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

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

Identification of QTL for kernel weight and size and analysis of the pentatricopeptide repeat (PPR) gene family in cultivated peanut (Arachis hypogaea L.).

Peanut (Arachis hypogaea L.) is an important oilseed crop worldwide. Improving its yield is crucial for sustainable peanut production to meet increasing food and industrial requirements. Deciphering the genetic control underlying peanut kernel weight and size, which are essential components of peanut yield, would facilitate high-yield breeding. A high-density single nucleotide polymorphism (SNP)-based linkage map was constructed using a recombinant inbred lines (RIL) population derived from a cross between the variety Yuanza9102 and a germplasm accession wt09-0023. Kernel weight and size quantitative trait loci (QTLs) were co-localized to a 0.16 Mb interval on Arahy07 using inclusive composite interval mapping (ICIM). Analysis of SNP, and Insertion or Deletion (INDEL) markers in the QTL interval revealed a gene encoding a pentatricopeptide repeat (PPR) superfamily protein as a candidate closely linked with kernel weight and size in cultivated peanut. Examination of the PPR gene family indicated a high degree of collinearity of PPR genes between A. hypogaea and its diploid progenitors, Arachis duranensis and Arachis ipaensis. The candidate PPR gene, Arahy.JX1V6X, displayed a constitutive expression pattern in developing seeds. These findings lay a foundation for further fine mapping of QTLs related to kernel weight and size, as well as validation of candidate genes in cultivated peanut.

The determination of peanut (Arachis hypogaea L.) pod-sizes during the rapid-growth stage by phytohormones.

BACKGROUND: Pod size is an important yield target trait for peanut breeding. However, the molecular mechanism underlying the determination of peanut pod size still remains unclear. RESULTS: In this study, two peanut varieties with contrasting pod sizes were used for comparison of differences on the transcriptomic and endogenous hormonal levels. Developing peanut pods were sampled at 10, 15, 20, 25 and 30 days after pegging (DAP). Our results showed that the process of peanut pod-expansion could be divided into three stages: the gradual-growth stage, the rapid-growth stage and the slow-growth stage. Cytological analysis confirmed that the faster increase of cell-number during the rapid-growth stage was the main reason for the formation of larger pod size in Lps. Transcriptomic analyses showed that the expression of key genes related to the auxin, the cytokinin (CK) and the gibberellin (GA) were mostly up-regulated during the rapid-growth stage. Meanwhile, the cell division-related differentially expressed genes (DEGs) were mostly up-regulated at 10DAP which was consistent with the cytological-observation. Additionally, the absolute quantification of phytohormones were carried out by liquid-chromatography coupled with the tandem-mass-spectrometry (LC-MS/MS), and results supported the findings from comparative transcriptomic studies. CONCLUSIONS: It was speculated that the differential expression levels of TAA1 and ARF (auxin-related), IPT and B-ARR (CK-related), KAO, GA20ox and GA3ox (GA-related), and certain cell division-related genes (gene-LOC112747313 and gene-LOC112754661) were important participating factors of the determination-mechanism of peanut pod sizes. These results were informative for the elucidation of the underlying regulatory network in peanut pod-growth and would facilitate further identification of valuable target genes.

Safety of peanut (Arachis hypogaea) allergen powder-dnfp in children and teenagers with peanut allergy: Pooled summary of phase 3 and extension trials.

BACKGROUND: Peanut (Arachis hypogaea) allergen powder-dnfp (PTAH; previously known as AR101) is a daily oral immunotherapy approved to mitigate allergic reactions after accidental peanut exposure in peanut-allergic individuals aged 4-17 years. OBJECTIVE: We sought to comprehensively summarize the PTAH safety profile for up to ∼2 years of treatment. METHODS: Safety and adverse event (AE) data from participants aged 4-17 years from 3 controlled, phase 3 and 2 open-label extension trials were pooled and assessed. RESULTS: Of the 944 individuals receiving ≥1 PTAH dose, median exposure was ∼49 weeks; most participants experienced ≥1 treatment-related AE (TRAE; n = 853; 90.4%). A total of 829 participants experienced TRAEs with a maximum severity of mild (497, 52.6%) or moderate (332, 35.2%); 24 participants (2.5%) experienced TRAEs graded as severe. Overall, 80 participants (9.5%) discontinued as a result of AEs; most experienced gastrointestinal symptoms and discontinued during the first 6 months. When adjusted for exposure, AEs and TRAEs occurred at a rate of 76.4 and 58.7 events per participant-year of exposure (PYE), respectively, during updosing; AEs and TRAEs decreased to 23.0 and 14.2, respectively, during 300 mg maintenance. Overall, exposure-adjusted rates of systemic allergic reactions were 0.12 events/PYE (mild), 0.11 events/PYE (moderate), and 0.01 events/PYE (severe [anaphylaxis]). CONCLUSION: The safety profile of PTAH was consistent across trials, manageable, and improved over time. AEs were predominantly mild to moderate, and all grades declined in frequency with continued treatment. These data can be used to facilitate shared decision-making discussions with patients and families considering treatment with PTAH.

