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Background Heme is an iron/porphyrin complex compound, widely used in the health care, food, and pharmaceutical industries. It is more advantageous and attractive to develop microbial cell factories to produce heme by fermentation, with lower production costs and environmentally more friendly procedures than those of the traditional extraction based on animal blood. In this study, Bacillus subtilis , a typical industrial model microorganism of food safety grade, was used for the first time as the host to synthesize heme. Results The heme biosynthetic pathway was engineered as four modules, the endogenous C5 pathway, the heterologous C4 pathway, the uroporphyrinogen (urogen) III synthesis pathway, and the downstream synthesis pathway. Knockout of hemX encoding the negative effector of the concentration of HemA, overexpression of hemA encoding glutamyl-tRNA reductase, and knockout of rocG encoding the major glutamate dehydrogenase in the C5 pathway, resulted in an increase of 427% in heme production. Introduction of the heterologous C4 pathway showed a negligible effect on heme biosynthesis. Overexpression of hemCDB , which encoded hydroxymethylbilane synthase, urogen III synthase, and porphobilinogen synthase participating in the urogen III synthesis pathway, increased heme production by 39%. Knockouts of uroporphyrinogen methyltransferase gene nasF and both heme monooxygenase genes hmoA and hmoB in the downstream synthesis pathway increased heme production by 52%. The engineered B. subtilis produced 248.26 ± 6.97 mg/L of total heme with 221.83 ± 4.71 mg/L of extracellular heme during the fed-batch fermentation in 10 L fermenter. Conclusions Strengthening endogenous C5 pathway, urogen III synthesis pathway and downstream synthesis pathway promoted the biosynthesis of heme in B. subtilis . The engineered B. subtilis strain has great potential as a microbial cell factory for efficient industrial heme production.

Recent studies have shown that disruptions in the nicotinamide adenine dinucleotide (NAD+) de novo synthesis pathway accelerate ovarian aging, yet its role in spermatogenesis remains largely unknown. In this study, we investigated the impact of the NAD+ de novo synthesis pathway on spermatogenesis by generating Qprt‐deficient mice using CRISPR‐Cas9 to target quinolinate phosphoribosyl transferase (Qprt), a key enzyme predominantly expressed in spermatocytes. Our results revealed that the deletion of Qprt did not affect NAD+ levels or spermatogenesis in the testes of 3‐month‐old mice. However, from 6 months of age onward, Qprt‐deficient mice exhibited significantly reduced NAD+ levels in the testes compared to wild‐type (WT) controls, along with a notable decrease in germ cell numbers and increased apoptosis. Additionally, these mice demonstrated mitochondrial dysfunction in spermatocytes, impaired progression through prophase I of meiosis, defective double‐strand break (DSB) repair, and abnormal meiotic sex chromosome inactivation. Importantly, supplementation with the NAD+ precursor nicotinamide riboside (NR) in Qprt‐deficient mice restored NAD+ levels and rescued the spermatogenic defects. These findings underscore the critical role of NAD+ de novo synthesis in maintaining NAD+ homeostasis and highlight its importance in meiotic recombination and meiotic sex chromosome inactivation in spermatogenesis. In Qprt‐deficient mice, testicular NAD+ levels decline accelerated with age, leading to mitochondrial dysfunction, disrupted DSB repair, impaired meiotic sex chromosome inactivation, and germ cell loss. Nicotinamide riboside (NR) supplementation restores NAD+ levels and alleviates these defects, highlighting the importance of NAD+ de novo synthesis in age‐related reproductive health.

Acute intermittent porphyria (AIP) is a rare disease caused by a deficiency of hydroxymethylbilane synthase (HMBS), the third enzyme of the heme‐synthesis pathway. Decreased enzymatic activity in the liver induces an overproduction of heme‐precursors and acute neurological attacks. We report a 36‐years‐old female with AIP with a long‐term history of severe, disabling, recurrent attacks, who underwent curative liver transplantation. Tissue samples from the explant were obtained for transcriptome analysis. Whole RNA was extracted and 16 gene‐transcripts were selected and investigated by quantitative polymerase chain reaction. These included nine genes encoding enzymes that consecutively catalyze heme‐synthesis and catabolism in the liver (ALAS1; ALAD; HMBS; UROS; UROD; CPOX; PPOX; FECH; HMOX1). Additionally, we studied genes related to inflammation (IL6; TNF) insulin signaling (PGC‐1α; IGF‐1; FOXO‐1) and tryptophan metabolism (TDO2; IDO). Transcripts of eight house‐keeping genes were co‐measured for normalization. All transcripts were also measured in five control samples from healthy living liver donors. The transcriptome of the controls showed important differences between the various genes, with the first two genes of the heme‐synthesis pathway, ALAS1 and ALAD showing strikingly high mRNA levels compared to the consecutive HMBS gene. Transcripts of several genes significantly differed in the AIP liver compared to controls. Transcripts of HMOX1 and UROS were increased in the AIP liver whereas transcripts of UROD; CPOX, PPOX, and TDO2 were decreased. ALAS1 expression was not increased, possibly due to hemin administered to the patient before transplantation. These results highlight several transcriptomic changes related to heme homeostasis in AIP.

