Lycopene Alleviates Endoplasmic Reticulum Stress in Steatohepatitis through Inhibition of the ASK1-JNK Signaling Pathway.

Lycopene has been proven to alleviate nonalcoholic steatohepatitis (NASH), but the precise mechanisms are inadequately elucidated. In this study, we found a previously unknown regulatory effect of lycopene on the apoptosis signal-regulating kinase 1 (ASK1) signaling pathway in both in vivo and in vitro models. Lycopene supplementation (3 and 6 mg/kg/day) exhibited a significant reduction in lipid accumulation, inflammation, and fibrosis of the liver in mice fed with a high-fat/high-cholesterol diet or a methionine-choline-deficient diet. RNA sequencing uncovered that the mitogen-activated protein kinases signaling pathway, which is closely associated with inflammation and endoplasmic reticulum (ER) stress, was significantly downregulated by lycopene. Furthermore, we found lycopene ameliorated ER swelling and decreased the expression levels of ER stress markers (i.e., immunoglobulin heavy chain binding protein, C/EBP homologous protein, and X-box binding protein 1s). Especially, the inositol-requiring enzyme 1α involved in the ASK1 phosphorylation was inhibited by lycopene, resulting in the decline of the subsequent c-Jun N-terminal kinase (JNK) signaling cascade. ASK1 inhibitor DQOP-1 eliminated the lycopene-induced inhibition of the ASK1-JNK pathway in oleic acid and palmitic acid-induced HepG2 cells. Molecular docking further indicated hydrophobic interactions between lycopene and ASK1. Collectively, our research indicates that lycopene can alleviate ER stress and attenuate inflammation cascades and lipid accumulation by inhibiting the ASK1-JNK pathway.

MAPK/NF-κB signaling mediates atrazine-induced cardiorenal syndrome and antagonism of lycopene.

Atrazine (ATZ) is the most prevalent herbicide that has been widely used in agriculture to control broadleaf weeds and improve crop yield and quality. The heavy use of ATZ has caused serious environmental pollution and toxicity to human health. Lycopene (LYC), is a carotenoid that exhibits numerous health benefits, such as prevention of cardiovascular diseases and nephropathy. However, it remains unclear that whether ATZ causes cardiorenal injury or even cardiorenal syndrome (CRS) and the beneficial role of LYC on it. To test this hypothesis, mice were treated with LYC and/or ATZ for 21 days by oral gavage. This study demonstrated that ATZ exposure caused cardiorenal morphological alterations, and several inflammatory cell infiltrations mediated by activating NF-κB signaling pathways. Interestingly, dysregulation of MAPK signaling pathways and MAPK phosphorylation caused by ATZ have been implicated in cardiorenal diseases. ATZ exposure up-regulated cardiac and renal injury associated biomarkers levels that suggested the occurrence of CRS. However, these all changes were reverted, and the phenomenon of CAR was disappeared by LYC co-treatment. Based on our findings, we postulated a novel mechanism to elucidate pesticide-induced CRS and indicated that LYC can be a preventive and therapeutic agent for treating CRS by targeting MAPK/NF-κB signaling pathways.

Enhancing the functionality of plant-based Yogurt: Integration of lycopene through dual-stage fermentation of soymilk.

Well-defined compositional assemblies of plant-based yogurt are of fast-growing awareness for world population concerning environmental sustainability, economic burdens and health risks. Soybean is an attractive candidate for plant yogurt, suffering from poor flavor, limited nutrition, and undesired allergens to offer healthy-functional segments. Herein, we deciphered a novel lycopene-soy yogurt by efficient two-stage fermentation of engineered B. subtilis and LAB. The fortified sogurt was ensured with redundant lycopene of 22.67 ± 2.95 mg/g DCW by engineered B. subtilis and enriched soy isoflavone from synergistic effects of engineered B. subtilis and LAB, possessing strong antioxidant capacity for upgrading functionality. Moreover, the desired pH, accelerated protein hydrolysis, enhanced amino acid availability, and expected sensory attributes cooperatively conferred lycopene-soy yogurt as healthy functional food. High potential is firstly ascribed to sequential dual culture of engineered B. subtilis and LAB in lycopene-soy yogurt, in which flavorful, hypoallergenic and antioxidative ingredients enabled functionalities for plant-based yogurt.

