BnUC1 Is a Key Regulator of Epidermal Wax Biosynthesis and Lipid Transport in Brassica napus.

The bHLH (basic helix-loop-helix) transcription factor AtCFLAP2 regulates epidermal wax accumulation, but the underlying molecular mechanism remains unknown. We obtained BnUC1mut (BnaA05g18250D homologous to AtCFLAP2) from a Brassica napus mutant with up-curling leaves (Bnuc1) and epidermal wax deficiency via map-based cloning. BnUC1mut contains a point mutation (N200S) in the conserved dimerization domain. Overexpressing BnUC1mut in ZS11 (Zhongshuang11) significantly decreased the leaf epidermal wax content, resulting in up-curled and glossy leaves. In contrast, knocking out BnUC1mut in ZS11-NIL (Zhongshuang11-near-isogenic line) restored the normal leaf phenotype (i.e., flat) and significantly increased the leaf epidermal wax content. The point mutation weakens the ability of BnUC1mut to bind to the promoters of VLCFA (very-long-chain fatty acids) synthesis-related genes, including KCS (ß-ketoacyl coenzyme synthase) and LACS (long-chain acyl CoA synthetase), as well as lipid transport-related genes, including LTP (non-specific lipid transfer protein). The resulting sharp decrease in the transcription of genes affecting VLCFA biosynthesis and lipid transport disrupts the normal accumulation of leaf epidermal wax. Thus, BnUC1 influences epidermal wax formation by regulating the expression of LTP and genes associated with VLCFA biosynthesis. Our findings provide a foundation for future investigations on the mechanism mediating plant epidermal wax accumulation.

The dominant white color trait of the melon fruit rind is associated with epicuticular wax accumulation.

MAIN CONCLUSION: Microscopic analyses and chemical profiling demonstrate that the white rind phenotype in melon fruit is associated with the accumulation of n-alkanes, fatty alcohols, aldehydes and wax esters. Serving as an indicator of quality, the rind (or external) color of fruit directly affects consumer choice. A fruit's color is influenced by factors such as the levels of pigments and deposited epicuticular waxes. The latter produces a white-grayish coating often referred to as "wax bloom". Previous reports have suggested that some melon (Cucumis melo L.) accessions may produce wax blooms, where a dominant white rind color trait was genetically mapped to a major locus on chromosome 7 and suggested to be inherited as a single gene named Wi. We here provide the first direct evidence of the contribution of epicuticular waxes to the dominant white rind trait in melon fruit. Our light and electron microscopy and gas chromatography-mass spectrometry (GC-MS) comparative analysis of melon accessions with white or green rinds reveals that the rind of melon fruit is rich in epicuticular waxes. These waxes are composed of various biochemical classes, including fatty acids, fatty alcohols, aldehydes, fatty amides, n-alkanes, tocopherols, triterpenoids, and wax esters. We show that the dominant white rind phenotype in melon fruit is associated with increased accumulation of n-alkanes, fatty alcohols, aldehydes and wax esters, which are linked with the deposition of crystal-like wax platelets on their surfaces. Together, this study broadens the understanding of natural variation in an important quality trait of melon fruit and promotes the future identification of the causative gene for the dominant white rind trait.

ACL1-ROC4/5 complex reveals a common mechanism in rice response to brown planthopper infestation and drought.

Brown planthopper (BPH) is the most destructive insect pest of rice. Drought is the most detrimental environmental stress. BPH infestation causes adaxial leaf-rolling and bulliform cells (BCs) shrinkage similar to drought. The BC-related abaxially curled leaf1 (ACL1) gene negatively regulates BPH resistance and drought tolerance, with decreased cuticular wax in the gain-of-function mutant ACL1-D. ACL1 shows an epidermis-specific expression. The TurboID system and multiple biochemical assays reveal that ACL1 interacts with the epidermal-characteristic rice outermost cell-specific (ROC) proteins. ROC4 and ROC5 positively regulate BPH resistance and drought tolerance through modulating cuticular wax and BCs, respectively. Overexpression of ROC4 and ROC5 both rescue ACL1-D mutant in various related phenotypes. ACL1 competes with ROC4/ROC5 in homo-dimer and hetero-dimer formation, and interacts with the repressive TOPLESS-related proteins. Altogether, we illustrate that ACL1-ROC4/5 complexes synergistically mediate drought tolerance and BPH resistance through regulating cuticular wax content and BC development in rice, a mechanism that might facilitate BPH-resistant breeding.

