Os02g56000, named OsRPT5A, as the causative gene. A point mutation in OsRPT5A, substituting valine for glutamic acid, was identified as the critical factor for the phenotype. Functional complementation and the generation of CRISPR/Cas9-mediated knockout lines in the IR64 background confirmed the central role of OsRPT5A in controlling this trait. The qPCR results from different parts of the rice plant revealed that OsRPT5A is constitutively expressed across various tissues, with its subcellular localization unaffected by the mutation. Notably, we observed an abnormal accumulation of reactive oxygen species (ROS) in spl43 mutants by examining the physiological indexes of leaves, suggesting a disruption in the ROS system. Complementation studies indicated OsRPT5A's involvement in ROS homeostasis and catalase activity regulation. Moreover, the spl43 mutant exhibited enhanced resistance to Xanthomonas oryzae pv. oryzae (Xoo), highlighting OsRPT5A's role in rice pathogen resistance mechanisms. Overall, our results suggest that OsRPT5A plays a critical role in regulating ROS homeostasis and enhancing pathogen resistance in rice. [ABSTRACT FROM AUTHOR]

Leaf senescence is an essential physiological process related to grain yield potential and nutritional quality. Green leaf duration (GLD) after anthesis directly reflects the leaf senescence process and exhibits large genotypic differences in common wheat; however, the underlying gene regulatory mechanism is still lacking. Here, we identified TaNAM‐A1 as the causal gene of the major loci qGLD‐6A for GLD during grain filling by map‐based cloning. Transgenic assays and TILLING mutant analyses demonstrated that TaNAM‐A1 played a critical role in regulating leaf senescence, and also affected spike length and grain size. Furthermore, the functional divergences among the three haplotypes of TaNAM‐A1 were systematically evaluated. Wheat varieties with TaNAM‐A1d (containing two mutations in the coding DNA sequence of TaNAM‐A1) exhibited a longer GLD and superior yield‐related traits compared to those with the wild type TaNAM‐A1a. All three haplotypes were functional in activating the expression of genes involved in macromolecule degradation and mineral nutrient remobilization, with TaNAM‐A1a showing the strongest activity and TaNAM‐A1d the weakest. TaNAM‐A1 also modulated the expression of the senescence‐related transcription factors TaNAC‐S‐7A and TaNAC016‐3A. TaNAC016‐3A enhanced the transcriptional activation ability of TaNAM‐A1a by protein–protein interaction, thereby promoting the senescence process. Our study offers new insights into the fine‐tuning of the leaf functional period and grain yield formation for wheat breeding under various geographical climatic conditions. [ABSTRACT FROM AUTHOR]

Abstract Fruit length is a crucial agronomic trait of snake gourd (Trichosanthes anguina L); however, genes associated with fruit length have not been characterised. In this study, F2 snake gourd populations were generated by crossing the inbred lines, S1 and S2 (fruit lengths: 110 and 20 cm, respectively). Subsequently, bulk segregant analysis, sequencing, and fine-mapping were performed on the F2 population to identify target genes. Our findings suggest that the fruit length of snake gourd is regulated by a major-effect regulatory gene. Mining of genes regulating fruit length in snake gourd to provide a basis for subsequent selection and breeding of new varieties. Genotype-phenotype association analysis was performed on the segregating F2 population comprising 6,000 plants; the results indicate that the target gene is located on Chr4 (61,846,126–61,865,087 bp, 18.9-kb interval), which only carries the annotated candidate gene, Tan0010544 (designated TFL). TFL belongs to the MADS-box family, one of the largest transcription factor families. Sequence analysis revealed a non-synonymous mutation of base C to G at position 202 in the coding sequence of TFL, resulting in the substitution of amino acid Gln to Glu at position 68 in the protein sequence. Subsequently, an InDel marker was developed to aid the marker-assisted selection of TFL. The TFL in the expression parents within the same period was analysed using quantitative real-time PCR; the TFL expression was significantly higher in short fruits than long fruits. Therefore, TFL can be a candidate gene for determining the fruit length in snake gourd. Collectively, these findings improve our understanding of the genetic components associated with fruit length in snake gourds, which could aid the development of enhanced breeding strategies for plant species.

Panicle type is one of the important factors affecting rice (Oryza sativa L.) yield, and the identification of regulatory genes in panicle development can provide significant insights into the molecular network involved. This study identified a large and dense panicle 1 (ldp1) mutant produced from the Wuyunjing 7 (WYJ7) genotype, which displayed significant relative increases in panicle length, number of primary and secondary branches, number of grains per panicle, grain width, and grain yield per plant. Scanning electron microscopy results showed that the shoot apical meristem (SAM) of ldp1 was relatively larger at the bract stage (BM), with a significantly increased number of primary (PBM) and secondary branch (SBM) meristematic centers, indicating that the ldp1 mutation affects early stages in SAM development Comparative RNA-Seq analysis of meristem tissues from WYJ7 and ldp1 at the BM, PBM, and SBM developmental stages indicated that the number of differentially expressed genes (DEGs) were highest (1407) during the BM stage. Weighted gene coexpression network analysis (WGCNA) revealed that genes in one module (turquoise) are associated with the ldp1 phenotype and highly expressed during the BM stage, suggesting their roles in the identity transition and branch differentiation stages of rice inflorescences. Hub genes involved in auxin synthesis and transport pathways, such as OsAUX1, OsAUX4, and OsSAUR25, were identified. Moreover, GO and KEGG analysis of the DEGs in the turquoise module and the 1407 DEGs in the BM stage revealed that a majority of genes involved in tryptophan metabolism and auxin signaling pathway were differentially expressed between WYJ and ldp1. The genetic analysis indicated that the ldp1 phenotype is controlled by a recessive monogene (LDP1), which was mapped to a region between 16.9 and 18.1 Mb on chromosome seven. This study suggests that the ldp1 mutation may affect the expression of key genes in auxin synthesis and signal transduction, enhance the size of SAM, and thus affect panicle development. This study provides insights into the molecular regulatory network underlying rice panicle morphogenesis and lays an important foundation for further understanding the function and molecular mechanism of LDP1 during panicle development. [ABSTRACT FROM AUTHOR]

Abstract Leaf shape is considered to be one of the most significant agronomic traits in crop breeding. However, the molecular basis underlying leaf morphogenesis in cotton is still largely unknown. In this study, through genetic mapping and molecular investigation using a natural cotton mutant cu with leaves curling upward, the causal gene GHCU is successfully identified as the key regulator of leaf flattening. Knockout of GHCU or its homolog in cotton and tobacco using CRISPR results in abnormal leaf shape. It is further discovered that GHCU facilitates the transport of the HD protein KNOTTED1‐like (KNGH1) from the adaxial to the abaxial domain. Loss of GHCU function restricts KNGH1 to the adaxial epidermal region, leading to lower auxin response levels in the adaxial boundary compared to the abaxial. This spatial asymmetry in auxin distribution produces the upward‐curled leaf phenotype of the cu mutant. By analysis of single‐cell RNA sequencing and spatiotemporal transcriptomic data, auxin biosynthesis genes are confirmed to be expressed asymmetrically in the adaxial‐abaxial epidermal cells. Overall, these findings suggest that GHCU plays a crucial role in the regulation of leaf flattening through facilitating cell‐to‐cell trafficking of KNGH1 and hence influencing the auxin response level.