Flux optimization using multiple promoters in Halomonas bluephagenesis as a model chassis of the next generation industrial biotechnology.

Predictability and robustness are challenges for bioproduction because of the unstable intracellular synthetic activities. With the deeper understanding of the gene expression process, fine-tuning has become a meaningful tool for biosynthesis optimization. This study characterized several gene expression elements and constructed a multiple inducible system that responds to ten different small chemical inducers in halophile bacterium Halomonas bluephagenesis. Genome insertion of regulators was conducted for the purpose of gene cluster stabilization and regulatory plasmid simplification. Additionally, dynamic ranges of the multiple inducible systems were tuned by promoter sequence mutations to achieve diverse scopes for high-resolution gene expression control. The multiple inducible system was successfully employed to precisely control chromoprotein expression, lycopene and poly-3-hydroxybutyrate (PHB) biosynthesis, resulting in colorful bacterial pictures, optimized cell growth, lycopene and PHB accumulation. This study demonstrates a desirable approach for fine-tuning of rational and efficient gene expressions, displaying the significance for metabolic pathway optimization.

Lycopene Alleviates Chronic Stress-Induced Hippocampal Microglial Pyroptosis by Inhibiting the Cathepsin B/NLRP3 Signaling Pathway.

Lycopene (LYC) exerts a strong neuroprotective and antipyroptotic effects. This study explored the effects and mechanisms of LYC on chronic stress-induced hippocampal microglial damage and depression-like behaviors. The caspase-1 inhibitor VX-765 attenuated chronic restrain stress (CRS)-induced hippocampal microglial pyroptosis and depression-like behaviors. Moreover, the alleviation of CRS-induced hippocampal microglial pyroptosis and depression-like behaviors by LYC was associated with the cathepsin B/NLRP3 pathway. In vitro, the caspase-1 inhibitor Z-YVAD-FMK alleviated pyroptosis in highly aggressively proliferating immortalized (HAPI) cells. Additionally, the alleviation of corticosterone-induced HAPI cell damage and pyroptosis by LYC was associated with the cathepsin B/NLRP3 pathway. Furthermore, the cathepsin B agonist pazopanib promoted HAPI cell pyroptosis, whereas LYC inhibited pazopanib-induced pyroptosis via the cathepsin B/NLRP3 pathway. Similarly, Z-YVAD-FMK inhibited pazopanib-induced HAPI cell pyroptosis. These results suggest that LYC alleviates chronic stress-induced hippocampal microglial pyroptosis via the cathepsin B/NLRP3 pathway inhibition. This study provides a new strategy for treating chronic stress encephalopathy.

Holistic Assessment Based On Hepatocyte Mitochondria: Lycopene Repairs Oxidized mtDNA to Alleviate Mitochondrial Stress Induced by Atrazine.

Atrazine (ATZ) is a highly persistent herbicide that harms organism health. Lycopene (LYC) is an antioxidant found in plants and fruits. The aim of this study is to investigate the mechanisms of atrazine-induced mitochondrial damage and lycopene antagonism in the liver. The mice were divided into seven groups by randomization: blank control (Con group), vehicle control (Vcon group), 5 mg/kg lycopene (LYC group), 50 mg/kg atrazine (ATZ1 group), ATZ1+LYC group, 200 mg/kg atrazine (ATZ2 group), and ATZ2+LYC group. The present study performed a holistic assessment based on mitochondria to show that ATZ causes the excessive fission of mitochondria and disrupts mitochondrial biogenesis. However, the LYC supplementation reverses these changes. ATZ causes increased mitophagy and exacerbates the production of oxidized mitochondrial DNA (Ox-mtDNA) and mitochondrial stress. This study reveals that LYC could act as an antioxidant to repair Ox-mtDNA and restore the disordered mitochondrial function caused by ATZ.

Characterization of prolycopene-accumulated Tan406 mutant of Solanum lycopersicum.