A wax chalk and self-heating paper-based analytical device (SH-PAD) for the detection of bisphenol A.

Bisphenol A (BPA) is a synthetic xenoestrogen widely present in the environment, known for its toxicity, endocrine-disrupting nature, carcinogenicity, and mutagenic effects on living organisms. The detection of BPA is essential as it infiltrates the human body through food, water, dust and dermal contact. Conventional methods currently in use are inadequate for on-the-spot detection. Consequently, there is a pressing need to build an all-in-one device that can be quickly fabricated using readily available and cost-effective off-the-shelf materials for the detection of BPA. Firstly, we have leveraged wax chalk for fabricating hydrophobic barriers on paper, which offers a hydrophilic channel resolution of 1.64 mm ± 0.05 mm and also the ability to confine major aqueous solvents without leakage. The fabricated device was used to detect BPA using the Folin-Ciocalteu reagent and sodium carbonate (in the presence of heat). Secondly, we have developed a self-heating paper-based analytical device (SH-PAD) using masking tape, lamination paper and Whatman filter paper. This cost-effective approach (0.017$) is based on an exothermic reaction caused by sodium hydroxide and a small quantity of aluminium in the paper layers and can retain heat adequately for more than 5 minutes, addressing the challenge of external heat sources and enabling effective and rapid colorimetric detection of BPA using the Folin-Ciocalteu reagent and sodium carbonate. Both methods can detect up to 2 µg mL-1 in spiked water samples. This developed method's user-friendliness and cost-effectiveness make it a promising candidate for point-of-care diagnostics or detection, providing testing capabilities in areas with limited resources.

Identification of long-chain acyl-CoA synthetase gene family reveals that SlLACS1 is essential for cuticular wax biosynthesis in tomato.

Long-chain acyl-CoA synthetases (LACSs), belonging to the acyl-activating enzyme superfamily, play crucial roles in lipid biosynthesis and fatty acid catabolism. Here, we identified 11 LACS genes in the tomato reference genome, and these genes were clustered into six subfamilies. Gene structure and conserved motif analyses indicated that LACSs from the same subfamily shared conserved gene and protein structures. Expression analysis revealed that SlLACS1 was highly expressed in the outer epidermis of tomato fruits and leaves. Subcellular localization assay results showed that SlLACS1 was located in the endoplasmic reticulum. Compared with wild-type plants, the wax content on leaves and fruits decreased by 22.5-34.2 % in SlLACS1 knockout lines, confirming that SlLACS1 was involved in wax biosynthesis in both leaves and fruits. Water loss, chlorophyll extraction, water-deficit, and toluidine blue assays suggested that cuticle permeability was elevated in SlLACS1 knockout lines, resulting in reduction in both drought stress resistance and fruit shelf-life. Overall, our analysis of the LACSs in tomato, coupled with investigations of SlLACS1 function, yielded a deeper understanding of the evolutionary patterns of LACS members and revealed the involvement of SlLACS1 in wax accumulation contribute to drought resistance and extended fruit shelf-life in tomato.

A high-quality genome assembly reveals adaptations underlying glossy, wax-coated leaves in the heat-tolerant wild raspberry Rubus leucanthus.