Abstract Background Tobacco mosaic virus (TMV) is a widely distributed viral disease that threatens many vegetables and horticultural species. Using the resistance gene N which induces a hypersensitivity reaction, is a common strategy for controlling this disease in tobacco (Nicotiana tabacum L.). However, N gene-mediated resistance has its limitations, consequently, identifying resistance genes from resistant germplasms and developing resistant cultivars is an ideal strategy for controlling the damage caused by TMV. Results Here, we identified highly TMV-resistant tobacco germplasm, JT88, with markedly reduced viral accumulation following TMV infection. We mapped and cloned two tobamovirus multiplication protein 2A (TOM2A) homeologs responsible for TMV replication using an F2 population derived from a cross between the TMV-susceptible cultivar K326 and the TMV-resistant cultivar JT88. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9)-mediated loss-of-function mutations of two NtTOM2A homeologs almost completely suppressed TMV replication; however, the single gene mutants showed symptoms similar to those of the wild type. Moreover, NtTOM2A natural mutations were rarely detected in 577 tobacco germplasms, and CRISPR/Cas9-mediated variation of NtTOM2A led to shortened plant height, these results indicating that the natural variations in NtTOM2A were rarely applied in tobacco breeding and the NtTOM2A maybe has an impact on growth and development. Conclusions The two NtTOM2A homeologs are functionally redundant and negatively regulate TMV resistance. These results deepen our understanding of the molecular mechanisms underlying TMV resistance in tobacco and provide important information for the potential application of NtTOM2A in TMV resistance breeding.

The yield of soybean is positively correlated with the duration of reproductive growth, which delays the aging of leaves after flowering, enhances their physiological performance, and supports growing plants with heavier grains. Leaf yellowing is one of the distinctive features of plant aging. Studies on leaf yellowing at the late stage of soybean drum grain have rarely been reported. The early yellowing feature of soybean late tympanic leaves was controlled by a single recessive nuclear gene, according to the genetic analysis of the hybrid of the early yellowing mutant ly and wild-type ofc in this study. Using Molecular Marker to Map-Based Cloning, a 2.23 Mb preliminary localization interval on chromosome 19 was obtained. The interval was shortened to 1.75 Mb and contained 219 genes. When this interval was combined with RNA-Seq analysis, 12 candidate genes were detected, including 4 SNP variant genes and 8 differentially expressed genes. The findings of this study provides the framework for the cloning of genes that cause aging and yellowing during the filling later period in soybean. [ABSTRACT FROM AUTHOR]

Background: Tobacco mosaic virus (TMV) is a widely distributed viral disease that threatens many vegetables and horticultural species. Using the resistance gene N which induces a hypersensitivity reaction, is a common strategy for controlling this disease in tobacco (Nicotiana tabacum L.). However, N gene-mediated resistance has its limitations, consequently, identifying resistance genes from resistant germplasms and developing resistant cultivars is an ideal strategy for controlling the damage caused by TMV. Results: Here, we identified highly TMV-resistant tobacco germplasm, JT88, with markedly reduced viral accumulation following TMV infection. We mapped and cloned two tobamovirus multiplication protein 2A (TOM2A) homeologs responsible for TMV replication using an F2 population derived from a cross between the TMV-susceptible cultivar K326 and the TMV-resistant cultivar JT88. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9)-mediated loss-of-function mutations of two NtTOM2A homeologs almost completely suppressed TMV replication; however, the single gene mutants showed symptoms similar to those of the wild type. Moreover, NtTOM2A natural mutations were rarely detected in 577 tobacco germplasms, and CRISPR/Cas9-mediated variation of NtTOM2A led to shortened plant height, these results indicating that the natural variations in NtTOM2A were rarely applied in tobacco breeding and the NtTOM2A maybe has an impact on growth and development. Conclusions: The two NtTOM2A homeologs are functionally redundant and negatively regulate TMV resistance. These results deepen our understanding of the molecular mechanisms underlying TMV resistance in tobacco and provide important information for the potential application of NtTOM2A in TMV resistance breeding. [ABSTRACT FROM AUTHOR]

SUMMARY: Seed color is one of the key target traits of domestication and artificial selection in chickpeas due to its implications on consumer preference and market value. The complex seed color trait has been well dissected in several crop species; however, the genetic mechanism underlying seed color variation in chickpea remains poorly understood. Here, we employed an integrated genomics strategy involving QTL mapping, high‐density mapping, map‐based cloning, association analysis, and molecular haplotyping in an inter‐specific RIL mapping population, association panel, wild accessions, and introgression lines (ILs) of Cicer gene pool. This delineated a MATE gene, CaMATE23, encoding a Transparent Testa (TT) and its natural allele (8‐bp insertion) and haplotype underlying a major QTL governing seed color on chickpea chromosome 4. Signatures of selective sweep and a strong purifying selection reflected that CaMATE23, especially its 8‐bp insertion natural allelic variant, underwent selection during chickpea domestication. Functional investigations revealed that the 8‐bp insertion containing the third cis‐regulatory RY‐motif element in the CaMATE23 promoter is critical for enhanced binding of CaFUSCA3 transcription factor, a key regulator of seed development and flavonoid biosynthesis, thereby affecting CaMATE23 expression and proanthocyanidin (PA) accumulation in the seed coat to impart varied seed color in chickpea. Consequently, overexpression of CaMATE23 in Arabidopsis tt12 mutant partially restored the seed color phenotype to brown pigmentation, ascertaining its functional role in PA accumulation in the seed coat. These findings shed new light on the seed color regulation and evolutionary history, and highlight the transcriptional regulation of CaMATE23 by CaFUSCA3 in modulating seed color in chickpea. The functionally relevant InDel variation, natural allele, and haplotype from CaMATE23 are vital for translational genomic research, including marker‐assisted breeding, for developing chickpea cultivars with desirable seed color that appeal to consumers and meet global market demand. Significance Statement: Integrated genomic strategy provides novel insights into the transcriptional regulatory role of functional allele and haplotype delineated in a MATE gene selected during Cicer domestication, in regulating varied seed color of chickpea. [ABSTRACT FROM AUTHOR]