Significant progress has been made in understanding carotenoid biosynthesis in tomato (Solanum lycopersicum), and most pathway genes have been cloned and characterized. However, isolation and characterization of novel fruit ripening mutants is a continuous and essential process. This study describes the characterization of the Tan406 (Tangerine406) mutant of Solanum lycopersicum. Fruits of Tan406-mutant plants have a unique orange color and accumulate prolycopene instead of lycopene. Genetic analysis revealed that a monogenic recessive mutation affects fruit pigmentation in the mutant, which inhibits the conversion of prolycopene to lycopene. Further, molecular analysis indicates that fruit phenotype is attributed to loss of CRTISO gene function, which encodes a carotenoid isomerase enzyme that converts prolycopene to lycopene. The loss of gene function is due to the deletion of 406 bp from the CRTISO promoter region. Analysis of genome-wide transcriptome expression profiling identified several hundreds of differentially expressed genes in the fruit ripening stages. The results of microarray studies showed a tendency for upregulation of the genes at the mature green stage and downregulation at the fully ripened stage in the mutant. The isolated mutant can be used for the development of varieties having altered nutritional value.

Development of lycopene-based whole-cell biosensors for the visual detection of trace explosives and heavy metals.

Residual explosives in conflicting zones have caused irreversible damage to human safety and the environment. Whole-cell biosensors can to detect remnants of buried explosives, such as 2,4-dinitrotoluene (DNT), a stable and highly volatile compound in explosives. However, all the reported whole-cell biosensors utilize fluorescence or luminescence as the biological markers, making their detection difficult in real minefields. Here, we presented a lycopene-based whole-cell biosensor in Escherichia coli to output visible signals in response to DNT, which can help in the visual detection of buried explosives. To construct the whole-cell biosensor, the DNT-responsive promoter yqjF was used as the sensing element, and the lycopene synthetic gene cassette crtEBI was served as the reporting element. Then, the metabolic flux for lycopene production was enhanced to improve the output signal of the whole-cell biosensor, and a terminator was utilized to reduce the background interference. The optimized biosensor LSZ05 could perceive at least 1 mg/L DNT. The DNT-specificity and robust performance of the biosensor under different environmental factors were confirmed. Our results showed that converting the biosensor into a lyophilized powder was an effective storage method. The biosensor LSZ05 could effectively detect DNT in two kinds of soil samples. The lycopene-based whole-cell biosensor could also be used to visually detect heavy metals. Our findings laid the foundation for visually detecting buried explosives in minefields, which was a valuable supplement to the reported biosensors. The methods used for optimizing the lycopene-based whole-cell biosensor, including the improvement of the output signal and reduction of background interference, were quite effective.

The Functional Characteristics and Soluble Expression of Saffron CsCCD2.

Crocins are important natural products predominantly obtained from the stigma of saffron, and that can be utilized as a medicinal compound, spice, and colorant with significant promise in the pharmaceutical, food, and cosmetic industries. Carotenoid cleavage dioxygenase 2 (CsCCD2) is a crucial limiting enzyme that has been reported to be responsible for the cleavage of zeaxanthin in the crocin biosynthetic pathway. However, the catalytic activity of CsCCD2 on ß-carotene/lycopene remains elusive, and the soluble expression of CsCCD2 remains a big challenge. In this study, we reported the functional characteristics of CsCCD2, that can catalyze not only zeaxanthin cleavage but also ß-carotene and lycopene cleavage. The molecular basis of the divergent functionality of CsCCD2 was elucidated using bioinformatic analysis and truncation studies. The protein expression optimization results demonstrated that the use of a maltose-binding protein (MBP) tag and the optimization of the induction conditions resulted in the production of more soluble protein. Correspondingly, the catalytic efficiency of soluble CsCCD2 was higher than that of the insoluble one, and the results further validated its functional verification. This study not only broadened the substrate profile of CsCCD2, but also achieved the soluble expression of CsCCD2. It provides a firm platform for CsCCD2 crystal structure resolution and facilitates the synthesis of crocetin and crocins.