With glossy, wax-coated leaves, Rubus leucanthus is one of the few heat-tolerant wild raspberry trees. To ascertain the underlying mechanism of heat tolerance, we generated a high-quality genome assembly with a genome size of 230.9 Mb and 24,918 protein-coding genes. Significantly expanded gene families were enriched in the flavonoid biosynthesis pathway and the circadian rhythm-plant pathway, enabling survival in subtropical areas by accumulating protective flavonoids and modifying photoperiodic responses. In contrast, plant-pathogen interaction and MAPK signaling involved in response to pathogens were significantly contracted. The well-known heat response elements (HSP70, HSP90, and HSFs) were reduced in R. leucanthus compared to two other heat-intolerant species, R. chingii and R. occidentalis, with transcriptome profiles further demonstrating their dispensable roles in heat stress response. At the same time, three significantly positively selected genes in the pathway of cuticular wax biosynthesis were identified, and may contribute to the glossy, wax-coated leaves of R. leucanthus. The thick, leathery, waxy leaves protect R. leucanthus against pathogens and herbivores, supported by the reduced R gene repertoire in R. leucanthus (355) compared to R. chingii (376) and R. occidentalis (449). Our study provides some insights into adaptive divergence between R. leucanthus and other raspberry species on heat tolerance.

Enhancing thermal energy storage properties of blend phase change materials using beeswax.

This study aims to use beeswax, a readily available and cost-effective organic material, as a novel phase change material (PCM) within blends of low-density polyethylene (LDPE) and styrene-b-(ethylene-co-butylene)-b-styrene (SEBS). LDPE and SEBS act as support materials to prevent beeswax leakage. The physicochemical properties of new blended phase change materials (B-PCM) were determined using an X-ray diffractometer and an infrared spectrometer, confirming the absence of a chemical reaction within the materials. A scanning electron microscope was used for microstructural analysis, indicating that the interconnection of the structure allowed better thermal conductivity. Thermal gravimetric analysis revealed enhanced thermal stability for the B-PCM when combined with SEBS, especially within its operating temperature range. Analysis of phase change temperature and latent heat with differential scanning calorimetry showed no major difference in the melting point of the various PCM blends created. During the melting/solidification process, the B-PCMs possess excellent performance as characterized by W70/P30 (112.45 J.g-1) > W70/P20/S10 (94.28 J.g-1) > W70/P10/S20 (96.21 J.g-1) of latent heat storage. Additionally, the blends tend to reduce supercooling compared to pure beeswax. During heating and cooling cycles, the B-PCM exhibited minimal leakage and degradation, especially in blends containing SEBS. In comparison to the rapid temperature drop observed during the cooling process of W70/P30, the temperature decline of W70/P30 was slower and longer, as demonstrated by infrared thermography. The addition of LDPE to the PCM reduced melting time, indicating an improvement in the thermal energy storage reaction time to the demand. According to the obtained findings, increasing the SEBS concentration in the composite increased the thermal stability of the resulting PCM blends significantly. Despite the challenges mentioned earlier, SEBS proved to be an effective encapsulating material for beeswax, whereas LDPE served well as a supporting material. Leak tests were performed to find the ideal mass ratio, and weight loss was analyzed after multiple cycles of cooling and heating at 70 °C. The morphology, thermal characteristics, and chemical composition of the beeswax/LDPE/SEBS composite were all examined. Beeswax proves to be a highly effective phase change material for storing thermal energy within LDPE/SEBS blends.

Fabrication of a graphite-paraffin carbon paste electrode and demonstration of its use in electrochemical detection strategies.

Electrochemical detection methods hold many advantages over their optical counterparts, such as operation in complex sample matrices, low-cost and high volume manufacture and possible equipment miniaturisation. Despite these advantages, the use of electrochemical detection is currently limited in the clinical setting. There is a wide range of potential electrode materials, selected for optimal signal-to-noise ratios and reproducibility when detecting target analytes. The use of carbon paste electrodes (CPEs) for electrochemical detection can be limited by their analytical performance, however they remain very attractive due to their low cost and biocompatibility. This paper presents the fabrication of an easy-to-make and use graphite powder/paraffin wax paste combined with a substrate produced via additive manufacturing and confirms its functionality for both direct and indirect electrochemical measurements. The produced CPEs enable the direct voltammetric detection of hexaammineruthenium(III) chloride and dopamine at an experimental limit of detection (ELoD) of 62.5 µM. The key inflammatory biomarker Interleukin-6 through an enzyme-linked immunosorbant assay (ELISA) was also quantified, yielding a clinically-relevant ELoD of 150 pg ml-1 in 10% human serum. The performance of low-cost and easy-to-use CPEs obtained in 0.5 hours is showcased in this study, demonstrating the platform's potential uses for point-of-need electroanalytical applications.