Summary: Plant height and grain size are two important agronomic traits that are closely related to crop yield. Numerous dwarf and grain‐shape mutants have been studied to identify genes that can be used to increase crop yield and improve breeding programs.In this study, we characterized a dominant mutant, dwarf and round grain 1 (drg1‐D), in bread wheat (Triticum aestivum L.). drg1‐D plants exhibit multiple phenotypic changes, including dwarfism, round grains, and insensitivity to brassinosteroids (BR). Cell structure observation in drg1‐D mutant plants showed that the reduced organ size is due to irregular cell shape.Using map‐based cloning and verification in transgenic plants, we found that a Glu209Lys substitution in the DRG1 protein is responsible for the irregular cell size and arrangement in the drg1‐D mutant. DRG1/TaACT7 encodes an actin family protein that is essential for polymerization stability and microfilament (MF) formation. In addition, the BR response and vesicular transport were altered by the abnormal actin cytoskeleton in drg1‐D mutant plants.Our study demonstrates that DRG1/TaACT7 plays an important role in wheat cell shape determination by modulating actin organization and intracellular material transport, which could in the longer term provide tools to better understand the polymerization of actin and its assembly into filaments and arrays. [ABSTRACT FROM AUTHOR]

Significance Statement: Superior natural alleles of Mediator subunit genes have efficacy to develop desirable semi‐dwarf chickpea cultivars by modulating lignin and phenylpropanoid biosynthesis without compromising agronomic performance. [ABSTRACT FROM AUTHOR]

Chloroplast RNA splicing and ribosome maturation (CRM) domain proteins are a family of plant-specific proteins associated with RNA binding. In this study, we have conducted a detailed characterization of a novel rice CRM gene (LOC_Os04g39060) mutant, yl4, which showed yellow-green leaves at all the stages, had fewer tillers, and had a decreased plant height. Map-based cloning and CRISPR/Cas9 editing techniques all showed that YL4 encoded a CRM domain protein in rice. In addition, subcellular localization revealed that YL4 was in chloroplasts. YL4 transcripts were highly expressed in all leaves and undetectable in roots and stems, and the mutation of YL4 affected the transcription of chloroplast-development-related genes. This study indicated that YL4 is essential for chloroplast development and affects some agronomic traits.

Our study investigates the genetic mechanisms underlying the spotted leaf phenotype in rice, focusing on the spl43 mutant. This mutant is characterized by persistent reddish-brown leaf spots from the seedling stage to maturity, leading to extensive leaf necrosis. Using map-based cloning, we localized the responsible locus to a 330 Kb region on chromosome 2. We identified LOC_Os02g56000, named OsRPT5A, as the causative gene. A point mutation in OsRPT5A, substituting valine for glutamic acid, was identified as the critical factor for the phenotype. Functional complementation and the generation of CRISPR/Cas9-mediated knockout lines in the IR64 background confirmed the central role of OsRPT5A in controlling this trait. The qPCR results from different parts of the rice plant revealed that OsRPT5A is constitutively expressed across various tissues, with its subcellular localization unaffected by the mutation. Notably, we observed an abnormal accumulation of reactive oxygen species (ROS) in spl43 mutants by examining the physiological indexes of leaves, suggesting a disruption in the ROS system. Complementation studies indicated OsRPT5A’s involvement in ROS homeostasis and catalase activity regulation. Moreover, the spl43 mutant exhibited enhanced resistance to Xanthomonas oryzae pv. oryzae (Xoo), highlighting OsRPT5A’s role in rice pathogen resistance mechanisms. Overall, our results suggest that OsRPT5A plays a critical role in regulating ROS homeostasis and enhancing pathogen resistance in rice.

Plant architecture, one of the key factors that determine grain yield in rice, is mainly affected by components such as plant height, tiller number, and panicle morphology. For this paper, we obtained a multi-tillering and lazy mutant from a japonica rice cultivar, Wuyunjing 7 (WYJ7), via treatment with a heavy ion beam. Compared to WYJ7, the mutant showed a significant increase in tiller angle, tiller number, number of primary and secondary branches, and number of grains; however, the plant height and grain thickness of the mutant was significantly decreased. Phenotypic analysis of the F1 hybrids revealed that the multi-tillering and lazy mutant phenotypes were regulated by a recessive gene. The segregation ratio of 1׃3 of the mutant phenotype and the wild-type plant in the F2 population indicated that the former was controlled by a single gene named Multi-Tillering and Lazy 1 (MTL1). Bulked segregant analysis was performed using the individual plants with extremely typical tiller angles in the F2 population. The MTL1 gene was initially mapped within a region of 5.58–17.64 Mb on chromosome 11. By using the F2 segregated population for fine mapping, the MTL1 gene was ultimately fine mapped within the range of 66.67 kb on chromosome 11. The analysis of genes in this region revealed the presence of the previously identified LAZY1 (LA1) gene. Genomic PCR amplification and semi-quantitative RT-PCR assays showed that the LA1 gene could not be amplified and was not expressed, thus indicating that the MTL1 gene might be identical to the LA1 gene. This study suggests that the multi-tillering and lazy mutant phenotypes might be caused by the deletion of LA1 function. This finding can guide further investigations on the functional mechanisms of the LA1 gene, thus enriching the theoretical knowledge of plant architecture in relation to rice. [ABSTRACT FROM AUTHOR]

SUMMARY: Trichomes, the outward projection of plant epidermal tissue, provide an effective defense against stress and insect pests. Although numerous genes have been identified to be involved in trichome development, the molecular mechanism for trichome cell fate determination is not well enunciated. Here, we reported GoSTR functions as a master repressor for stem trichome formation, which was isolated by map‐based cloning based on a large F2 segregating population derived from a cross between TM‐1 (pubescent stem) and J220 (smooth stem). Sequence alignment revealed a critical G‐to‐T point mutation in GoSTR's coding region that converted codon 2 from GCA (Alanine) to TCA (Serine). This mutation occurred between the majority of Gossypium hirsutum with pubescent stem (GG‐haplotype) and G. barbadense with glabrous stem (TT‐haplotype). Silencing of GoSTR in J220 and Hai7124 via virus‐induced gene silencing resulted in the pubescent stems but no visible change in leaf trichomes, suggesting stem trichomes and leaf trichomes are genetically distinct. Yeast two‐hybrid assay and luciferase complementation imaging assay showed GoSTR interacts with GoHD1 and GoHOX3, two key regulators of trichome development. Comparative transcriptomic analysis further indicated that many transcription factors such as GhMYB109, GhTTG1, and GhMYC1/GhDEL65 which function as positive regulators of trichomes were significantly upregulated in the stem from the GoSTR‐silencing plant. Taken together, these results indicate that GoSTR functions as an essential negative modulator of stem trichomes and its transcripts will greatly repress trichome cell differentiation and growth. This study provided valuable insights for plant epidermal hair initiation and differentiation research. [ABSTRACT FROM AUTHOR]

