ACO Genes Research Articles

Insights into ACO genes across Rosaceae: evolution, expression, and regulatory networks in fruit development.

ACO (1-aminocyclopropane-1-carboxylic acid) serves as a pivotal enzyme within the plant ethylene synthesis pathway, exerting influence over critical facets of plant biology such as flowering, fruit ripening, and seed development. This study aims to identify ACO genes from representative Rosaceae genomes, reconstruct their phylogenetic relationships by integrating synteny information, and investigate their expression patterns and networks during fruit development. we utilize a specialized Hidden Markov Model (HMM), crafted on the sequence attributes of ACO gene-encoded proteins, to systematically identify and analyze ACO gene family members across 12 representative species within the Rosaceae botanical family. Through transcriptome analysis, we delineate the expression patterns of ACO genes in six distinct Rosaceae fruits. Our investigation reveals the presence of 62 ACO genes distributed among the surveyed Rosaceae species, characterized by hydrophilic proteins predominantly expressed within the cytoplasm. Phylogenetic analysis categorizes these ACO genes into three discernible classes, namely Class I, Class II, and Class III. Further scrutiny via collinearity assessment indicates a lack of collinearity relationships among these classes, highlighting variations in conserved motifs and promoter types within each class. Transcriptome analysis unveils significant disparities in both expression levels and trends of ACO genes in fruits exhibiting respiratory bursts compared to those that do not. Employing Weighted Gene Co-Expression Network Analysis (WGCNA), we discern that the co-expression correlation of ACO genes within loquat fruit notably differs from that observed in apples. Our findings, derived from Gene Ontology (GO) enrichment results, signify the involvement of ACO genes and their co-expressed counterparts in biological processes linked to terpenoid metabolism and carbohydrate synthesis in loquat. Moreover, our exploration of gene regulatory networks (GRN) highlights the potential pivotal role of the GNAT transcription factor (Ejapchr1G00010380) in governing the overexpression of the ACO gene (Ejapchr10G00001110) within loquat fruits. The constructed HMM of ACO proteins offers a precise and systematic method for identifying plant ACO proteins, facilitating phylogenetic reconstruction. ACO genes from representative Rosaceae fruits exhibit diverse expression and regulative patterns, warranting further function characterizations.

AMF inhibit the production of phenolic acid autotoxins at the seed-filling stage in soybeans with continuous monocropping

BackgroundSoybean is the main oil crop in Northeast China. Continuous monocropping is more commonly used for soybean production due to rising market demand and arable land constraints. However, autotoxic substances, such as phenolic acids, produced by continuously cropped soybean can reduce yield and quality. The mycorrhiza formed of Arbuscular mycorrhizal fungi (AMF) and plant roots regulate the metabolic activities of the host plant and increase its disease resistance. The main purpose of this study was to inhibit the production of phenolic acids and determine the adverse effects on the growth of continuous monocropping soybean by inoculating Funneliformis mosseae (F. mosseae).ResultsTranscriptomics results showed that the production of phenolic acids in continuous monocropping soybean roots was mainly regulated by the expression of the CHS6, PCL1, SAMT, SRG1, and ACO1 genes, and the expression of these genes was significantly downregulated after inoculation with F. mosseae. Metabolomics results showed that continuous monocropping soybean roots inoculated with F. mosseae inhibited phenolic acid production through the phenylpropane biosynthetic, α-linoleic acid, linoleic acid, and other metabolic pathways. Phenolic acids in the phenylpropane metabolic pathway, such as 4-hydroxybenzoic acid, phthalic acid, and vanillic acid, decreased significantly after inoculation with F. mosseae. The combined analysis of the two showed that genes such as YLS9 and ARF3 were positively correlated with 4-hydroxybenzoic acid and so on, while genes such as CHS6 and SRG1 were negatively correlated with butyric acid and so on.ConclusionF. mosseae regulated the expression of functional genes and related phenolic acid metabolic pathways produced by continuous monocropping soybean roots, inhibiting the production of phenolic acid autotoxic substances in continuous cropped soybean, and slowing down the disturbance of continuous monocropping. This study provides a new solution for continuous monocropping of plants to overcome the autotoxicity barrier and provides a new basis for the development and utilization of AMF as a biological agent.

Optimization of Phloretin-loaded Nanospanlastics for Targeting of FAS/SREBP1c/AMPK/ OB-Rb Signaling Pathway in HFD-induced Obesity.