Lycopene Prevents Phthalate-Induced Cognitive Impairment via Modulating Ferroptosis.

Di-(2-ethylhexyl) phthalate (DEHP) is frequently used as a plasticizer in industrial and agricultural products. DEHP can cause severe neurotoxicity, such as impaired learning and memory function. Lycopene (LYC) as a carotenoid exerts excellent antioxidant capacity and therapeutic effects in neurodegenerative diseases. However, whether LYC can prevent the cognitive impairment induced by DEHP and the specific mechanisms are unclear. In the present study, the behavioral test results suggested that LYC alleviated the learning and memory impairment induced by DEHP. The histopathological data revealed that LYC attenuated DEHP-induced disordered arrangement of the neurons in the CA1 and CA3 regions of the hippocampus tissue. Moreover, LYC inhibited the occurrence of DEHP-induced ferroptosis via regulating iron metabolism, inhibiting lipid peroxidation, and activating the cysteine transporter and nuclear factor erythroid 2-related factor 2/heme oxygenase-1 (NrF2/HO-1) signaling pathway. Overall, the study contributes novel perspectives into the potential mechanisms of LYC preventing phthalate-induced cognitive impairment in the hippocampus.

Metabolic Engineering of Pichia pastoris for High-Level Production of Lycopene.

Lycopene is widely used in cosmetics, food, and nutritional supplements. Microbial production of lycopene has been intensively studied. However, few metabolic engineering studies on Pichia pastoris have been aimed at achieving high-yield lycopene production. In this study, the CRISPR/Cpf1-based gene repression system was developed and the gene editing system was optimized, which were applied to improve lycopene production successfully. In addition, the sterol regulatory element-binding protein SREBP (Sre) was used for the regulation of lipid metabolic pathways to promote lycopene overproduction in P. pastoris for the first time. The final engineered strain produced lycopene at 7.24 g/L and 75.48 mg/g DCW in fed-batch fermentation, representing the highest lycopene yield in P. pastoris reported to date. These findings provide effective strategies for extended metabolic engineering assisted by the CRISPR/Cpf1 system and new insights into metabolic engineering through transcriptional regulation of related metabolic pathways to enhance carotenoid production in P. pastoris.

Clorf Encodes Carotenoid Isomerase and Regulates Orange Flesh Color in Watermelon (Citrullus lanatus L.).

Flesh color is a significant characteristic of watermelon. Although various flesh-color genes have been identified, the inheritance and molecular basis of the orange flesh trait remain relatively unexplored. In the present study, the genetic analysis of six generations derived from W1-1 (red flesh) and W1-61 (orange flesh) revealed that the orange flesh color trait was regulated by a single recessive gene, Clorf (orange flesh). Bulk segregant analysis (BSA) locked the range to ∼4.66 Mb, and initial mapping situated the Clorf locus within a 688.35-kb region of watermelon chromosome 10. Another 1,026 F2 plants narrowed the Clorf locus to a 304.62-kb region containing 32 candidate genes. Subsequently, genome sequence variations in this 304.62-kb region were extracted for in silico BSA strategy among 11 resequenced lines (one orange flesh and ten nonorange flesh) and finally narrowed the Clorf locus into an 82.51-kb region containing nine candidate genes. Sequence variation analysis of coding regions and gene expression levels supports Cla97C10G200950 as the most possible candidate for Clorf, which encodes carotenoid isomerase (Crtiso). This study provides a genetic resource for investigating the orange flesh color of watermelon, with Clorf malfunction resulting in low lycopene accumulation and, thus, orange flesh.

Lycopene attenuates silver nanoparticle-induced liver injury in albino mice.