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Panicle type is one of the important factors affecting rice (Oryza sativa L.) yield, and the identification of regulatory genes in panicle development can provide significant insights into the molecular network involved. This study identified a large and dense panicle 1 (ldp1) mutant produced from the Wuyunjing 7 (WYJ7) genotype, which displayed significant relative increases in panicle length, number of primary and secondary branches, number of grains per panicle, grain width, and grain yield per plant. Scanning electron microscopy results showed that the shoot apical meristem (SAM) of ldp1 was relatively larger at the bract stage (BM), with a significantly increased number of primary (PBM) and secondary branch (SBM) meristematic centers, indicating that the ldp1 mutation affects early stages in SAM development Comparative RNA-Seq analysis of meristem tissues from WYJ7 and ldp1 at the BM, PBM, and SBM developmental stages indicated that the number of differentially expressed genes (DEGs) were highest (1407) during the BM stage. Weighted gene coexpression network analysis (WGCNA) revealed that genes in one module (turquoise) are associated with the ldp1 phenotype and highly expressed during the BM stage, suggesting their roles in the identity transition and branch differentiation stages of rice inflorescences. Hub genes involved in auxin synthesis and transport pathways, such as OsAUX1, OsAUX4, and OsSAUR25, were identified. Moreover, GO and KEGG analysis of the DEGs in the turquoise module and the 1407 DEGs in the BM stage revealed that a majority of genes involved in tryptophan metabolism and auxin signaling pathway were differentially expressed between WYJ and ldp1. The genetic analysis indicated that the ldp1 phenotype is controlled by a recessive monogene (LDP1), which was mapped to a region between 16.9 and 18.1 Mb on chromosome seven. This study suggests that the ldp1 mutation may affect the expression of key genes in auxin synthesis and signal transduction, enhance the size of SAM, and thus affect panicle development. This study provides insights into the molecular regulatory network underlying rice panicle morphogenesis and lays an important foundation for further understanding the function and molecular mechanism of LDP1 during panicle development.

Summary: African cultivated rice (Oryza glaberrima Steud.) was domesticated from its wild progenitor species (Oryza barthii) about 3000 years ago. Seed shattering is one of the main constraints on grain production in African cultivated rice, which causes severe grain losses during harvest. By contrast, Asian cultivated rice (Oryza sativa) displays greater resistance to seed shattering, allowing higher grain production. A better understanding in regulation of seed shattering would help to improve harvesting efficiency in African cultivated rice. Here, we report the map‐based cloning and characterization of OgSH11, a MYB transcription factor controlling seed shattering in O. glaberrima. OgSH11 represses the expression of lignin biosynthesis genes and lignin deposition by binding to the promoter of GH2. We successfully developed a new O. glaberrima material showing significantly reduced seed shattering by knockout of SH11 in O. glaberrima using CRISPR‐Cas9 mediated approach. Identification of SH11 not only supplies a new target for seed shattering improvement in African cultivated rice, but also provides new insights into the molecular mechanism of abscission layer development. [ABSTRACT FROM AUTHOR]

In pumpkin (Cucurbita moschata), the naked or hull-less seed phenotype has great benefits for breeding this crop for oil or snack use. We previously identified a naked seed mutant in this crop. In this study, we report genetic mapping, identification, and characterization of a candidate gene for this mutation. We showed that the naked seed phenotype is controlled by a single recessive gene (N). The bulked segregant analysis identified a 2.4 Mb region on Chromosome 17 with 15 predicted genes. Multiple lines of evidence suggested that CmoCh17G004790 is the most probable candidate gene for the N locus which encodes a NAC transcription factor WALL THICKENING PROMOTING FACTOR 1 (CmNST1). No nucleotide polymorphism or structural variation was found in the genomic DNA sequences of CmNST1 between the mutant and the wildtype inbred line (hulled seed). However, the cDNA sequence cloned from developing seed coat samples of the naked seed mutant was 112 bp shorter than that from the wildtype which is due to seed coat-specific alternative splicing in the second exon of the mutant CmNST1 transcript. The expression level of CmNST1 in the developing seed coat was higher in the mutant than in the wildtype during early seed coat development which was reversed later. Transcriptomic profiling with RNA-Seq at different stages of seed development in the mutant and wildtype revealed a critical role of CmNST1 as a master regulator for the lignin biosynthesis pathway during seed coat development while other NAC and MYB transcription factors were also involved in forming a regulatory network for the building of secondary cell walls. This work provides a novel mechanism for the well-characterized NST1 transcription factor gene in regulating secondary cell wall development. The cloned gene also provides a useful tool for marker-assisted breeding of hull-less C. moschata varieties. [ABSTRACT FROM AUTHOR]

Chloroplasts are essential sites for plant photosynthesis, and the biogenesis of the photosynthetic complexes involves the interaction of nuclear genes and chloroplast genes. In this study, we identified a rice pale green leaf mutant, crs2. The crs2 mutant showed different degrees of low chlorophyll phenotypes at different growth stages, especially at the seedling stage. Fine mapping and DNA sequencing of crs2 revealed a single nucleotide substitution (G4120A) in the eighth exons of CRS2, causing a G-to-R mutation of the 229th amino acid of CRS2 (G229R). The results of complementation experiments confirmed that this single-base mutation in crs2 is responsible for the phenotype of the crs2 mutant. CRS2 encodes a chloroplast RNA splicing 2 protein localized in the chloroplast. Western blot results revealed an abnormality in the abundance of the photosynthesis-related protein in crs2. However, the mutation of CRS2 leads to the enhancement of antioxidant enzyme activity, which could reduce ROS levels. Meanwhile, with the release of Rubisco activity, the photosynthetic performance of crs2 was improved. In summary, the G229R mutation in CRS2 causes chloroplast protein abnormalities and affects photosystem performance in rice; the above findings facilitate the elucidation of the physiological mechanism of chloroplast proteins affecting photosynthesis. [ABSTRACT FROM AUTHOR]

In grass, the lemma is a unique floral organ structure that directly determines grain size and yield. Despite a great deal of research on grain enlargement caused by changes in glume cells, the importance of normal development of the glume for normal grain development has been poorly studied. In this study, we investigated a rice spikelet mutant, degenerated lemma (del), which developed florets with a slightly degenerated or rod-like lemma. More importantly, del also showed a significant reduction in grain length and width, seed setting rate, and 1000-grain weight, which led to a reduction in yield. The results indicate that the mutation of the DEL gene further affects rice grain yield. Map-based cloning shows a single-nucleotide substitution from T to A within Os01g0527600/DEL/OsRDR6, causing an amino acid mutation of Leu-34 to His-34 in the del mutant. Compared with the wild type, the expression of DEL in del was significantly reduced, which might be caused by single base substitution. In addition, the expression level of tasiR-ARF in del was lower than that of the wild type. RT-qPCR results show that the expression of some floral organ identity genes was changed, which indicates that the DEL gene regulates lemma development by modulating the expression of these genes. The present results suggest that the normal expression of DEL is necessary for the formation of lemma and the normal development of grain morphology and therefore has an important effect on the yield. [ABSTRACT FROM AUTHOR]

