Advances and perspectives in genetic expression and operation for the oleaginous yeast Yarrowia lipolytica.

The utilization of industrial biomanufacturing has emerged as a viable and sustainable alternative to fossil-based resources for producing functional chemicals. Moreover, advancements in synthetic biology have created new opportunities for the development of innovative cell factories. Notably, Yarrowia lipolytica, an oleaginous yeast that is generally regarded as safe, possesses several advantageous characteristics, including the ability to utilize inexpensive renewable carbon sources, well-established genetic backgrounds, and mature genetic manipulation methods. Consequently, there is increasing interest in manipulating the metabolism of this yeast to enhance its potential as a biomanufacturing platform. Here, we reviewed the latest developments in genetic expression strategies and manipulation tools related to Y. lipolytica, particularly focusing on gene expression, chromosomal operation, CRISPR-based tool, and dynamic biosensors. The purpose of this review is to serve as a valuable reference for those interested in the development of a Y. lipolytica microbial factory.

High titer production of gastrodin enabled by systematic refactoring of yeast genome and an antisense-transcriptional regulation toolkit.

Gastrodin, a phenolic glycoside, is a prominent component of Gastrodia elata, which is renowned for its sedative, hypnotic, anticonvulsant, and neuroprotective activities. Engineering heterologous production of plant natural products in microbial host represents a safe, cost-effective, and scalable alternative to plant extraction. Here, we present the construction of an engineered Yarrowia lipolytica yeast that achieves a high-titer production of gastrodin. We systematically refactored the yeast genome by enhancing the flux of the shikimate pathway and optimizing the glucosyl transfer system. We introduced more than five dozen of genetic modifications onto the yeast genome, including enzyme screening, alleviation of rate-limiting steps, promoter selection, genomic integration site optimization, downregulation of competing pathways, and elimination of gastrodin degradation. Meanwhile, we developed a Copper-induced Antisense-Transcriptional Regulation (CATR) tool. The developed CATR toolkit achieved dynamic repression and activation of violacein synthesis through the addition of copper in Y. lipolytica. This strategy was further used to dynamically regulate the pyruvate kinase node to effectively redirect glycolytic flux towards the shikimate pathway while maintaining cell growth at proper rate. Taken together, these efforts resulted in 9477.1 mg/L of gastrodin in shaking flaks and 13.4 g/L of gastrodin with a yield of 0.149 g/g glucose in a 5-L bioreactor, highlighting the potential for large-scale and sustainable production of gastrodin from microbial fermentation.

Molecular characteristics of circulating B cells and kidney cells at the single-cell level in special types of primary membranous nephropathy.

Background: Although primary membranous nephropathy (pMN) associated with podocyte autoantibodies (POS) is becoming well-known, the molecular characteristics of the specific type of pMN that is negative for podocyte autoantibodies (NEG) is still unclear. Methods: We performed single-cell transcriptome sequencing and single-cell B cell receptor sequencing on circulating CD19+ cells and kidney cells of a NEG paediatric patient with pMN. The single-cell datasets of POS patients and healthy control individuals were included for integrative analysis. Results: The gene expression characteristics and clonal expansion of naïve and memory B cells in the NEG patient changed significantly. We found that a group of CD38+ naïve B cells expanded in the NEG patient, which had the functional characteristics of cell activation. In addition, the conversion between immunoglobulin M (IgM)/IgD and IgG1 in the NEG patient was increased. Parietal epithelial cells (PECs) and podocytes shared similar signature genes (WT1, CLIC5), and new candidate marker genes for PECs, such as NID2, CAV1 and THY1, might contribute to the definition of cell subsets. PECs might have undergone significant changes in the disease, mainly manifested by changes in the expression of CCN2, PLAAT4 and SEPTIN2. The scores of gene sets related to extracellular matrix, cell adhesion and calcium channel in podocytes of the NEG patient was significantly increased. The gene expression of sodium transporter in a group of proximal tubule cells in the disease was significantly increased, especially SLC5A12, which might be related to the oedema of patients. Conclusions: Our research demonstrated the cell type-specific molecular features in the circulation and kidney of the NEG pMN patient.

MXene-BN-Introduced Artificial SEI to Inhibit Dendrite Growth of Lithium Metal Batteries.

Lithiophilic substrates have been shown to improve the electrochemical performance of lithium metal anodes. The MXene-BN/Cu 3D current collector was prepared by a filtration method. The artificial solid electrolyte interface (SEI) layer composed of Li3N and LiF was formed on the surface of MXene-BN/Cu during the Li deposition process. Volume changes can be effectively relieved by this special 3D structure. The artificial SEI film reduced the critical dendrite growth length, inhibited Li dendrite growth, and stabilized the electrochemical cycle. MXene-BN/Cu exhibited highly reversible cycling properties during lithium metal deposition with a high Coulombic efficiency of ∼ 98.0% over 500 cycles. Furthermore, LiBH4 was produced during the Li deposition process. This study presents a promising strategy for developing dendrite-free Li anodes for use in lithium metal batteries.