Selective capture of PM2.5 by urban trees: The role of leaf wax composition and physiological traits in air quality enhancement.

Human health risks from particles with a diameter of less than 2.5 µm (PM2.5) highlight the role of urban trees as bio-filters in air pollution control. However, whether the size and composition of particles captured by various tree species differ or not remain unclear. This study investigates how leaf attributes affect the capture of PM2.5, which can penetrate deep into the lungs and pose significant health risks. Using a self-developed particulate matter (PM) resuspension chamber and single-particle aerosol mass spectrometer, we measured the size distribution and mass spectra of particles captured by ten tree species. Notably, Cinnamomum camphora (L.) J.Presl and Osmanthus fragrans Lour. are more effective at capturing particles under 1 µm, which are most harmful because they can reach the alveoli, whereas Ginkgo biloba L. and Platanus × acerifolia (Aiton) Willd. tend to capture larger particles, up to 1.6 µm, which are prone to being trapped in the upper respiratory tract. Leaf physiological traits such as stomatal conductance and water potential significantly enhance the capture of larger particles. The Adaptive Resonance Theory neural network (ART-2a) algorithm classified a large number of single particles to determine their composition. Results indicate distinct inter-species variations in chemical composition of particles captured by leaves. Moreover, we identified how specific leaf wax compositions-beyond the known sticky nature of hydrophobic waxes-contribute to particle adhesion, particularly highlighting the roles of fatty acids and alkanes in adhering particles rich in organic carbon and heavy metals, respectively. This research advances our understanding by linking leaf physiological and wax characteristics to the selective capture of PM2.5, providing actionable insights for urban forestry management. The detailed exploration of particle size and composition, tied to specific tree species, enriches the current literature by quantifying how and why different species contribute variably to air quality improvement. This adds a crucial layer of specificity to the general knowledge that trees serve as bio-filters, offering a refined strategy for planting urban trees based on their particulate capture profiles.

Underexplored food safety hazards of beekeeping products: Key knowledge gaps and suggestions for future research.

These days, a growing consumer demand and scientific interest can be observed for nutraceuticals of natural origin, including apiculture products. Due to the growing emphasis on environmental protection, extensive research has been conducted on the pesticide and heavy metal contamination of bee products; however, less attention is devoted on other food safety aspects. In our review, scientific information on the less-researched food safety hazards of honey, bee bread, royal jelly, propolis, and beeswax are summarized. Bee products originating from certain plants may inherently contain phytotoxins, like pyrrolizidine alkaloids, tropane alkaloids, matrine alkaloids, grayanotoxins, gelsemium alkaloids, or tutin. Several case studies evidence that bee products can induce allergic responses to sensitive individuals, varying from mild to severe symptoms, including the potentially lethal anaphylaxis. Exposure to high temperature or long storage may lead to the formation of the potentially toxic 5-hydroxymethylfurfural. Persistent organic pollutants, radionuclides, and microplastics can potentially be transferred to bee products from contaminated environmental sources. And lastly, inappropriate beekeeping practices can lead to the contamination of beekeeping products with harmful microorganisms and mycotoxins. Our review demonstrates the necessity of applying good beekeeping practices in order to protect honeybees and consumers of their products. An important aim of our work is to identify key knowledge gaps regarding the food safety of apiculture products.

Effectiveness and Biocompatibility Evaluation of a Novel Absorbable Bone Wax Used in Bone Tissue.

This study aims to determine the hemostatic effectivity and biocompatibility of a novel absorbable bone wax in comparison with a commercially available product. Eighteen small fat-tail sheep were used to simulate clinical surface bleeding of sternal injury. Hemostasis effectiveness, the degree of bone healing, micro-computed tomography, and histopathology were evaluated over a period after the application of the material to the surgically created wound. The absorbable bone wax used in the study stopped bleeding immediately and did not affect bone healing. The histopathological results also showed that there were no complications associated with the new material. The results showed that the new absorbable bone wax used in this study was effective and biocompatible.