Obese patients are at increased risk for CVD, which is the main cause of premature death and has been a major cause of disability and ill health in recent years. PTN, a natural dihydrochalcone flavonoid, has a variety of pharmacological characteristics. This article aimed to prepare PTN-NSLs to evaluate their anti-obesity activity. Morphology, Particle size, zeta potential, UV-vis, entrapment efficiency, FT-IR spectra, and an in vitro release study of PTN-NSLs were described. PTN-NSLs were also tested for their anti-obesity properties in obese rats. The LD50 of PTN-NSLs was calculated, as was the 1/20 LD50 prepared for the treatment of obese rats. Also, the level of glycemic, oxidative stress and inflammatory biomarkers were estimated in the obese rat's model. The synthesized PTN-NSLs were uniform, spherically shaped, and well dispersed with no aggregation noted, with a size range of 114.06 ± 8.35 nm. The measured zeta potential value of PTN-NSLs was -32.50.8 mv. Also, the UV spectra of PTN and PTN-NSLs have strong absorption at 225 and 285 nm. Also, the LD50 of PTN-NSLs was found to be 2750 mg/kg.b.w. Moreover, administrating obese rats with PTN-NSLs resulted in improved glycemic features as well as GSH, SOD, GPx, GR, IL10, TBARs, and IL-6 levels, as well as attenuated FAS, SREBP1c, AMPK, ACO, CPT1, and OB-Rb gene expression. Administration of PTN-NSLs significantly attenuated the levels of glycemic, oxidative stress, and inflammatory biomarkers. The biochemical and PCR findings are aided by histological investigations. Also, the present findings imply that PTN-NSLs might be a promising pharmacological tool for the treatment of obesity-related diseases.

The transcription factor WRKY22 modulates ethylene biosynthesis and root development through transactivating the transcription of ACS5 and ACO5 in Arabidopsis.

The WRKY transcription factor (TF) genes form a large family in higher plants, with 72 members in Arabidopsis (Arabidopsis thaliana). The gaseous phytohormone ethylene (ET) regulates multiple physiological processes in plants. It is known that 1-aminocyclopropane-1-carboxylic acid (ACC) synthases (ACSs, EC 4.4.1.14) limit the enzymatic reaction rate of ethylene synthesis. However, whether WRKY TFs regulate the expression of ACSs and/or ACC oxidases (ACOs, EC 1.14.17.4) remains largely elusive. Here, we demonstrated that Arabidopsis WRKY22 positively regulated the expression of a few ACS and ACO genes, thus promoting ethylene production. Inducible overexpression of WRKY22 caused shorter hypocotyls without ACC treatment. A qRT-PCR screening demonstrated that overexpression of WRKY22 activates the expression of several ACS and ACO genes. The promoter regions of ACS5, ACS11, and ACO5 were also activated by WRKY22, which was revealed by a dual luciferase reporter assay. A follow-up chromatin immunoprecipitation coupled with quantitative PCR (ChIP-qPCR) and electrophoretic mobility shift assay (EMSA) showed that the promoter regions of ACS5 and ACO5 could be bound by WRKY22 directly. Moreover, wrky22 mutants had longer primary roots and more lateral roots than wild type, while WRKY22-overexpressing lines showed the opposite phenotype. In conclusion, this study revealed that WRKY22 acts as a novel TF activating, at least, the expression of ACS5 and ACO5 to increase ethylene synthesis and modulate root development.

Characterization and functional analysis of gerbera plant defensin (PDF) genes reveal the role of GhPDF2.4 in defense against the root rot pathogen Phytophthora cryptogea

Gerbera (Gerbera hybrida), a major fresh cut flower crop, is very susceptible to root rot disease. Although plant defensins (PDFs), a major group of plant antimicrobial peptides, display broad-spectrum antifungal and antibacterial activities, PDF genes in gerbera have not been systematically characterized. Here, we identified and cloned nine PDF genes from gerbera and divided them into two classes based on phylogenetic analysis. Most Class I GhPDF genes were highly expressed in petioles, whereas all Class II GhPDF genes were highly expressed in roots. Phytophthora cryptogea inoculation strongly upregulated all Class II GhPDF genes in roots and upregulated all Class I GhPDF genes in petioles. Transient overexpression of GhPDF1.5 and GhPDF2.4 inhibited P. cryptogea infection in tobacco (Nicotiana benthamiana) leaves. Transient overexpression of GhPDF2.4, but not GhPDF1.5, significantly upregulated ACO and LOX gene expression in tobacco leaves, indicating that overexpressing GhPDF2.4 activated the jasmonic acid/ethylene defense pathway and that the two types of GhPDFs have different modes of action. Prokaryotically expressed recombinant GhPDF2.4 inhibited mycelial growth and delayed the hyphal swelling of P. cryptogea, in vitro, indicating that GhPDF2.4 is a morphogenetic defensin. Moreover, the addition of GhPDF2.4 to plant culture medium alleviated the root rot symptoms of in vitro-grown gerbera seedlings and greatly reduced pathogen titer in P. cryptogea-inoculated gerbera roots in the early stages of treatment. Our study provides a basis for the use of GhPDFs, especially GhPDF2.4, for controlling root rot disease in gerbera.