Summary: Anthocyanin accumulations in the flowers can improve seed production of hybrid lines, and produce higher commodity value in cotton fibre. However, the genetic mechanism underlying the anthocyanin pigmentation in cotton petals is poorly understood. Here, we showed that the red petal phenotype was introgressed from Gossypium bickii through recombination with the segment containing the R3bic region in the A07 chromosome of Gossypium hirsutum variety LR compared with the near‐isogenic line of LW with white flower petals. The cyanidin‐3‐O‐glucoside (Cy3G) was the major anthocyanin in red petals of cotton. A GhTT19 encoding a TT19‐like GST was mapped to the R3bic site associated with red petals via map‐based cloning, but GhTT19 homologue gene from the D genome was not expressed in G. hirsutum. Intriguingly, allelic variations in the promoters between GhTT19LW and GhTT19LR, rather than genic regions, were found as genetic causal of petal colour variations. GhTT19‐GFP was found localized in both the endoplasmic reticulum and tonoplast for facilitating anthocyanin transport. An additional MYB binding element found only in the promoter of GhTT19LR, but not in that of GhTT19LW, enhanced its transactivation by the MYB activator GhPAP1. The transgenic analysis confirmed the function of GhTT19 in regulating the red flower phenotype in cotton. The essential light signalling component GhHY5 bonded to and activated the promoter of GhPAP1, and the GhHY5‐GhPAP1 module together regulated GhTT19 expression to mediate the light‐activation of petal anthocyanin pigmentation in cotton. This study provides new insights into the molecular mechanisms for anthocyanin accumulation and may lay a foundation for faster genetic improvement of cotton. [ABSTRACT FROM AUTHOR]

Zinc (Zn) deficiency, caused by inadequate Zn intake in the human diet, has serious health implications. Rice (Oryza sativa) is the staple food in regions with a high incidence of Zn deficiency, so raising Zn levels in rice grain could help alleviate Zn deficiency. The wild relatives of cultivated rice vary widely in grain Zn content and thus are suitable resources for improving this trait. However, few loci underlying grain Zn content have been identified in wild rice relatives. Here, we identified a major quantitative trait locus for grain Zn content, Grain Zn Content 1 (qGZnC1), from Yuanjiang common wild rice (Oryza rufipogon Griff.) using map-based cloning. Down-regulating GZnC1 expression reduced the grain Zn content, whereas the presence of GZnC1 had the opposite effect, indicating that GZnC1 is involved in grain Zn content in rice. Notably, GZnC1 is identical to a previously reported gene, EMBRYO SAC ABORTION 1 (ESA1), involved in seed setting rate. The mutation in GZnC1/ESA1 at position 1819 (T1819C) causes delayed termination of protein translation. In addition, GZnC1 is specifically expressed in developing panicles. Several genes related to Zn-transporter genes were up-regulated in the presence of GZnC1. Our results suggest that GZnC1 activates Zn transporters to promote Zn distribution in panicles. Our work thus sheds light on the genetic mechanism of Zn accumulation in rice grain and provides a new genetic resource for improving Zn content in rice. Key message: We identified an allele from the rice progenitor Oryza rufipogon, GZnC1, that influences grain zinc content. This allele is pleiotropic and is identical to EMBRYO SAC ABORTION 1, which is implicated in decreased seed setting rate. [ABSTRACT FROM AUTHOR]

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To further understand the molecular mechanism for rice male reproduction, a rice male sterile mutant paa1 was screened from the rice mutant library generated by treatment with 60Coγ-rays. Genetic analysis revealed that paa1 is controlled by a single- recessive nuclear gene, and the anthers of the paa1 mutant were smaller than those of WT plants with a white color. Histological analysis demonstrated that the anthers of the paa1 mutant began to turn abnormal at the microspore stage after meiosis, with abnormal degradation of tapetum, deformed Ubisch bodies, and defective pollen exine. TUNEL assay results also confirmed the delay of tapetum PCD in paa1. Map-based cloning was performed for the PAA1 location. As a result, PAA1 was located in a 88-kb region at the end of chromosome 10, which comprises a total of seven candidate genes, and no genes related to anther development have been reported in this region. The results indicate that PAA1 is an essential gene in regulating tapetum development and pollen/microspore formation after rice meiosis. [ABSTRACT FROM AUTHOR]

The fruit shape of cucumber is an important agronomic trait, and mining regulatory genes, especially dominant ones, is vital for cucumber breeding. In this study, we identified a short and fat fruit mutant, named sff , from an EMS mutagenized population. Compared to the CCMC (WT), sff (MT) exhibited reduced fruit length and increased dimeter. Segregation analysis revealed that the sff phenotype is controlled by a semi-dominant single gene with dosage effects. Through map-based cloning, the SFF locus was narrowed down to a 52.6 kb interval with two SNPs (G651A and C1072T) in the second and third exons of CsaV3_1G039870 , which encodes an IQD family protein, CsSUN. The G651A within the IQ domain of CsSUN was identified as the unique SNP among 114 cucumber accessions, and it was the primary cause of the functional alteration in CsSUN. By generating CsSUN knockout lines in cucumber, we confirmed that CsSUN was responsible for sff mutant phenotype. The CsSUN is localized to the plasma membrane. CsSUN exhibited the highest expression in the fruit with lower expression in sff compared to WT. Histological observations suggest that the sff mutant phenotype is due to increased transverse cell division and inhibited longitudinal cell division. Transcriptome analysis revealed that CsSUN significantly affected the expression of genes related to cell division, expansion, and auxin signal transduction. This study unveils CsSUN's crucial role in shaping cucumber fruit and offers novel insights for cucumber breeding. • Identification of a mutant with short and fat fruit (sff) in cucumber. • Cucumber transformation revealed that CsSUN is the causal gene for short and fat fruit. • Two SNPs are present inside CsSUN , which encodes an IQD family protein. • The SNP (G651A) inside the IQ domain of CsSUN is the primary cause of the functional alteration in CsSUN. [ABSTRACT FROM AUTHOR]

Plant architecture, one of the key factors that determine grain yield in rice, is mainly affected by components such as plant height, tiller number, and panicle morphology. For this paper, we obtained a multi-tillering and lazy mutant from a japonica rice cultivar, Wuyunjing 7 (WYJ7), via treatment with a heavy ion beam. Compared to WYJ7, the mutant showed a significant increase in tiller angle, tiller number, number of primary and secondary branches, and number of grains; however, the plant height and grain thickness of the mutant was significantly decreased. Phenotypic analysis of the F1 hybrids revealed that the multi-tillering and lazy mutant phenotypes were regulated by a recessive gene. The segregation ratio of 1׃3 of the mutant phenotype and the wild-type plant in the F2 population indicated that the former was controlled by a single gene named Multi-Tillering and Lazy 1 (MTL1). Bulked segregant analysis was performed using the individual plants with extremely typical tiller angles in the F2 population. The MTL1 gene was initially mapped within a region of 5.58–17.64 Mb on chromosome 11. By using the F2 segregated population for fine mapping, the MTL1 gene was ultimately fine mapped within the range of 66.67 kb on chromosome 11. The analysis of genes in this region revealed the presence of the previously identified LAZY1 (LA1) gene. Genomic PCR amplification and semi-quantitative RT-PCR assays showed that the LA1 gene could not be amplified and was not expressed, thus indicating that the MTL1 gene might be identical to the LA1 gene. This study suggests that the multi-tillering and lazy mutant phenotypes might be caused by the deletion of LA1 function. This finding can guide further investigations on the functional mechanisms of the LA1 gene, thus enriching the theoretical knowledge of plant architecture in relation to rice.