Lycopene is a carotenoid widely used for its dominant antioxidant properties and beneficial health effects. Silver nanoparticles (AgNP) have gained attention for use in many medicinal and consumer products, leading to animal, human, and environmental exposure. This study investigated the dose-dependent effects of lycopene on AgNP-induced hepatotoxicity in albino mice. The four experimental groups, comprising eight albino mice each, were as follows: Group I, vehicle control (C); Group II, AgNP-treated (5 mg/kg/day) (AgNP); Group III, AgNP/lycopene-treated (5 + 10 mg/kg/day) (AgNP + LP10); and Group IV, AgNP/lycopene-treated (5 + 100 mg/kg/day) (AgNP + LP100). All solutions were orally administered to the mice once in a day for consecutive 14 days. The levels of serum aspartate transaminase, alanine transaminase, alkaline phosphatase, and total bilirubin were significantly higher in the AgNP-treated group than in the control group but significantly lower in the AgNP + LP100 group than in the AgNP-treated group. A significant decrease in reduced glutathione level and superoxide dismutase activity and an increase in lipid peroxidation were observed in the AgNP-treated group; these were significantly suppressed in the AgNP+LP100 as compared to AgNP-treated group. Histopathological examination showed substantial morphological alterations in hepatic tissues in the AgNP, which were adequately improved in the low and high dose lycopene-treated groups. The dose of 100 mg/kg/day of lycopene was more effective than 10 mg/kg/day, as pretreatment with high dose lycopene significantly diminished the adverse changes occurred due to AgNP in liver weight, hepatic architecture, serum functional markers, and antioxidant markers. Thus, present study shows that pretreatment with lycopene offers protection against AgNP-induced hepatotoxicity and oxidative stress.

Antioxidant and anti-apoptotic potency of allicin and lycopene against methotrexate-induced cardiac injury in rats.

This study aimed to explore whether allicin (ALC) and lycopene (LP) could offer protection against the harmful effects of methotrexate (MTX), a type of chemotherapy drug known for its severe side effects, on the heart of rats. In this experiment, seven groups of rats (n = 7) were used. The first group was given saline as a control vehicle, the second group was given ALC at a dosage of 20 mg/kg orally, the third group was given LP at a dosage of 10 mg/kg orally, and the fourth group was given MTX at a dosage of 20 mg/kg intraperitoneally on the 15th day of the experiment. The remaining three groups received treatments, including ALC + MTX, LP + MTX, and ALC + LP + MTX. After the administration of MTX, the concentrations of serum cardiac biomarkers, such as Creatine kinase (CK), Lactate dehydrogenase (LDH), and creatine kinase-myoglobin binding (CK-MB) were found to increase. Also, MTX caused a notable rise in malondialdehyde (MDA) levels and significant declines in the levels of glutathione (GSH), superoxide dismutase (SOD), and catalase (CAT) in the heart tissues of rats. In addition, MTX caused alterations in the cardiac histopathology and enhanced the caspase-3 expression in the cardiac tissues, indicating the occurrence of apoptosis. The antioxidant properties of ALC and/or LP were effectively reduced cardiac toxicity and apoptosis induced by MTX. The administration of ALC and/or LP was found to alleviate these effects caused by MTX.

Functional Characterization of Lycopene ß- and ε-Cyclases from a Lutein-Enriched Green Microalga Chlorella sorokiniana FZU60.

Lutein is a high-value carotenoid with many human health benefits. Lycopene ß- and ε-cyclases (LCYB and LCYE, respectively) catalyze the cyclization of lycopene into distinct downstream branches, one of which is the lutein biosynthesis pathway, via α-carotene. Hence, LCYB and LCYE are key enzymes in lutein biosynthesis. In this study, the coding genes of two lycopene cyclases (CsLCYB and CsLCYE) of a lutein-enriched marine green microalga, Chlorella sorokiniana FZU60, were isolated and identified. A sequence analysis and computational modeling of CsLCYB and CsLCYE were performed using bioinformatics to identify the key structural domains. Further, a phylogenetic analysis revealed that CsLCYB and CsLCYE were homogeneous to the proteins of other green microalgae. Subcellular localization tests in Nicotiana benthamiana showed that CsLCYB and CsLCYE localized in chloroplasts. A pigment complementation assay in Escherichia coli revealed that CsLCYB could efficiently ß-cyclize both ends of lycopene to produce ß-carotene. On the other hand, CsLCYE possessed a strong ε-monocyclase activity for the production of δ-carotene and a weak ε-bicyclic activity for the production of ε-carotene. In addition, CsLCYE was able to catalyze lycopene into ß-monocyclic γ-carotene and ultimately produced α-carotene with a ß-ring and an ε-ring via γ-carotene or δ-carotene. Moreover, the co-expression of CsLCYB and CsLCYE in E. coli revealed that α-carotene was a major product, which might lead to the production of a high level of lutein in C. sorokiniana FZU60. The findings provide a theoretical foundation for performing metabolic engineering to improve lutein biosynthesis and accumulation in C. sorokiniana FZU60.