Detoxification of fermentation inhibitors using a waste biomass-based adsorbent of AEPA250 to enhance bioethanol production: The adsorption analysis and microbial mechanism

Environmental pollution and energy scarcity have promoted the development of renewable alternative energy of rice straw-based bioethanol. However, the production of main fermentation inhibitors from the pretreatment of rice straw raw material reduces the bioethanol yield. In order to improve bioethanol production, a waste biomass-based adsorbent (AEPA250) was used to adsorb three main inhibitors of ferulic acid, vanillin, and furfural from the rice straw hydrolysate in this study. The adsorption percentages were calculated using the high-performance liquid chromatography method. The results showed that the adsorption percentages of three inhibitors were all above 70% in 10 min, demonstrating the efficient detoxification of AEPA250 on rice straw hydrolysate. The adsorption contributions related to the attribute features of three inhibitors and AEPA250 were evaluated for the first time. The density functional theory calculations and AEPA250 characterizations were used to obtain these attribute parameters. Analysis of the correlation heat map between adsorption efficiencies and attribute features of AEPA250 (or fermentation inhibitors) indicated that AEPA250 possessed the lower activation energy (Ea) and pre-exponential factor (A) when it was prepared, as well as fermentation inhibitors had the higher ionization constant (pKa) and lower electrophilicity index (ω), resulting in the enhancement of bioethanol production. The contribution of pKa was greater than A followed by Ea and ω for adsorption process. Meanwhile, the key metabolic pathways involved in central carbon metabolism were analyzed for AEPA250 detoxification using proteomics method. The results indicated that the up-regulation of SDH3, ACO1, CIT2 and MLS1 (or down-regulation of SCT1) genes contributed to inhibitors detoxification using AEPA250, which effectively provided more nicotinamide adenine dinucleotide (NADH) for the reduction of these inhibitors. This study explores the detoxification mechanisms at the molecular level for boosting bioethanol production in the future, which lays the foundation for the development of efficient adsorbents and resistant engineering strains.

Characterization and Potential Action Mode Divergences of Homologous ACO1 Genes during the Organ Development and Ripening Process between Non-Climacteric Grape and Climacteric Peach.

Ethylene is one crucial phytohormone modulating plants' organ development and ripening process, especially in fruits, but its action modes and discrepancies in non-climacteric grape and climacteric peach in these processes remain elusive. This work is focused on the action mode divergences of ethylene during the modulation of the organ development and ripening process in climacteric/non-climacteric plants. We characterized the key enzyme genes in the ethylene synthesis pathway, VvACO1 and PpACO1, and uncovered that their sequence structures are highly conserved, although their promoters exhibit important divergences in the numbers and types of the cis-elements responsive to hormones, implying various responses to hormone signals. Subsequently, we found the two have similar expression modes in vegetative organ development but inverse patterns in reproductive ones, especially in fruits. Then, VvACO1 and PpACO1 were further validated in promoting fruit ripening functions through their transient over-expression/RNAi-expression in tomatoes, of which the former possesses a weaker role than the latter in the fruit ripening process. Our findings illuminated the divergence in the action patterns and function traits of the key VvACO1/PpACO1 genes in the tissue development of climacteric/non-climacteric plants, and they have implications for further gaining insight into the interaction mechanism of ethylene signaling during the modulation of the organ development and ripening process in climacteric/non-climacteric plants.