A nuclear-encoded sigma (σ) factor is essential for the transcriptional regulation of plant chloroplast-encoded genes. Five putative maize σ factors have been identified by database searches, but their functions are unknown. We report a maize leaf color mutant etiolated/albino leaf 1 (eal1) that was derived from space mutation breeding. The eal1 mutant displays etiolated or albino leaves that then gradually turn to normal green at the seedling stage. The changes in eal1 leaf color are associated with changes in photosynthetic pigment content and chloroplast development. Map-based cloning revealed that a single amino-acid deletion changing Val480-Val481-Val482 to Val480-Val481, in the C-terminal domain σ4 of the putative σ factor ZmSig2A, is responsible for the eal1 mutation. In comparison with the expression level of the wild-type (WT) allele ZmSig2A+ in WT plants, much higher expression of the mutant allele ZmSig2AΔV in eal1 plants was detected before the eal1 plants turned to normal green. ZmSig2A shows the highest similarity to rice OsSig2A and Arabidopsis SIG2. Ectopic expression of ZmSig2A+ or ZmSig2AΔV driven by the cauliflower mosaic virus 35S promoter rescued the pale green leaf of the sig2 mutant, but ectopic expression of ZmSig2AΔV driven by the SIG2 promoter did not. We propose that the Val deletion generated a new weak allele of ZmSig2A that cannot completely abolish the ZmSig2A function. Some genes involved in chloroplast development and photosynthesis-associated nuclear genes showed significant expression differences between eal1 and WT plants. We conclude that ZmSig2A encoding a σ factor is essential for maize chloroplast development. The eal1 mutant with a weak allele of ZmSig2A represents a valuable genetic resource for investigating the regulation of ZmSig2A-mediated chloroplast development in maize. The eal1 mutation may be useful as a marker for early identification and elimination of false hybrids or transgene transmission in the application of genetic male sterility to commercial hybrid seed production.

The molecular mechanisms underlying anthocyanin-based flower coloration remain unknown in Brassica napus. To identify the key genes and metabolites associated with apricot and pink flower colors, metabolome, BSA-seq, and RNA-seq analyses were conducted on apricot-, pink-, yellow-, and white-flowered F2 B. napus. Yellow carotenoids and red anthocyanins were abundant in apricot petals, while colorless carotenoids and red anthocyanins accumulated in pink petals. Most carotenoid genes were not differentially regulated between apricot and yellow or between pink and white petals. Three regulator genes, BnaMYBL2 , BnaA07.PAP2 , and BnaTT8 , and structural genes in anthocyanin biosynthesis were dramatically enhanced in apricot and pink petals in comparison with yellow and white petals. Map-based cloning revealed that BnaA07.PAP2 is responsible for anthocyanin-based flower color and encodes a nucleus-localized protein predominantly expressed in apricot and pink flowers. Two insertions in the promoter region are responsible for the transcriptional activation of BnaA07.PAP2 in flowers. Introducing the BnaA07.PAP2 In-184-317 allele broadly activated the expression of anthocyanin-related genes and promoted anthocyanin accumulation in flowers, yielding color change from yellow to apricot. These findings illustrate the genetic basis of anthocyanin-based flower coloration and provide a valuable genetic resource for breeding varieties with novel flower colors in B. napus. [ABSTRACT FROM AUTHOR]

Leaf-color mutants play an important role in the study of chlorophyll metabolism, chloroplast development, and photosynthesis system. In this study, the yellow leaf 1 (yl1) rice mutant was identified from the ethyl methane sulfonate-treated mutant progeny of Lailong, a glutinous japonica rice landrace cultivated in Guizhou Province, China. Results showed that yl1 exhibited yellow leaves with decreased chlorophyll content throughout the growth period. Chloroplast development in the yl1 mutant was disrupted, and the grana lamellae was loosely packed and disordered. RNA sequencing and real-time quantitative polymerase chain reaction (qRT-PCR) analysis revealed that the chlorophyll synthesis-related genes OsCHLH, OsCHLM, OsCHLG, PORB, and YGL8, as well as the chloroplast development-related genes FtsZ, OsRpoTp, and RbcL, were down-regulated in the yl1 mutant. Genetic analysis revealed that the yellow leaf phenotype of yl1 was controlled by recessive nuclear gene. By employing the MutMap method, the mutation responsible for the phenotype was mapped to a 6.17 Mb region between 17.34 and 23.51 Mb on chromosome 3. Two non-synonymous single-nucleotide polymorphisms (SNPs) located in the gene locus LOC_Os03g31210 and LOC_Os03g36760 were detected in this region. The two SNPs were further confirmed by PCR and Sanger sequencing. The expression patterns of the two candidate genes indicated that LOC_Os03g36760 showed greater potential for functional verification. Subcellular protein localization revealed that the encoded product of LOC_Os03g36760 was localized in the nucleus, cytoplasm, and plasma membrane. These results will be useful for further characterization and cloning of the yl1 gene, and for research on the molecular mechanisms controlling biogenesis and chloroplast biochemical processes. [ABSTRACT FROM AUTHOR]

Background: As one of the most important staple food crops, rice produces large of agronomic biomass residues that contain lots of secondary cell walls (SCWs). Membrane trafcking plays key roles in SCWs biosynthesis, but information association membrane trafcking and SCWs formation in plants is limited. Results: In this study, we report the function characterization of a rice mutant, culm easily fragile 3 (cef3), that exhibits growth retardation and fragile culm phenotype with signifcantly altered cell wall composition and reduced second-ary wall thickness. Map-based cloning revealed that CEF3 encodes a homologous protein of Arabidopsis STOMATAL CYTOKINESIS DEFECTIVE2 (SCD2). The saccharifcation assays revealed that CEF3 mutation can improve biomass enzymatic saccharifcation. Expression pattern analysis indicated that CEF3 is ubiquitously expressed in many organs at diferent developmental stages. Subcellular localization revealed that CEF3 is a Golgi-localized protein. The FM4-64 uptake assay revealed CEF3 is involved in endocytosis. Furthermore, mutation of CEF3 not only afected cellulose synthesis-related genes expression, but also altered the abundance of cellulose synthase catalytic subunit 9 (OsC-ESA9) in the PM and in the endomembrane systems. Conclusions: This study has demonstrated that CEF3 participates in the membrane trafcking that is essential for normal cellulose and other polysaccharides biosynthesis of the secondary cell wall, thereby manipulation of CEF3 could alter cellulose content and enhance biomass enzymatic saccharifcation in rice plants. Therefore, the study of the function of CEF3 can provide a strategy for genetic modifcation of SCWs in bioenergy crops. [ABSTRACT FROM AUTHOR]