Nanoscale mesh acts as anti-adhesive surface against particulate contamination in eyes of whiteflies.

In many insects the surface of the eye is nanostructured by arrays of protuberances termed ommatidial gratings which provide the cuticle with anti-reflective, anti-wetting and self-cleaning properties. The hypothesised anti-contamination role of the gratings against dust and pollen results from theoretical predictions on grating geometry and experiments on synthetic replicas of ommatidia surfaces but has not yet been proven in an animal. Whiteflies are biological test beds for anti-contamination surfaces because they deliberately distribute wax particles extruded from abdominal plates over their entire bodies. The numerous particles protect the animal against water evaporation and radiation, but may severely impair vision. Using scanning electron microscopy (SEM) and CryoSEM, we here show that the cornea of whiteflies exhibits ~ 220 nm wide mesh-like structures forming hexagonal gratings with thin ~ 40 nm connecting walls. Quantitative measurements of wax particles on the eye show that the nanostructures reduce particle contamination by more than ~ 96% compared to other areas of the cuticle. Altogether, our study is the first description of a predicted optimized grating geometry for anti-contamination in an arthropod. The findings serve as evidence of the high effectiveness of nanostructured surfaces for reducing contact area and thus adhesion forces between biological surfaces and contaminating particles.

Transcription factors and plant hormones mediate wax metabolism in response to drought stress.

Plants have, throughout evolution, developed a hydrophobic cuticle to protect them from various stresses in the terrestrial environment. The cuticle layer is mainly composed of cutin and cuticular wax, a mixture of very-long-chain fatty acids and their derivatives. With the progress of transcriptome sequencing and other research methods, the key enzymes, transporters and regulatory factors in wax synthesis and metabolism have been gradually identified, especially the study on the regulation of wax metabolism by transcription factors and others in response to plant stress has become a hot topic. Drought is a major abiotic stress that limits plant growth and crop productivity. Plant epidermal wax prevents non-stomatal water loss and improves water use efficiency to adapt to arid environments. In this study, the ways of wax synthesis, transport, metabolism and regulation at different levels are reviewed. At the same time, the regulation of wax by different transcription factors and plant hormones in response to drought is elaborated, and key research questions and important directions for future solutions are proposed to enhance the potential application of epidermal wax in agriculture and the environment.

PpyLTP36 and PpyLTP39 are involved in the transmembrane transport of cuticular wax and are associated with the occurrence of pear fruit russeting.

Non-specific lipid-transfer proteins (nsLTPs) are a group of small, cysteine-rich proteins that are involved in the transport of cuticular wax and other lipid compounds. Accumulating evidence suggests that dynamic changes in cuticular waxes are strongly associated with fruit russeting, an undesirable visual quality that negatively affects consumer appeal in pears. Currently, the regulatory role of nsLTPs in cuticular wax deposition and pear fruit skin russeting remains unclear. Here, we characterized the variations of cuticular waxes in non-treated (russeted) and preharvest bagging treated (non-russeted) pear fruits throughout fruit development and confirmed that the contents of cuticular waxes were significantly negatively correlated with the occurrence of pear fruit russeting. Based on RNA-Sequencing (RNA-Seq) and quantitative real-time PCR (qRT-PCR) analyses, two nsLTP genes (PpyLTP36 and PpyLTP39) were identified, which exhibited high expression levels in non-russeted pear fruit skins and were significantly repressed during fruit skin russeting. Subcellular localization analysis demonstrated that PpyLTP36 and PpyLTP39 were localized to the plasma membrane (PM). Further, transient Virus-Induced Gene Silencing (VIGS) analyses of PpyLTP36 and PpyLTP39 in pear fruits significantly reduced cuticular wax deposition. In conclusion, PpyLTP36 and PpyLTP39 are involved in the transmembrane transport of cuticular wax and are associated with pear fruit skin russeting.

The lipid-amylose complexes enhance resistant starch content in candelilla wax-based oleogels cookies.