The compound heterozygous mutations of c.607G>a and c.657delC in the FAH gene are associated with renal damage with hereditary tyrosinemia type 1 (HT1).

BACKGROUND: Hereditary tyrosinemia type 1 (HT1) is a rare inherited metabolic disease characterized by severe liver and renal dysfunction. Early identification in affected children is critical for improved treatment options and prognosis. METHODS: In this study, we identified novel compound heterozygous mutations (NM_000137: c.657delC (p.K220Rfs*12) and c.607G>A (p.A203T)) in the fumarylacetoacetate hydrolase (FAH) gene in a family. We also characterized the clinical phenotype of the proband and verified the pathogenic effects of the mutations. Furthermore, we explored the pathogenic mechanism of renal injury through renal biopsy pathology and cell-based in vitro assays. Our study aims to verify the association between novel fumarylacetoacetate hydrolase (FAH) variants and HT1, confirm the pathogenic effects of the mutations and explore the pathogenic mechanism of renal injury. RESULTS: We showed these FAH mutations were inherited in an autosomal recessive manner and resulted in abnormal FAH protein expression and dysfunction, leading to fumarylacetoacetate (FAA) accumulation. The proband also showed apparent renal injury, including glomerular filtration barrier dysfunction and abnormal tubular protein reabsorption. CONCLUSIONS: These observations may provide deeper insights on disease pathogenesis and identify potential therapeutic approaches for HT1 from a genetic perspective. Similarly, we hope to provide valuable information for genetic counseling and prenatal diagnostics.

Expression Characteristics and Significant Prognostic Values of PGK1 in Breast Cancer.

It was proven that PGK1 plays a vital role in the proliferation, migration, and invasion of human breast cancer. However, the correlation of PGK1 mRNA and protein expression with clinicopathologic characteristics and prognostic values according to various kinds of breast cancer patient classifications remains unsufficient. Here, we analyzed data from the Oncomine database, Breast cancer Gene-Expression Miner v4.5, TNMplot, MuTarget, PrognoScan database, and clinical bioinformatics to investigate PGK1 expression distribution and prognostic value in breast cancer patients. Our study revealed that the mRNA and protein expression levels of PGK1 were up-regulated in various clinicopathologic types of breast cancer. Moreover, the expression of PGK1 was correlated with mutations of common tumor suppressor genes TP53 and CDH1. In addition, we found that high mRNA level of PGK1 was significantly associated with poor OS, RFS, and DMFS. Notably, Cox regression analysis showed that PGK1 could be used as an independent prognostic marker. In summary, the aforementioned findings suggested that PGK1 might be not only explored as a potential biomarker, but also combined with TP53/CDH1 for chemotherapy in breast cancer.

Biotransformation mechanism of Vibrio diabolicus to sulfamethoxazole at transcriptional level.

Sulfamethoxazole (SMX) has attracted much attention due to its high probability of detection in the environment. Marine bacteria Vibrio diabolicus strain L2-2 has been proven to be able to transform SMX. In this study, the potential resistance and biotransformation mechanism of strain L2-2 to SMX, and key genes responses to SMX at environmental concentrations were researched. KEGG pathways were enriched by down-regulated genes including degradation of L-Leucine, L-Isoleucine, and fatty acid metabolism. Resistance mechanism could be concluded as the enhancement of membrane transport, antioxidation, response regulator, repair proteins, and ribosome protection. Biotransformation genes might involve in arylamine N-acetyltransferases (nat), cytochrome c553 (cyc-553) and acyl-CoA synthetase (acs). At the environmental concentration of SMX (0.1-10 µg/L), nat was not be activated, which meant the acetylation of SMX might not occur in the environment; however, cyc-553 was up-regulated under SMX stress of 1 µg/L, which indicated the hydroxylation of SMX could occur in the environment. Besides, the membrane transport and antioxidation of strain L2-2 could be activated under SMX stress of 10 µg/L. The results provided a better understanding of resistance and biotransformation of bacteria to SMX and would support related researches about the impacts of environmental antibiotics.

LncRNA GAS5 is upregulated in polycystic ovary syndrome and regulates cell apoptosis and the expression of IL-6.