Summary: Lettuce (Lactuca sativa) is one of the most important vegetables worldwide and an ideal plant for producing protein drugs. Both well‐functioning chloroplasts that perform robust photosynthesis and small leaf angles that enable dense planting are essential for high yields. In this study, we used an F2 population derived from a cross between a lettuce cultivar with pale‐green leaves and large leaf angles to a cultivar with dark‐green leaves and small leaf angles to clone LsNRL4, which encodes an NPH3/RPT2‐Like (NRL) protein. Unlike other NRL proteins in lettuce, the LsNRL4 lacks the BTB domain. Knockout mutants engineered using CRISPR/Cas9 and transgenic lines overexpressing LsNRL4 verified that LsNRL4 contributes to chloroplast development, photosynthesis and leaf angle. The LsNRL4 gene was not present in the parent with pale‐green leaves and enlarged leaf angles. Loss of LsNRL4 results in the enlargement of chloroplasts, decreases in the amount of cellular space allocated to chloroplasts and defects in secondary cell wall biosynthesis in lamina joints. Overexpressing LsNRL4 significantly improved photosynthesis and decreased leaf angles. Indeed, the plant architecture of the overexpressing lines is ideal for dense planting. In summary, we identified a novel NRL gene that enhances photosynthesis and influences plant architecture. Our study provides new approaches for the breeding of lettuce that can be grown in dense planting in the open field or in modern plant factories. LsNRL4 homologues may also be used in other crops to increase photosynthesis and improve plant architecture. [ABSTRACT FROM AUTHOR]

In this study, we identified a novel glossy mutant from Chinese cabbage, named SD369, and all wax monomers longer than 26 carbons were significantly decreased. Inheritance analysis revealed that the glossy trait of SD369 was controlled by a single recessive locus, BrWAX3. We fine-mapped the BrWAX3 locus to an interval of 161.82 kb on chromosome A09. According to the annotated genome of Brassica rapa, Bra024749 (BrCER60.A09), encoding a β-ketoacyl-CoA synthase, was identified as the candidate gene. Expression analysis showed that BrCER60.A09 was significantly downregulated in all aerial organs of glossy plants. Subcellular localization indicated that the BrCER60.A09 protein functions in the endoplasmic reticulum. A 5567-bp insertion was identified in exon 1 of BrCER60.A09 in SD369, which lead to a premature stop codon, thus causing a loss of function of the BrCER60.A09 enzyme. Moreover, comparative transcriptome analysis revealed that the 'cutin, suberine, and wax biosynthesis' pathway was significantly enriched, and genes involved in this pathway were almost upregulated in glossy plants. Further, two functional markers, BrWAX3-InDel and BrWAX3-KASP1, were developed and validated. Overall, these results provide a new information for the cuticular wax biosynthesis and provide applicable markers for marker-assisted selection (MAS)-based breeding of Brassica rapa. [ABSTRACT FROM AUTHOR]

SUMMARY: Canola (Brassica napus) is an important oil crop worldwide. The seed‐setting rate (SS) is a critical factor in determining its yield, and the development of pistils affects pollination and seed sets. However, research on seed‐setting defects has been limited owing to difficulties in the identification of phenotypes, mutations, and complex genetic mechanisms. In this study, we found a stigma defect (sd) mutant in B. napus, which had no nectary. The SS of sd mutants in the field was approximately 93.4% lower than that of the wild type. Scanning and transmission electron microscopy imaging of sd mutants showed a low density of stigma papillary cells and stigma papillary cell vacuoles that disappeared 16 h after flowering. Genetic analysis of segregated populations showed that two recessive nuclear genes are responsible for the mutant phenotype of sd. Based on re‐sequencing and map‐based cloning, we reduced the candidate sites on ChrA07 (BnaSSA07) and ChrC06 (BnaSSC06) to 30 and 67 kb, including six and eight predicted genes, respectively. Gene analyses showed that a pair of CRABS CLAW (CRC) homeologous genes at BnaSSA07 and BnaSSC06 were associated with the development of carpel and nectary. BnaSSA07.CRC and BnaSSC06.CRC candidate genes were found to be expressed in flower organs only, with significant differences in their expression in the pistils of the near‐isogenic lines. DNA sequencing showed transposon insertions in the upstream region and intron of the candidate gene BnaSSA07.crc. We also found that BnaSSC06.crc exists widely in the natural population and we give possible reasons for its widespread existence. Significance Statement: The BnaCRCs of domesticated preference are located at A07 and C06 within 30‐ and 67‐kb regions, respectively, as determined by re‐sequencing and map‐based cloning. They were positively correlated with the Brassica napus seed‐setting rate. [ABSTRACT FROM AUTHOR]

Spike compactness (SC) is strongly associated with wheat (Triticum aestivum L.) grain yield. In this study, we conducted a quantitative trait locus (QTL) analysis using a doubled haploid (DH) population derived from a cross between two common wheat varieties with contrasting spike morphology, revealing 16 stable QTLs associated with SC. The effect of a major QTL, QSc.cau-6B.1, was validated in 231 F7 recombinant inbred lines (RILs) derived from the same cross as the DH population. Using two residual heterozygous lines (RHLs), we delimited QSc.cau-6B.1 to an approximately 0.5-Mbp physical interval containing four high-confidence genes. The tetratricopeptide repeat-TraesCS6B03G1214400 (TaTPR-B1) was the priority candidate gene according to sequence and expression variations between near-isogenic lines. Accordingly, TaTPR-B1 knockout in the common wheat variety 'CB037' significantly increased SC compared to the wild type (WT). Conversely, TaTPR-B1 overexpression in the common wheat variety 'Fielder' significantly decreased SC compared to the WT. Moreover, we developed a PCR-based marker targeting the 32-bp insertion/deletion (InDel) between the two TaTPR-B1 alleles, which could be practical and valuable in modern wheat breeding programs for diagnostic purposes. Collectively, these findings provide insight into the genetic basis of SC in common wheat and present a valuable target with a breeder-friendly diagnostic marker for gene pyramid breeding., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

DNA methylation patterns in plants are dynamically shaped by the antagonistic actions of DNA methylation and demethylation pathways. Although the DNA methylation pathway has been well studied, the DNA demethylation pathway, however, are not fully understood so far. To gain deeper insights into the mechanisms of DNA demethylation pathway, we conducted a genetic screening for proteins that were involved in preventing epigenetic gene silencing, and then the ones, which were also implicated in DNA demethylation pathway, were used for further studies. Eventually, a mutant with low luciferase luminescence (low LUC luminescence) was recovered, and named reduced LUC luminescence 6–1 (rll6-1). Map-based cloning revealed that rll6-1 mutation was located on chromosome 4, and there were a total of 10 candidate genes residing within such a region. Analyses of genome-wide methylation patterns of rll6-1 mutant showed that mutation of RLL6 locus led to 3,863 hyper-DMRs (DMRs for differentially methylated regions) throughout five Arabidopsis chromosomes, and elevated DNA methylation level of 2 × 35S promoter, which was similar to that found in the ros1 (repressor of silencing 1) mutant. Further analysis demonstrated that there were 1,456 common hyper-DMRs shared by rll6-1 and ros1-7 mutants, suggesting that both proteins acted together in a synergistic manner to remove DNA methylation. Further investigations demonstrated that mutation of RLL6 locus did not affect the expression of the four genes of the DNA glycosylase/lyase family. Thus, our results demonstrate that RLL6 locus-encoded protein not only participates in transcriptional anti-silencing of a transgene, but is also involved in DNA demethylation pathway.