The substitution of margarine with candelilla wax (CW)-based oleogel is currently a prominent focus of research in the bakery industry. However, the use of CW-based oleogel in cookies increased starch digestibility, potentially posing a risk to human health. Thus, the anti-enzymatic mechanism of lipid-amylose complexes was used to evaluate the influence of olive diacylglycerol stearin (ODS) on starch digestibility in CW-based oleogel cookies. The in vitro digestibility analysis demonstrated that the DCW/ODS-35 cookie exhibited a increase of 27.72 % in slowly digestible starch (SDS) and resistant starch (RS) contents, compared to cookie formulated with margarine. The in-vivo glycemic index analysis revealed that the DCW/ODS-35 cookie had a medium glycemic index of 68. XRD pattern suggested that the presence of ODS in oleogels facilitated the formation of lipid-amylose complexes. The DSC analysis revealed that the addition of ODS resulted in the gelatinization enthalpy of DCW-based cookies increased from 389.9 to 3314.9 J/g. The FTIR spectra indicated that the combination of ODS could promote a short-range ordered structure in DCW-based cookies. Overall, these findings demonstrated that the utilization of DCW-based oleogel presented a viable alternative to commercial margarine in the development of CW-based cookies with reduced starch digestibility.

Phase inversion regulable bigels co-stabilized by Chlorella pyrenoidosa protein and beeswax: In-vitro digestion and food 3D printing.

Algal proteins are an emerging source of functional foods. Herein, Chlorella pyrenoidosa protein (CPP)/xanthan gum-based hydrogels (HG) and beeswax-gelled oleogels (OG) are adopted to fabricate bigels. The phase inversion of bigels can be regulated by the ratio of OG and HG: As the OG increased, bigels turn from OG-in-HG (OG/HG) to a semicontinuous state and then HG-in-OG (HG/OG). In OG/HG bigels (OG ≤ 50 %), hydrophilic CPP acts as the emulsifier at the interface of OG and HG, while beeswax emulsifies the system in HG/OG bigels (OG = 80 %). A semicontinuous bigel appears during the transition between HG/OG and OG/HG. The increase of OG can enhance the viscoelasticity, hardness, adhesiveness, chewiness, and thermal stability. OG/HG bigels exhibit stronger thixotropic recovery and oil-holding capacity than HG/OG bigels. In the in-vitro digestion and food 3D printing, the high specific surface area and the highest thixotropic recovery caused by the emulsion structure of the OG/HG bigel (OG = 50 %) are conducive to the release of free fatty acids and molding of 3D-printed objects, respectively. This study provides a new approach to structure the gelled water-oil system with CPP and helps to develop edible algal proteins-based multiphase systems in food engineering or pharmacy.

Development and characterization of eco-friendly guar gum-agar-beeswax-based active packaging film for cheese preservation.

In this work, an attempt has been made to develop a novel natural polysaccharide-based composite packaging biofilm prepared through a solution casting method. The biofilm is prepared from guar gum (GG) and agar-agar (AA) beeswax (BE). The incorporation of 20 % wt./wt.glycerol BE in the blended polymer GG/AA (50:50) (GG/AA/BE20 (50:50)) film shows a reduction in water solubility (66.67 %), water vapour permeability (69.28 %) and oxygen permeability (72.23 %). Moreover, GG/AA/BE20 (50:50) shows an increment in the tensile strength and elongation of a break by 48.32 % and 26.05 %, respectively, compared to pristine GG film. The scanning electron microscopy (SEM) image reveals defects-free smooth surfaces of the film. The Fourier transform-infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) demonstrated the strong hydrogen bonding between GG, AA, and BE. The biodegradable film shows 99 % degradation within 28 days when placed in the soil. The developed film plays a crucial role in extending the shelf life of cheese, effectively maintaining its moisture content, texture, colour, and pH over a span of up to two months from the point of packaging. These results suggest that GG/AA/BE20 (50:50) composite film is a promising packaging film for cheese preservation.