Genome-wide in-silico analysis of ethylene biosynthesis gene family in Musa acuminata L. and their response under nutrient stress

Ethylene is a gaseous phytohormone involved in plants’ growth and developmental processes, including seed germination, root initiation, fruit ripening, flower and leaf senescence, abscission, and stress responses. Ethylene biosynthesis (EB) gene analysis in response to nitrogen (N) and potassium (K) stress has not yet been conducted in Musa acuminata (banana) roots. The genome mining of banana (Musa acuminata L.) revealed 14 putative 1-aminocyclopropane-1-carboxylate synthase (ACS), 10 1-aminocyclopropane-1-carboxylate oxidase (ACO), and 3 Ethylene overproducer 1 (ETO1) genes. ACS, ACO, and ETO1 proteins possessed amino acid residues ranging from 422–684, 636–2670, and 893–969, respectively, with molecular weight (Mw) ranging from 4.93–7.55 kD, 10.1–8.3 kD and 10.1–10.78 kD. The number of introns present in ACS, ACO, and ETO1 gene sequences ranges from 0–14, 1–6, and 0–6, respectively. The cis-regulatory element analysis revealed the presence of light-responsive, abscisic acid, seed regulation, auxin-responsive, gibberellin element, endosperm-specific, anoxic inducibility, low-temperature responsiveness, salicylic acid responsiveness, meristem-specific and stress-responsive elements. Comprehensive phylogenetic analyses ACS, ACO, and ETO1 genes of Banana with Arabidopsis thaliana revealed several orthologs and paralogs assisting in understanding the putative functions of these genes. The expression profile of Musa acuminata genes in root under normal and low levels of nitrogen and potassium shows that MaACS14 and MaACO6 expressed highly at normal nitrogen supply. MaACS1 expression was significantly upregulated at low potassium levels, whereas, MaACO6 gene expression was significantly downregulated. The functional divergence and site-specific selective pressures on specific gene sequences of banana have been investigated. The bioinformatics-based genome-wide assessment of the family of banana attempted in the present study could be a significant step for deciphering novel ACS, ACO, and ETO1 genes based on genome-wide expression profiling.

Integration of Metabolomics and Transcriptomics to Explore Dynamic Alterations in Fruit Color and Quality in 'Comte de Paris' Pineapples during Ripening Processes.

Pineapple color yellowing and quality promotion gradually manifest as pineapple fruit ripening progresses. To understand the molecular mechanism underlying yellowing in pineapples during ripening, coupled with alterations in fruit quality, comprehensive metabolome and transcriptome investigations were carried out. These investigations were conducted using pulp samples collected at three distinct stages of maturity: young fruit (YF), mature fruit (MF), and fully mature fruit (FMF). This study revealed a noteworthy increase in the levels of total phenols and flavones, coupled with a concurrent decline in lignin and total acid contents as the fruit transitioned from YF to FMF. Furthermore, the analysis yielded 167 differentially accumulated metabolites (DAMs) and 2194 differentially expressed genes (DEGs). Integration analysis based on DAMs and DEGs revealed that the biosynthesis of plant secondary metabolites, particularly the flavonol, flavonoid, and phenypropanoid pathways, plays a pivotal role in fruit yellowing. Additionally, RNA-seq analysis showed that structural genes, such as FLS, FNS, F3H, DFR, ANR, and GST, in the flavonoid biosynthetic pathway were upregulated, whereas the COMT, CCR, and CAD genes involved in lignin metabolism were downregulated as fruit ripening progressed. APX as well as PPO, and ACO genes related to the organic acid accumulations were upregulated and downregulated, respectively. Importantly, a comprehensive regulatory network encompassing genes that contribute to the metabolism of flavones, flavonols, lignin, and organic acids was proposed. This network sheds light on the intricate processes that underlie fruit yellowing and quality alterations. These findings enhance our understanding of the regulatory pathways governing pineapple ripening and offer valuable scientific insight into the molecular breeding of pineapples.

Investigating the Effects of Alltech Crop Science (ACS) Products on Plant Defence against Root-Knot Nematode Infestation.

Two formulations of Alltech Crop Science products (ACS), a proprietary blend of fermentation products and plant extracts with micronutrients (ACS5075), and a microbial based product (ACS3048), were tested to understand (1) their impact on the tomato plant immune response and (2) whether they are priming a resistance response in plants against root knot nematodes (RKN). Research findings reported previously indicate that tomato plants pre-treated with ACS5075 and ACS3048 were found less sensitive to Meloidogyne javanica infection. In the current study, the expression of six defence-related genes (PR-1, PR-3, PR-5T, ACO, CAT and JERF 3), relative to a housekeeping gene, were monitored via RT-PCR. Results suggest that the treatment with ACS5075 enhanced ACO and PR-1 gene expression levels, both post- treatment and post-infection with M. javanica. Reduced M. javanica infestation that was reported in the previous study could be attributed to the increased expression of these genes in the ACS5075-treated plants. Tomato plants treated with ACS3048, but without RKN infection, also demonstrated higher levels of ACO and PR-1 gene expression. Subsequently, 2D-gel electrophoresis was performed to study the differential protein expression in leaf tissues of treated tomato plants in an effort to elucidate a possible mechanism of action for these products. Protein spot 1 was identified as 'disease resistance protein RPP13-like', protein spot 2 as 'phosphatidylinositol 4-phosphate 5-kinase 2', spot 3 as 'protein SABRE like' and protein spot 4 as 'uncharacterized protein'. Overall research findings indicate that the ACS products could be used as plant immunity-boosting agents, as they play a significant role in the expression of certain genes and proteins associated with plant defence.