The oleaginous yeast Lipomyces starkeyi is an excellent sustainable lipid producer, which can convert industrial wastes into lipids and accumulate triacylglycerols (TAG) by > 70% of its dry cell weight. Recent studies using omics technologies applied in L. starkeyi have aided in obtaining greater understanding of the important mechanisms of lipid metabolism in L. starkeyi. Therefore, the development of genetic engineering tools for L. starkeyi has led to accelerated efforts for a highly efficient production of lipids.This review focuses on the aspects of TAG and fatty acid synthesis pathways in L. starkeyi. We also present a quite effective strategy to obtain L. starkeyi mutants accumulating a larger amount of lipids and having a higher lipid production rate than the wild-type strain. The analysis of these mutants exhibiting high lipid production has led to the identification of important genes for achieving highly effective lipid production and thus advanced improvement in lipid production. Herein, our aim was to provide useful information to advance the development of L. starkeyi as a cost-effective TAG feedstock.Key Points•Oleaginous yeast Lipomyces starkeyi is an excellent sustainable lipid producer.•Efficient isolation of lipid-enriched L. starkeyi mutants depends on the low density of lipids.•Increased acyl-CoA synthesis pathway is important for improving lipid productivity.

Recent advances highlight the pivotal role of nicotinamide adenine dinucleotide (NAD + ) in ovarian aging. However, the roles of de novo NAD + biosynthesis on ovarian aging are still unknown. Here, we found that genetic ablation of Ido1 (indoleamine‐2,3‐dioxygenase 1) or Qprt (Quinolinate phosphoribosyl transferase), two critical genes in de novo NAD + biosynthesis, resulted in decreased ovarian NAD + levels in middle‐aged mice, leading to subfertility, irregular estrous cycles, reduced ovarian reserve, and accelerated aging. Moreover, we observed impaired oocyte quality, characterized by increased reactive oxygen species and spindle anomalies, which ultimately led to reduced fertilization ability and impaired early embryonic development. A transcriptomic analysis of ovaries in both mutant and wild‐type mice revealed alterations in gene expression related to mitochondrial metabolism. Our findings were further supported by the observation of impaired mitochondrial distribution and decreased mitochondrial membrane potential in the oocytes of knockout mice. Supplementation with nicotinamide riboside (NR), an NAD + booster, in mutant mice increased ovarian reserve and improved oocyte quality. Our study highlights the importance of the NAD + de novo pathway in middle‐aged female fertility.

Fragrant rice is regarded as a premium variety due to its distinctive aroma, delicate texture, and rich nutritional value. This aroma primarily originates from 2-acetyl-1-pyrroline (2-AP), but the metabolic basis of 2-AP remains elusive to this day, and the genetic basis for metabolite accumulation is largely unknown. While several researchers have investigated differences in 2-AP synthesis pathways between fragrant and non-fragrant rice, few studies have examined the 2-AP synthesis pathways in fragrant rice varieties exhibiting 2-AP differences. Therefore, after conducting gene similarity analyses on six fragrant rice varieties, we measured the expression levels of substances and related genes involved in multiple metabolic pathways within the 2-AP synthesis pathway, along with the specific enzyme activities associated with these pathways. Results indicate that XG12 (Guizhou Fragrant Rice Variety) exhibits the highest 2-AP content, yet its efficiency in synthesizing 2-AP is not the highest across any individual metabolic pathway. This finding reveals that among fragrant rice varieties, 2-AP content negatively correlates with OsBadh2 expression levels and GABA content, while showing no linear correlation with other related substances or metabolic genes. At this point, variations in the major effect gene OsBadh2 no longer dominate; instead, subtle differences in 2-AP content are jointly determined by numerous minor effect genes and environmental factors. This phenomenon not only resolves apparent contradictions but profoundly illuminates the complex regulatory mechanisms governing 2-AP biosynthesis. 2-AP synthesis represents a dynamic equilibrium process, with different fragrant rice varieties potentially accumulating 2-AP through distinct metabolic pathways. Additionally, this study analyzed the volatile organic compounds (VOCs) of six fragrant rice varieties through metabolomics. Results revealed that DLX, which exhibited the lowest 2-AP content, contained the richest array of aggregated VOCs, indicating no correlation between 2-AP and numerous VOCs. Our findings provide a clear research direction for elucidating the genetic regulatory mechanisms of 2-AP underlying fragrant rice and lay the foundation for technological research aimed at enhancing the aroma of fragrant rice varieties.