Summary: Global warming is a major abiotic stress factor, which limit rice production. Exploiting the genetic basis of the natural variation in heat resistance at different reproductive stages among diverse exotic Oryza germplasms can help breeding heat‐resistant rice cultivars. Here, we identified a stable quantitative trait locus (QTL) for heat tolerance at the heading stage on chromosome 5 (qHTH5) in O. rufipogon Griff. The corresponding gene, HTH5, pertains to the pyridoxal phosphate‐binding protein PLPBP (formerly called PROSC) family, which is predicted to encode pyridoxal phosphate homeostasis protein (PLPHP) localized to the mitochondrion. Overexpression of HTH5 increased the seed‐setting rate of rice plants under heat stress at the heading stage, whereas suppression of HTH5 resulted in greater susceptibility to heat stress. Further investigation indicated that HTH5 reduces reactive oxygen species accumulation at high temperatures by increasing the heat‐induced pyridoxal 5'‐phosphate (PLP) content. Moreover, we found that two SNPs located in the HTH5 promoter region are involved with its expression level and associated with heat tolerance diversity. These findings suggest that the novel gene HTH5 might have great potential value for heightening rice tolerance to heat stress to the on‐going threat of global warming. [ABSTRACT FROM AUTHOR]

SUMMARY: Though Medicago truncatula Tnt1 mutants are widely used by researchers in the legume community, they are mainly used for reverse genetics because of the availability of the BLAST‐searchable large‐scale flanking sequence tags database. However, these mutants should have also been used extensively for forward genetic screens, an effort that has been hindered due to the lack of a compatible genetic crossing partner for the M. truncatula genotype R108, from which Tnt1 mutants were generated. In this study, we selected three Medicago HapMap lines (HM017, HM018 and HM022) and performed reciprocal genetic crosses with R108. After phenotypic analyses in F1 and F2 progenies, HM017 was identified as a compatible crossing partner with R108. By comparing the assembled genomic sequences of HM017 and R108, we developed and confirmed 318 Indel markers evenly distributed across the eight chromosomes of the M. truncatula genome. To validate the effectiveness of these markers, by employing the map‐based cloning approach, we cloned the causative gene in the dwarf mutant crs isolated from the Tnt1 mutant population, identifying it as gibberellin 3‐β‐dioxygenase 1, using some of the confirmed Indel markers. The primer sequences and the size difference of each marker were made available for users in the web‐based database. The identification of the crossing partner for R108 and the generation of Indel markers will enhance the forward genetics and the overall usage of the Tnt1 mutants. Significance Statement: We identified the Medicago truncatula HapMap line HM017 as a compatible crossing partner with the genotype R108 and Tnt1 mutants. We further developed more than 300 Indel markers with at least 100‐bp amplicon size difference between HM017 and R108. The developed resources are instrumental to enhancing the forward genetics of Tnt1 mutants. [ABSTRACT FROM AUTHOR]

Abstract Background Mustard (Brassica juncea) is an important economic vegetable, and some cultivars have purple leaves and accumulate more anthocyanins than the green. The genetic and evolution of purple trait in mustard has not been well studied. Result In this study, free-hand sections and metabolomics showed that the purple leaves of mustard accumulated more anthocyanins than green ones. The gene controlling purple leaves in mustard, Mustard Purple Leaves (MPL), was genetically mapped and a MYB113-like homolog was identified as the candidate gene. We identified three alleles of the MYB113-like gene, BjMYB113a from a purple cultivar, BjMYB113b and BjMYB113c from green cultivars. A total of 45 single nucleotide polymorphisms (SNPs) and 8 InDels were found between the promoter sequences of the purple allele BjMYB113a and the green allele BjMYB113b. On the other hand, the only sequence variation between the purple allele BjMYB113a and the green allele BjMYB113c is an insertion of 1,033-bp fragment in the 3’region of BjMYB113c. Transgenic assay and promoter activity studies showed that the polymorphism in the promoter region was responsible for the up-regulation of the purple allele BjMYB113a and high accumulation of anthocyanin in the purple cultivar. The up-regulation of BjMYB113a increased the expression of genes in the anthocyanin biosynthesis pathway including BjCHS, BjF3H, BjF3’H, BjDFR, BjANS and BjUGFT, and consequently led to high accumulation of anthocyanin. However, the up-regulation of BjMYB113 was compromised by the insertion of 1,033-bp in 3’region of the allele BjMYB113c. Conclusions Our results contribute to a better understanding of the genetics and evolution of the BjMYB113 gene controlling purple leaves and provide useful information for further breeding programs of mustard.

Abstract Background The rice (Oryza sativa) gene Xa7 has been hypothesized to be a typical executor resistance gene against Xanthomonas oryzae pv. oryzae (Xoo), and has conferred durable resistance in the field for decades. Its identity and the molecular mechanisms underlying this resistance remain elusive. Results Here, we filled in gaps of genome in Xa7 mapping locus via BAC library construction, revealing the presence of a 100-kb non-collinear sequence in the line IRBB7 compared with Nipponbare reference genomes. Complementary transformation with sequentially overlapping subclones of the BACs demonstrated that Xa7 is an orphan gene, encoding a small novel protein distinct from any other resistance proteins reported. A 27-bp effector binding element (EBE) in the Xa7 promoter is essential for AvrXa7-inducing expression model. XA7 is anchored in the endoplasmic reticulum membrane and triggers programmed cell death in rice and tobacco (Nicotiana benthamiana). The Xa7 gene is absent in most cultivars, landraces, and wild rice accessions, but highly homologs of XA7 were identified in Leersia perrieri, the nearest outgroup of the genus Oryza. Conclusions Xa7 acts as a trap to perceive AvrXa7 via EBEAvrXa7 in its promoter, leading to the initiation of resistant reaction. Since EBEAvrXa7 is ubiquitous in promoter of rice susceptible gene SWEET14, the elevated expression of which is conducive to the proliferation of Xoo, that lends a great benefit for the Xoo strains retaining AvrXa7. As a result, varieties harboring Xa7 would show more durable resistance in the field. Xa7 alleles analysis suggests that the discovery of new resistance genes could be extended beyond wild rice, to include wild grasses such as Leersia species.