Compatible Consortium of Endophytic Bacillus halotolerans Strains Cal.l.30 and Cal.f.4 Promotes Plant Growth and Induces Systemic Resistance against Botrytis cinerea.

Evaluating microbial-based alternatives to conventional fungicides and biofertilizers enables us to gain a deeper understanding of the biocontrol and plant growth-promoting activities. Two genetically distinct Bacillus halotolerans strains (Cal.l.30, Cal.f.4) were evaluated for the levels of their compatibility. They were applied individually or in combination under in vitro and greenhouse conditions, using seed bio-priming and soil drenching as inoculum delivery systems, for their plant growth-promoting effect. Our data indicate that application of Cal.l.30 and Cal.f.4 as single strains and as a mixture significantly enhanced growth parameters of Arabidopsis and tomato plants. We investigated whether seed and an additional soil treatment with these strains could induce the expression of defense-related genes in leaves of young tomato seedling plants. These treatments mediated a long lasting, bacterial-mediated, systemic-induced resistance as evidenced by the high levels of expression of RP3, ACO1 and ERF1 genes in the leaves of young tomato seedlings. Furthermore, we presented data showing that seed and soil treatment with B. halotolerans strains resulted in an effective inhibition of Botrytis cinerea attack and development on tomato leaves. Our findings highlighted the potential of B. halotolerans strains as they combine both direct antifungal activity against plant pathogens and the ability to prime plant innate immunity and enhance plant growth.

Genome scale analysis of 1-aminocyclopropane-1-carboxylate oxidase gene family in G. barbadense and its functions in cotton fiber development

A class of proteins, 1-aminocyclopropane-1-carboxylate oxidase (ACO), is required in the final step of production of ethylene from its immediate precursor 1-aminocyclopropane-1-carboxylic acid (ACC). Despite the crucial and regulatory role of ACO gene family in the fiber development, it has not been thoroughly analyzed and annotated in G. barbadense genome. In the present study, we have identified and characterized all isoforms of ACO gene family from genomes of Gossypium arboreum, G. barbadense, G. hirsutum and G. raimondii. Phylogenetic analysis classified all ACO proteins into six distinct groups on the basis of maximum likelihood. Gene locus analysis and circos plots indicated the distribution and relationship of these genes in cotton genomes. Transcriptional profiling of ACO isoforms in G. arboreum, G. barbadense and G. hirsutum fiber development exhibited the highest expression in G. barbadense during early fiber elongation. Moreover, the accumulation of ACC was found highest in developing fibers of G. barbadense in comparison with other cotton species. ACO expression and ACC accumulation correlated with the fiber length in cotton species. Addition of ACC to the ovule cultures of G. barbadense significantly increased fiber elongation while ethylene inhibitors hindered fiber elongation. These findings will be helpful in dissecting the role of ACOs in cotton fiber development and pave a way towards genetic manipulations for fiber quality improvement.

Prion Mutations in Republic of Republic of Korea, China, and Japan.