Surfactin has many biological activities, such as inhibiting plant diseases, resisting bacteria, fungi, viruses, tumors, mycoplasma, anti-adhesion, etc. It has great application potential in agricultural biological control, clinical medical treatment, environmental treatment and other fields. However, the low yield has been the bottleneck of its popularization and application. It is very important to understand the synthesis route and control strategy of surfactin to improve its yield and purity. In this paper, based on the biosynthetic pathway and regulatory factors of surfactin, its biosynthesis regulation strategy was comprehensively summarized, involving enhancement of endogenous and exogenous precursor supply, modification of the synthesis pathway of lipid chain and peptide chain, improvement of secretion and efflux, and manipulation some global regulatory factors, such as Spo0A, AbrB, ComQXP, phrCSF, etc. to directly or indirectly stimulate surfactin synthesis. And the current production and separation and purification process of surfactin are briefly described. This review also provides a scientific reference for promoting surfactin production and its applications in various fields.

Medicinal plant microRNAs (miRNAs) are an endogenous class of small RNA central to the posttranscriptional regulation of gene expression. Biosynthetic research has shown that the mature miRNAs in medicinal plants can be produced from either the standard messenger RNA splicing mechanism or the pre-ribosomal RNA splicing process. The medicinal plant miRNA function is separated into two levels: (1) the cross-kingdom level, which is the regulation of disease-related genes in animal cells by oral intake, and (2) the intra-kingdom level, which is the participation of metabolism, development, and stress adaptation in homologous or heterologous plants. Increasing research continues to enrich the biosynthesis and function of medicinal plant miRNAs. In this review, peer-reviewed papers on medicinal plant miRNAs published on the Web of Science were discussed, covering a total of 78 species. The feasibility of the emerging role of medicinal plant miRNAs in regulating animal gene function was critically evaluated. Staged progress in intra-kingdom miRNA research has only been found in a few medicinal plants, which may be mainly inhibited by their long growth cycle, high demand for growth environment, immature genetic transformation, and difficult RNA extraction. The present review clarifies the research significance, opportunities, and challenges of medicinal plant miRNAs in drug development and agricultural production. The discussion of the latest results furthers the understanding of medicinal plant miRNAs and helps the rational design of the corresponding miRNA/target genes functional modules.

Aroma compounds are important flavor components in pear fruit. Among cultivated pears, fruits from (hereafter referred to as ) cultivars are famous for their abundant aroma, while the fruits of most (hereafter referred to as ) cultivars lack aroma compounds. A comparative study on the formation of differential aroma compounds between the two species could provide a theoretical foundation for improving the aroma quality of cultivars. However, there is a lack of systematic research on this subject. An analysis of volatile and non-volatile metabolites was combined with transcriptome analysis to explore the formation mechanism of differential aroma compounds between three and three cultivars. In this study, a total of 510 volatile compounds were identified in the six cultivars. Of these, sixteen ester and alcohol compounds, including butyl acetate, hexyl acetate, ethyl-2-methylbutyrate, ethanol, butanol, propanol, and 2-methylbutanol, with higher contents in the cultivars than in the cultivars were identified as the primary differential aroma compounds. Among the possible synthesis pathways for these 16 aroma compounds, certain amino acid degradation processes, including isoleucine, valine, and alanine oxidation and threonine dehydration, were found to provide important intermediate substances for synthesis. Within the key enzyme genes in the synthesis pathway, several critical enzyme genes, including monoacylglycerol lipase (PcMAGL, pycom08g09340), threonine dehydrase (PcTD, pycom12g10020), and acyl CoA dehydrogenase (PcACD, pycom16g13880), might be important factors contributing to the disparity in aromatic compounds between and cultivars. The aforementioned results provide valuable information into the formation mechanisms of differential aroma compounds and offer novel target sites for enhancing pear aroma quality through gene editing.

We have developed an innovative nanoparticle preparation technique. It utilised a high-pressure steam sterilisation pot as the reaction equipment, simplifying biomedical nanoparticle synthesis and sterilisation processes. Gold nanoparticles (AuNPs), silver nanoparticles (AgNPs), and two types of hydrotalcite-layered nanoparticles (LDHs) were prepared successfully. The nanoparticles had a regular morphology and good dispersion, which is consistent with the characteristics of products prepared using traditional methods. The average particle sizes of AuNPs and AgNPs were 25 and 38 nm. Ultraviolet absorption peaks of AuNPs and AgNPs were observed at 520 and 436 nm, respectively. The average particle sizes of the LDHs used for anion and cation intercalation were 82 and 90 nm, respectively. The elemental composition of the LDHs included Co, Mg, Al, C, and O. Functional group characterisation results showed that the surfaces of the four nanoparticles contained abundant functional groups. The high-pressure sterilisation method used in this study produced sterile nanoparticles with good performance in a single step. It avoided the problems associated with the physicochemical property changes in the nanoparticles caused by the secondary sterilisation operations, thereby ensuring their usability in biomedical applications. Thus, the method provides a scalable synthesis pathway for nanotechnology and inspiration for further research on nanotechnology.