GWAS and bulked segregant analysis reveal the Loci controlling growth habit-related traits in cultivated Peanut (Arachis hypogaea L.).

BACKGROUND: Peanut (Arachis hypogaea L.) is a grain legume crop that originated from South America and is now grown around the world. Peanut growth habit affects the variety's adaptability, planting patterns, mechanized harvesting, disease resistance, and yield. The objective of this study was to map the quantitative trait locus (QTL) associated with peanut growth habit-related traits by combining the genome-wide association analysis (GWAS) and bulked segregant analysis sequencing (BSA-seq) methods. RESULTS: GWAS was performed with 17,223 single nucleotide polymorphisms (SNPs) in 103 accessions of the U.S. mini core collection genotyped using an Affymetrix version 2.0 SNP array. With a total of 12,342 high-quality polymorphic SNPs, the 90 suggestive and significant SNPs associated with lateral branch angle (LBA), main stem height (MSH), lateral branch height (LBL), extent radius (ER), and the index of plant type (IOPT) were identified. These SNPs were distributed among 15 chromosomes. A total of 597 associated candidate genes may have important roles in biological processes, hormone signaling, growth, and development. BSA-seq coupled with specific length amplified fragment sequencing (SLAF-seq) method was used to find the association with LBA, an important trait of the peanut growth habit. A 4.08 Mb genomic region on B05 was associated with LBA. Based on the linkage disequilibrium (LD) decay distance, we narrowed down and confirmed the region within the 160 kb region (144,193,467-144,513,467) on B05. Four candidate genes in this region were involved in plant growth. The expression levels of Araip.E64SW detected by qRT-PCR showed significant difference between 'Jihua 5' and 'M130'. CONCLUSIONS: In this study, the SNP (AX-147,251,085 and AX-144,353,467) associated with LBA by GWAS was overlapped with the results in BSA-seq through combined analysis of GWAS and BSA-seq. Based on LD decay distance, the genome range related to LBA on B05 was shortened to 144,193,467-144,513,467. Three candidate genes related to F-box family proteins (Araip.E64SW, Araip.YG1LK, and Araip.JJ6RA) and one candidate gene related to PPP family proteins (Araip.YU281) may be involved in plant growth and development in this genome region. The expression analysis revealed that Araip.E64SW was involved in peanut growth habits. These candidate genes will provide molecular targets in marker-assisted selection for peanut growth habits.

Priming of rhizobial nodulation signaling in the mycosphere accelerates nodulation of legume hosts.

The simultaneous symbiosis of leguminous plants with two root mutualists, endophytic fungi and rhizobia is common in nature, yet how two mutualists interact and co-exist before infecting plants and the concomitant effects on nodulation are less understood. Using a combination of metabolic analysis, fungal deletion mutants and comparative transcriptomics, we demonstrated that Bradyrhizobium and a facultatively biotrophic fungus, Phomopsis liquidambaris, interacted to stimulate fungal flavonoid production, and thereby primed Bradyrhizobial nodulation signaling, enhancing Bradyrhizobial responses to root exudates and leading to early nodulation of peanut (Arachis hypogaea), and such effects were compromised when disturbing fungal flavonoid biosynthesis. Stress sensitivity assays and reactive oxygen species (ROS) determination revealed that flavonoid production acted as a strategy to alleviate hyphal oxidative stress during P. liquidambaris-Bradyrhizobial interactions. By investigating the interactions between P. liquidambaris and a collection of 38 rhizobacteria, from distinct bacterial genera, we additionally showed that the flavonoid-ROS module contributed to the maintenance of fungal and bacterial co-existence, and fungal niche colonization under soil conditions. Our results demonstrate for the first time that rhizobial nodulation signaling can be primed by fungi before symbiosis with host plants and highlight the importance of flavonoid in tripartite interactions between legumes, beneficial fungi and rhizobia.

Nodule INception-independent epidermal events lead to bacterial entry during nodule development in peanut (Arachis hypogaea).