Prion gene (PRNP) mutations are associated with diverse disease phenotypes, including familiar Creutzfeldt-Jakob Disease (CJD), Gerstmann-Sträussler-Scheinker disease (GSS), and fatal familial insomnia (FFI). Interestingly, PRNP mutations have been reported in patients diagnosed with Alzheimer's disease, dementia with Lewy bodies, Parkinson's disease, and frontotemporal dementia. In this review, we describe prion mutations in Asian countries, including Republic of Republic of Korea, China, and Japan. Clinical phenotypes and imaging data related to these mutations have also been introduced in detail. Several prion mutations are specific to Asians and have rarely been reported in countries outside Asia. For example, PRNP V180I and M232R, which are rare in other countries, are frequently detected in Republic of Korea and Japan. PRNP T188K is common in China, and E200K is significantly more common among Libyan Jews in Israel. The A117V mutation has not been detected in any Asian population, although it is commonly reported among European GSS patients. In addition, V210I or octapeptide insertion is common among European CJD patients, but relatively rare among Asian patients. The reason for these differences may be geographical or ethical isolation. In terms of clinical phenotypes, V180I, P102L, and E200K present diverse clinical symptoms with disease duration, which could be due to other genetic and environmental influences. For example, rs189305274 in the ACO1 gene may be associated with neuroprotective effects in cases of V180I mutation, leading to longer disease survival. Additional neuroprotective variants may be possible in cases featuring the E200K mutation, such as KLKB1, KARS, NRXN2, LAMA3, or CYP4X1. E219K has been suggested to modify the disease course in cases featuring the P102L mutation, as it may result in the absence of prion protein-positive plaques in tissue stained with Congo red. However, these studies analyzed only a few patients and may be too preliminary. The findings need to be verified in studies with larger sample sizes or in other populations. It would be interesting to probe additional genetic factors that cause disease progression or act as neuroprotective factors. Further studies are needed on genetic modifiers working with prions and alterations from mutations.

Pseudomonas putida induces resistance to Fusarium oxysporum f. sp. radicis-lycopersici in tomato plants by activating expression of defense-related genes

Plant growth-promoting rhizobacteria (PGPR) may prevent attack from pathogenic microorganisms by eliciting induced systemic resistance (ISR). In the present work, Pseudomonas putida isolate TR21/1 showed significant biological control of tomato seedlings inoculated with Fusarium oxysporum f.sp. radicis-lycopersici (FORL). Here, the SA-responsive genes PR-1, PR-4, PR-6 and CH9 were downregulated upon induction of ISR by P. putida strain TR21/1 and induced when bacterized tomato roots were inoculated with FORL. This indicates that SAR involves the accumulation of SA-responsive genes but ISR does not. Similarly, expression of ET-regulated genes such as ACO1, ACO3, ACO4 were not induced in ISR-expressing tomato roots and P. putida treatment induced only ACO2 expression suggesting that ACO2 expression is involved in ISR-expressing tomato seedlings. In contrast, the infection of ISR expressing plants by FORL strongly induced ACO3, ACO2, and ACO1 indicating the transcriptional regulation of ACO genes in response to FORL attack which may be related to possible ethylene synthesis in response to pathogen. Here P. putida treatment increased ETR1 gene expression in roots and this induction was upregulated in presence of FORL indicating that ETR1 plays a role in the protection of plants against FORL by reducing ethylene sensitivity. Activation of SA-and ET- regulated genes in bacterized plants in the presence of FORL implies that not only SA but other signals as well, may play an important role in inducing resistance.

WRKY29 transcription factor regulates ethylene biosynthesis and response in arabidopsis

The gaseous phytohormone ethylene participates in a lot of physiological processes in plants. 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS, EC 4.4.1.14) and the ACC oxidase (ACO, EC 1.14.17.4) are key enzymes in ethylene biosynthesis. However, how ACSs and ACOs are regulated at the transcriptional level is largely unknown. In the present study, we showed that an Arabidopsis (Arabidopsis thaliana) WRKY-type transcription factor (TF), WRKY29 positively regulated the expression of ACS5, ACS6, ACS8, ACS11 and ACO5 genes and thus promoted basal ethylene production. WRKY29 protein was localized in nuclei and was a transcriptional activator. Overexpression of WRKY29 caused pleiotropic effect on plant growth, development and showed obvious response even without ACC treatment. Inducible overexpression of WRKY29 also reduced primary root elongation and lateral root growth. A triple response assay of overexpression and mutant seedlings of WRKY29 showed that overexpression seedlings had shorter hypocotyls than the transgenic GFP (Green Fluorescence Protein) control, while mutants had no difference from wild-type. A qRT-PCR assay demonstrated that expression of multiple ACSs and ACO5 was up-regulated in WRKY29 overexpression plants. A transactivation assay through dual luciferase reporter system confirmed the regulation of promoters of ACS5, ACS6, ACS8, ACS11 and ACO5 by WRKY29. Both in vivo chromatin immunoprecipitation (ChIP)- quantitative PCR and in vitro electrophoretic mobility shift assay (EMSA) revealed that WRKY29 directly bound to the promoter regions of its target genes. Taken together, these results suggest that WRKY29 is a novel TF positively regulating ethylene production by modulating the expression of ACS and ACO genes.