Rapid construction of Escherichia coli chassis with genome multi-position integration of isopentenol utilization pathway for efficient and stable terpenoid accumulation.

The isopentenol utilization pathway (IUP) is potential in terpenoids synthesis. This study aimed to construct IUP-employed Escherichia coli chassis for stably synthesizing terpenoids. As to effectiveness, promotor engineering strategy was employed to regulate IUP expression level, while ribosome-binding site (RBS) library of the key enzyme was constructed for screening the optimal RBS, followed by optimization of concentration of inducer and substrates, the titer of reporting production, lycopene, from 0.087 to 8.67 mg OD600 -1 . As about stability, the IUP expression cassette was integrated into the genome through transposition tool based on CRISPR-associated transposases. Results showed that the strain with 13 copies produced 1.78-fold lycopene titer that of the controlled strain with IUP-harbored plasmid, and it exhibited stable expression after ten successions while the plasmid loss was observed in the controlled strain in the 3rd succession. This strategy provides valuable information for rapid construction of highly effective and stable chassis employing IUP for terpenoids production.

A Programmable CRISPR/Cas9 Toolkit Improves Lycopene Production in Bacillus subtilis.

Bacillus subtilis has been widely used and generally recognized as a safe host for the production of recombinant proteins, high-value chemicals, and pharmaceuticals. Thus, its metabolic engineering attracts significant attention. Nevertheless, the limited availability of selective markers makes this process difficult and time-consuming, especially in the case of multistep biosynthetic pathways. Here, we employ CRISPR/Cas9 technology to build an easy cloning toolkit that addresses commonly encountered obstacles in the metabolic engineering of B. subtilis, including the chromosomal integration locus, promoter, terminator, and guide RNA (gRNA) target. Six promoters were characterized, and the promoter strengths ranged from 0.9- to 23-fold that of the commonly used strong promoter P43. We characterized seven terminators in B. subtilis, and the termination efficiencies (TEs) of the seven terminators are all more than 90%. Six gRNA targets were designed upstream of the promoter and downstream of the terminator. Using a green fluorescent protein (GFP) reporter, we confirmed integration efficiency with the single-locus integration site is up to 100%. We demonstrated the applicability of this toolkit by optimizing the expression of a challenging but industrially important product, lycopene. By heterologous expression of the essential genes for lycopene synthesis on the B. subtilis genome, a total of 13 key genes involved in the lycopene biosynthetic pathway were manipulated. Moreover, our findings showed that the gene cluster ispG-idi-dxs-ispD could positively affect the production of lycopene, while the cluster dxr-ispE-ispF-ispH had a negative effect on lycopene production. Hence, our multilocus integration strategy can facilitate the pathway assembly for production of complex chemicals and pharmaceuticals in B. subtilis. IMPORTANCE We present a toolkit that allows for rapid cloning procedures and one-step subcloning to move from plasmid-based expression to stable chromosome integration and expression in a production strain in less than a week. The utility of the customized tool was demonstrated by integrating the MEP (2C-methyl-d-erythritol-4-phosphate) pathway, part of the pentose phosphate pathway (PPP), and the hetero-lycopene biosynthesis genes by stable expression in the genome. The tool could be useful to engineer B. subtilis strains through diverse recombination events and ultimately improve its potential and scope of industrial application as biological chassis.

Development of Terminator-Promoter Bifunctional Elements for Application in Saccharomyces cerevisiae Pathway Engineering.