Comprehensive investigation of niosomal red palm wax gel encapsulating ginger (Zingiber officinale Roscoe): Network pharmacology, molecular docking, In vitro studies and phase 1 clinical trials.

Ginger, a Zingeberaceae family member, is notable for its anti-inflammatory properties. This study explores the pharmaceutical mechanisms of ginger and red palm wax co-extract, developing novel niosomal formulations for enhanced transdermal delivery. Evaluations included physical characteristics, drug loading, in vitro release, network pharmacology, molecular docking, and biocompatibility. The niosomal ginger with red palm wax gel (NGPW) exhibited non-Newtonian fluid properties. The optimized niosome formulation (cholesterol: Tween80: Span60 = 12.5: 20: 5 w/w) showed a high yield (93.23 %), high encapsulation efficiency (54.71 %), and small size (264.33 ± 5.84 nm), prolonging in vitro anti-inflammatory activity. Human skin irritation and biocompatibility tests on 1 % NGPW showed favorable cytotoxicity and hemocompatibility results (ISO10993). Network pharmacology identified potential targets, while molecular docking highlighted high affinities between gingerol and red palm wax compounds with TRPM8 and TRPV1 proteins, suggesting pain inhibition via serotonergic synapse pathways. NGPW presents a promising transdermal pain inhibitory drug delivery strategy.

Genome editing-based mutagenesis stably modifies composition of wax esters synthesized by Euglena gracilis under anaerobic conditions.

Microalgal oil production represents a promising renewable biofuel source. Metabolic engineering can enhance its utility, transforming it into an improved biofuel and expanding its applications as a feedstock for commodity chemicals, thereby increasing their value in biorefineries. This study focused on anaerobic wax ester production by the microalga Euglena gracilis, aiming to develop stable mutant strains with altered wax ester profiles through genome editing. Two enzymes in the fatty acid beta-oxidation pathway involved in wax ester production were targeted-3-ketoacyl-CoA thiolase and acyl-CoA dehydrogenase-using clustered regularly interspaced short palindromic repeats/Cas9. The results revealed one genetic mutation that lengthened and three that shortened the distribution of wax ester compositions compared to the wild-type (WT). The triple-knockout mutant, combining mutations that shorten wax ester chains, produced wax esters with acyl chains two carbons shorter than WT. This study established a methodology to stably modify wax ester composition in E. gracilis.

Cuticle deposition ceases during strawberry fruit development.

BACKGROUND: Ideally, the barrier properties of a fruit's cuticle persist throughout its development. This presents a challenge for strawberry fruit, with their rapid development and thin cuticles. The objective was to establish the developmental time course of cuticle deposition in strawberry fruit. RESULTS: Fruit mass and surface area increase rapidly, with peak growth rate coinciding with the onset of ripening. On a whole-fruit basis, the masses of cutin and wax increase but on a unit surface-area basis, they decrease. The decrease is associated with marked increases in elastic strain. The expressions of cuticle-associated genes involved in transcriptional regulation (FaSHN1, FaSHN2, FaSHN3), synthesis of cutin (FaLACS2, FaGPAT3) and wax (FaCER1, FaKCS10, FaKCR1), and those involved in transport of cutin monomers and wax constituents (FaABCG11, FaABCG32) decreased until maturity. The only exceptions were FaLACS6 and FaGPAT6 that are presumably involved in cutin synthesis, and FaCER1 involved in wax synthesis. This result was consistent across five strawberry cultivars. Strawberry cutin consists mainly of C16 and C18 monomers, plus minor amounts of C19, C20, C22 and C24 monomers, ω-hydroxy acids, dihydroxy acids, epoxy acids, primary alcohols, carboxylic acids and dicarboxylic acids. The most abundant monomer is 10,16-dihydroxyhexadecanoic acid. Waxes comprise mainly long-chain fatty acids C29 to C46, with smaller amounts of C16 to C28. Wax constituents are carboxylic acids, primary alcohols, alkanes, aldehydes, sterols and esters. CONCLUSION: The downregulation of cuticle deposition during development accounts for the marked cuticular strain, for the associated microcracking, and for their high susceptibility to the disorders of water soaking and cracking.