Genome-Wide Identification of the Aconitase Gene Family in Tomato (Solanum lycopersicum) and CRISPR-Based Functional Characterization of SlACO2 on Male-Sterility.

Tomato (Solanum lycopersicum) is one of the most cultivated vegetables in the world due to its consumption in a large variety of raw, cooked, or processed foods. Tomato breeding and productivity highly depend on the use of hybrid seeds and their higher yield, environmental adaption, and disease tolerance. However, the emasculation procedure during hybridization raises tomato seed production costs and labor expenses. Using male sterility is an effective way to reduce the cost of hybrid seeds and ensure cultivar purity. Recent developments in CRISPR genome editing technology enabled tomato breeders to investigate the male sterility genes and to develop male-sterile tomato lines. In the current study, the tomato Acotinase (SlACO) gene family was investigated via in silico tools and functionally characterized with CRISPR/Cas9-mediated gene disruption. Genome-wide blast and HMM search represented two SlACO genes located on different tomato chromosomes. Both genes were estimated to have a segmental duplication in the tomato genome due to their identical motif and domain structure. One of these genes, SlACO2, showed a high expression profile in all generative cells of tomato. Therefore, the SlACO2 gene was targeted with two different gRNA/Cas9 constructs to identify their functional role in tomatoes. The gene was mutated in a total of six genome-edited tomato lines, two of which were homozygous. Surprisingly, pollen viability was found to be extremely low in mutant plants compared to their wild-type (WT) counterparts. Likewise, the number of seeds per fruit also sharply decreased more than fivefold in mutant lines (10-12 seeds) compared to that in WT (67 seeds). The pollen shape, anther structures, and flower colors/shapes were not significantly varied between the mutant and WT tomatoes. The mutated lines were also subjected to salt and mannitol-mediated drought stress to test the effect of SlACO2 on abiotic stress tolerance. The results of the study indicated that mutant tomatoes have higher tolerance with significantly lower MDA content under stress conditions. This is the first CRISPR-mediated characterization of ACO genes on pollen viability, seed formation, and abiotic stress tolerance in tomatoes.

Nitric Oxide Negatively Regulates the Rapid Formation of Pleurotus ostreatus Primordia by Inhibiting the Mitochondrial aco Gene.

Nitric oxide (NO) is as a signaling molecule that participates in the regulation of plant development and in a number of physiological processes. However, the function and regulatory pathway of NO in the growth and development of edible mushrooms are still unknown. This study found that NO played a negative role in the transformation of Pleurotus ostreatus from vegetative growth to reproductive growth by the exogenous addition of NO donors and scavengers. Further studies showed that NO can inhibit the gene expression and enzyme activity of aconitase (ACO). Moreover, the overexpression (OE) of mitochondrial aco and RNA interference (RNAi) confirmed that ACO participates in the regulation of the primordia formation rate. The effects of aco OE and RNAi on the tricarboxylic acid (TCA) cycle and energy metabolism were further measured. The results showed that RNAi-aco mutant strains can affect the enzyme activities of isocitrate dehydrogenase of mitochondria (ICDHm) and α-ketoglutarate dehydrogenase (α-KGDH) in the TCA cycle, thereby reducing the production of nicotinamide adenine dinucleotide (NADH) in the TCA cycle, decreasing the contents of adenosine triphosphate (ATP) and hydrogen peroxide (H2O2), and negatively regulating the rapid formation of primordia. In addition, H2O2 was significantly increased during the transformation from vegetative growth to reproductive growth of P. ostreatus. Additionally, the exogenous addition of H2O2 and its scavengers further confirmed the positive regulation by H2O2 in primordia formation. This study shows that during the growth and development of P. ostreatus, NO can inhibit the expression of the mitochondrial aco gene and ACO protein in the TCA cycle, reduce the production of ATP and H2O2 in the respiratory chain, and negatively regulate the rate of primordia formation.

Overexpression of acdS gene encoding 1-aminocyclopropane-1-carboxylic acid deaminase enzyme in petunia negatively affects seed germination.