Legumes can host nitrogen-fixing rhizobia inside root nodules. In model legumes, rhizobia enter via infection threads (ITs) and develop nodules in which the infection zone contains a mixture of infected and uninfected cells. Peanut (Arachis hypogaea) diversified from model legumes c. 50-55 million years ago. Rhizobia enter through 'cracks' to form nodules in peanut roots where cells of the infection zone are uniformly infected. Phylogenomic studies have indicated symbiosis as a labile trait in peanut. These atypical features prompted us to investigate the molecular mechanism of peanut nodule development. Combining cell biology, genetics and genomic tools, we visualized the status of hormonal signaling in peanut nodule primordia. Moreover, we dissected the signaling modules of Nodule INception (NIN), a master regulator of both epidermal infection and cortical organogenesis. Cytokinin signaling operates in a broad zone, from the epidermis to the pericycle inside nodule primordia, while auxin signaling is narrower and focused. Nodule INception is involved in nodule organogenesis, but not in crack entry. Nodulation Pectate Lyase, which remodels cell walls during IT formation, is not required. By contrast, Nodule enhanced Glycosyl Hydrolases (AhNGHs) are recruited for cell wall modification during crack entry. While hormonal regulation is conserved, the function of the NIN signaling modules is diversified in peanut.

Comparative Multi-Omics Analysis Reveals Lignin Accumulation Affects Peanut Pod Size.

Pod size is one of the important factors affecting peanut yield. However, the metabolites relating to pod size and their biosynthesis regulatory mechanisms are still unclear. In the present study, two peanut varieties (Tif and Lps) with contrasting pod sizes were used for a comparative metabolome and transcriptome analysis. Developing peanut pods were sampled at 10, 20 and 30 days after pegging (DAP). A total of 720 metabolites were detected, most of which were lipids (20.3%), followed by phenolic acids (17.8%). There were 43, 64 and 99 metabolites identified as differentially accumulated metabolites (DAMs) at 10, 20 and 30 DAP, respectively, and flavonoids were the major DAMs between Tif and Lps at all three growth stages. Multi-omics analysis revealed that DAMs and DEGs (differentially expressed genes) were significantly enriched in the phenylpropanoid biosynthesis (ko00940) pathway, the main pathway of lignin biosynthesis, in each comparison group. The comparisons of the metabolites in the phenylpropanoid biosynthesis pathway accumulating in Tif and Lps at different growth stages revealed that the accumulation of p-coumaryl alcohol (H-monolignol) in Tif was significantly greater than that in Lps at 30 DAP. The differential expression of gene-LOC112771695, which is highly correlated with p-coumaryl alcohol and involved in the biosynthesis of monolignols, between Tif and Lps might explain the differential accumulation of p-coumaryl alcohol. The content of H-lignin in genetically diverse peanut varieties demonstrated that H-lignin content affected peanut pod size. Our findings would provide insights into the metabolic factors influencing peanut pod size and guidance for the genetic improvement of the peanut.

Optimization of Protoplast Isolation and Transformation for a Pilot Study of Genome Editing in Peanut by Targeting the Allergen Gene Ara h 2.

The cultivated peanut (Arachis hypogaea L.) is a legume consumed worldwide in the form of oil, nuts, peanut butter, and candy. Improving peanut production and nutrition will require new technologies to enable novel trait development. Clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9 (CRISPR-Cas9) is a powerful and versatile genome-editing tool for introducing genetic changes for studying gene expression and improving crops, including peanuts. An efficient in vivo transient CRISPR-Cas9- editing system using protoplasts as a testbed could be a versatile platform to optimize this technology. In this study, multiplex CRISPR-Cas9 genome editing was performed in peanut protoplasts to disrupt a major allergen gene with the help of an endogenous tRNA-processing system. In this process, we successfully optimized protoplast isolation and transformation with green fluorescent protein (GFP) plasmid, designed two sgRNAs for an allergen gene, Ara h 2, and tested their efficiency by in vitro digestion with Cas9. Finally, through deep-sequencing analysis, several edits were identified in our target gene after PEG-mediated transformation in protoplasts with a Cas9 and sgRNA-containing vector. These findings demonstrated that a polyethylene glycol (PEG)-mediated protoplast transformation system can serve as a rapid and effective tool for transient expression assays and sgRNA validation in peanut.

First comprehensive proteomics analysis of lysine crotonylation in leaves of peanut (Arachis hypogaea L.).

Lysine crotonylation is an important post-translational modification process. Most research in this area has been carried out on mammals and yeast, but there has been little research on it in plants. In the current study, large-scale lysine crotonylome analysis was performed by a combination of affinity enrichment and high-resolution LC-MS/MS analysis. Altogether, 6051 lysine crotonylation sites were identified in 2508 protein groups. Bioinformatics analysis showed that lysine-crotonylated proteins were involved in many biological processes, such as carbon fixation in photosynthetic organisms, biosynthesis of amino acids, ribosomes structure and function. In particular, subcellular localization analysis showed that 43% of the crotonylated proteins were located in the chloroplast. Twenty-nine crotonylation proteins were associated with photosynthesis and functional enrichment that these proteins were associated with the reaction center, photosynthetic electron transport, and ATP synthesis. Based on these results, further studies to expand on the lysine crotonylome analysis were suggested.