The construction of a genetic circuit requires the substitution and redesign of different promoters and terminators. The assembly efficiency of exogenous pathways will also decrease significantly when the number of regulatory elements and genes is increased. We speculated that a novel bifunctional element with promoter and terminator functions could be created via the fusion of a termination signal with a promoter sequence. In this study, the elements from a Saccharomyces cerevisiae promoter and terminator were employed to design a synthetic bifunctional element. The promoter strength of the synthetic element is apparently regulated through a spacer sequence and an upstream activating sequence (UAS) with a ~5-fold increase, and the terminator strength could be finely regulated by the efficiency element, with a ~5-fold increase. Furthermore, the use of a TATA box-like sequence resulted in the adequate execution of both functions of the TATA box and the efficiency element. By regulating the TATA box-like sequence, UAS, and spacer sequence, the strengths of the promoter-like and terminator-like bifunctional elements were optimally fine-tuned with ~8-fold and ~7-fold increases, respectively. The application of bifunctional elements in the lycopene biosynthetic pathway showed an improved pathway assembly efficiency and higher lycopene yield. The designed bifunctional elements effectively simplified pathway construction and can serve as a useful toolbox for yeast synthetic biology.

LYC loaded ferritin nanoparticles for intracerebral delivery and the attenuation of neurodegeneration in D-gal-induced mice.

Recombinant human H-ferritin nanocage (rHuHF) loaded with natural antioxidative lycopene molecules (LYC) was successfully constructed for the first time, aiming to enrich LYC in the brain and explore the regulation mechanism of this nanoparticles on neurodegeneration. Here, the mouse model was constructed via D-galactose-induced neurodegeneration based on behavioural analysis, histological observation, immunostaining analysis, Fourier transform infrared microscopy, and Western blotting analysis for the regulation of rHuHF-LYC. rHuHF-LYC improved the behaviour of mice in a dose-dependent manner. Besides, rHuHF-LYC can attenuate neuronal damage, maintain the number of Nissl body, increase the level of unsaturated fat, inhibit the activation of glial cells, and prevent excessive accumulation of neurotoxic proteins in the hippocampus of mice. More importantly, synaptic plasticity was activated in response to the regulation of rHuHF-LYC with excellent biocompatibility and biosafety. This study proved the validity of the direct use of natural antioxidant nano drugs for treating neurodegeneration, providing a promising therapeutic option against further imbalances in the degenerative brain microenvironment.

Lycopene ε-cyclase mediated transition of α-carotene and ß-carotene metabolic flow in carrot fleshy root.

The accumulation of carotenoids, such as xanthophylls, lycopene, and carotenes, is responsible for the color of carrot (Daucus carota subsp. sativus) fleshy roots. The potential role of DcLCYE, encoding a lycopene ε-cyclase associated with carrot root color, was investigated using cultivars with orange and red roots. The expression of DcLCYE in red carrot varieties was significantly lower than that in orange carrots at the mature stage. Furthermore, red carrots accumulated larger amounts of lycopene and lower levels of α-carotene. Sequence comparison and prokaryotic expression analysis revealed that amino acid differences in red carrots did not affect the cyclization function of DcLCYE. Analysis of the catalytic activity of DcLCYE revealed that it mainly formed ε-carotene, while a side activity on α-carotene and γ-carotene was also observed. Comparative analysis of the promoter region sequences indicated that differences in the promoter region may affect the transcription of DcLCYE. DcLCYE was overexpressed in the red carrot 'Benhongjinshi' under the control of the CaMV35S promoter. Lycopene in transgenic carrot roots was cyclized, resulting in the accumulation of higher levels of α-carotene and xanthophylls, while the ß-carotene content was significantly decreased. The expression levels of other genes in the carotenoid pathway were simultaneously upregulated. Knockout of DcLCYE in the orange carrot 'Kurodagosun' by CRISPR/Cas9 technology resulted in a decrease in the α-carotene and xanthophyll contents. The relative expression levels of DcPSY1, DcPSY2, and DcCHXE were sharply increased in DcLCYE knockout mutants. The results of this study provide insights into the function of DcLCYE in carrots, which could serve as a basis for creating colorful carrot germplasms.