Overexpression of acdS in petunia negatively affects seed germination by suppression of ethylene biosynthesis and signaling genes and induction of abscisic acid biosynthesis genes in the seeds. The acdS gene, which encodes 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, has been overexpressed in horticultural crops to improve their tolerance to abiotic stress. However, the role of acdS in the germination of crop seeds has not been investigated, despite its suppression of ethylene production. In this study, acdS overexpression significantly reduced seed weight and germination rate in transgenic petunia cv. Merage Rose (T5, T7, and T12) relative to wild type via the suppression of ethylene biosynthesis and signaling genes and induction of abscisic acid (ABA) biosynthesis genes. The germination rate of T7 was significantly lower than those of T5 and T12, which was linked to higher expression of acdS in the former than the latter. The addition of exogenous ACC and gibberellic acid (GA3) to the germination medium improved the germination rate of T5 seeds and GA3 promoted the germination rate of T12 seeds. However, neither ACC nor GA3 promoted the germination rate of T7 seeds. The improved germination rates in T5 and T12 were associated with the transcriptional regulation of ethylene biosynthesis genes, particularly that of the ACO1 gene, signaling genes, and ABA biosynthesis genes. In this study, we discovered a negative role of acdS in seed germination in petunia. Thus, we highlight the need to consider the negative effect of acdS on seed germination when overexpressing the gene in horticultural crops to improve tolerance to abiotic stress.

Ethylene regulates aerenchyma formation in cotton under hypoxia stress by inducing the accumulation of reactive oxygen species

Aerenchyma formation is one of the most effective mechanisms adapted by plants to cope with waterlogging (WL). It which reduces the negative effects of WL by facilitating gas diffusion within roots. To understand the response of cotton roots to WL, hypoxia-tolerant genotype Cx094 and hypoxia-sensitive genotype Wang18 were used in this study. The seedlings of two varieties were treated by three methods: flooding in pots as Treatment 1 (flooding), low oxygen with agar solution as Treatment 2 (flooding + agar) and hypoxia with agar solution and nitrogen gas as Treatment 3 (flooding + agar + N 2 ). The results showed that in Cx094, the highest proportion of aerenchyma was found in Treatment 2, with no corresponding aerenchyma formation in Wang18. Further studies were carried out in Treatment 2 to check the aerenchyma formation in six cotton accessions and the results showed that aerenchyma only formed in hypoxia-tolerance genotypes, confirming the important role of aerenchyma formation in WL-tolerant genotypes. The unique characteristic of cotton is that four symmetrical aerenchyma were formed under WL stress, and they appeared one by one. We also showed that hypoxia signal induced expression of ACS and ACO genes, resulting more ethylene synthesis in hypoxia-tolerant genotypes. Moreover, ethylene induced reactive oxygen species (ROS) accumulated by up-regulation of RBOHs expression and down-regulation of MT-3a expression. Finally, ROS caused up-regulation of CEL and XET expressions to induce programmed cell death, and result in lysigenous aerenchyma. Exogenous ethylene inhibitor decreased the formation of aerenchyma in Cx094, while ethephon, an ethylene precursor, promote the formation of aerenchyma in Wang18, signifying the important role of ethylene in the formation of aerenchyma in cotton. • The first time to report aerenchyma in cotton. • Ethylene induced ROS accumulation. • ROS promotes PCD occurrence. • Ethylene promotes aerenchyma formation in cotton.

Runs of homozygosity analysis reveals population characteristics of yellow-feathered chickens using re-sequencing data

ABSTRACT 1. To evaluate the inbreeding of yellow-feathered chickens (YFCs) and identify genes related to their unique characteristics, whole-genome re-sequencing data were applied to detect runs of homozygosity (ROH) in the genomes of 10 YFC chickens from each of 10 different YFC breeds. The number, length, distribution of ROH, and inbreeding coefficient in different YFC populations were calculated. Genomic regions with high frequency in ROH were annotated. 2. In total, 25,547 ROH with an average length of 335 kb were detected, with most being <1 Mb. The domination of short ROH reflected the long breeding history of this chicken. The number, length, frequency, and distribution of ROH varied among chicken populations, and high genetic diversity was maintained. 3. Numerous genes related to YFC characteristics were identified in the high-frequency ROH regions. Among these, IFNA, IFNB, IL11 RA, IL22 RA1, IFNLR1, and TRIF genes were involved in disease resistance. The AMY, G6PC, SDHB, GCNT4, and ACO genes were associated with energy material metabolism; and FABPL, AQP7, ACAA2, and RYR2 were related to meat quality and flavour. The KITLG, CREB3, RYR2, and LGR4 genes, related to pigmentation, were detected. 4. This ROH-based inbreeding evaluation lays the foundation for breeding and conservation of YFC populations, and the candidate genes identified can be used for marker-assisted selection.