BACKGROUND: GAS5 contains a hormone response element that can induce cell apoptosis in breast cancer. It is known that cell apoptosis and hormone response play crucial roles in polycystic ovary syndrome (PCOS), indicating the potential involvement of GAS5 in PCOS. This study was performed to investigate the potential involvement of GAS5 and IL-6 (a critical player in PCOS) in PCOS. METHODS: Research subjects of this study included 60 PCOS patients and 60 healthy controls. The expression levels of GAS5 and IL-6 in plasma of both patients and controls were measured by qPCR and ELISA, respectively. Cell transfections were performed to analyze the interaction between GAS5 and IL-6. Cell apoptosis was analyzed by cell apoptosis assay. RESULTS: GAS5 was upregulated in plasma of PCOS patients. The expression levels of GAS5 were positively correlated with the expression levels of IL-6. Altered expression levels of GAS5 and IL-6 distinguished PCOS patients from healthy controls. In cells of a granulosa-like tumor cell line (KGN), overexpression of GAS5 led to upregulated IL-6, while silencing of GAS5 played an opposite role. Cell apoptosis analysis showed that overexpression of GAS5 significantly decreased apoptosis rate of KGN cells. Silencing of GAS5 increased the rate of KGN cell apoptosis. CONCLUSIONS: GAS5 is upregulated in PCOS and regulates cell apoptosis and the expression of IL-6.

Isolation and characterization of a marine bacterium Vibrio diabolicus strain L2-2 capable of biotransforming sulfonamides.

Sulfonamides (SAs) have attracted much attention because of their high detection rates in natural water. In this study, a marine bacterium Vibrio diabolicus strain L2-2 was isolated which could metabolize 9 SAs to a different extent. Compared with SAs and their analogs, SAs with N-oxides of heterocyclic structure were easier to be transformed to their N4-acetylated metabolites or their isoxazole ring rearrangement isomers by strain L2-2. And, gene vdnatA and vdnatG were likely to be the key genes in SAs acetylation process, which might code Arylamine N-acetyltransferase. The biotransformation rates of sulfathiazole(STZ), sulfamonomethoxine(SMT), sulfadiazine(SDZ), sulfamethoxazole(SMX) and sulfisoxazole(SIX) could reach 29.39 ± 5.63, 24.97 ± 4.45, 79.41 ± 4.05, 64.64 ± 1.71, 32.82 ± 4.46% in 6 days, respectively. Besides, the overall optimal conditions for SAs biotransformation were less than 100 mg/L for total SAs in neutral or weakly alkaline medium with the salinity of 10-20‰ and additional nutrients like glucose, sucrose or glycerine. Furthermore, toxicity was demonstrated to be significantly reduced after biotransformation. Together, this study introduced a strategy to use V. diabolicus strain L2-2 to realize simultaneous removal and detoxification of multiple SAs in freshwater and seawater, and revealed SAs removal pathways and relevant molecular mechanism.

Long non-coding RNA TUG1 sponges miR-197 to enhance cisplatin sensitivity in triple negative breast cancer.

Breast cancer is the leading cause of women death worldwide. Several long non-coding RNAs (lncRNAs) have been identified as oncogenes or tumor suppressors during the progression of cancers. However, the role of taurine upregulated gene (TUG1) in mediating the chemotherapy sensitivity of triple negative breast cancer (TNBC) has not been studied yet. In TNBC patients, we observed a significant decrease of TUG1 in tumor tissues compared to the normal tissues. Similarly, TUG1 expression was significantly decreased in TNBC cell lines compared with normal breast epithelial cell line and cell lines of other subtypes of breast cancer. In MDA-MB-231 and BT549, cisplatin induced cell growth arrest was remarkably augmented by overexpression of TUG1 and was significantly reduced by TUG1 silencing. Moreover, very low concentration of cisplatin caused cell proliferation inhibition in TUG1-overexpressed-TNBC cells. In addition, we found that TUG1 negatively regulated miR-197 expression in the tested TNBC cell lines. Sponging of TUG1 to miR-197 was proved by a dual luciferase reporter assay. We further predicted and validated that nemo-like kinase (NLK), which was positively controlled by TUG1, was a target gene of miR-197. Via regulation of miR-197/NLK, TUG1 inactivated WNT signaling pathway and thus increasing chemotherapy sensitivity of TNBC cells. Analysis of TCGA database showed that higher expression of TUG1 was associated with better prognosis in breast cancer patients. Our current study drew a preliminary conclusion that TUG1 was involved in chemotherapy sensitivity in TNBC cells.

3D architecture of a graphene/CoMoO(4) composite for asymmetric supercapacitors usable at various temperatures.

Designing and optimizing the electrode materials and studying the electrochemical performance or cycle life of the supercapacitor under different working conditions are crucial to its practical application. Herein, we proposed a rational design of 3D-graphene/CoMoO4 nanoplates by a facile two-step hydrothermal method. Owing to the high electron transfer rate of graphene and the high activity of the CoMoO4 nanoplates, the three-dimensional electrode architectures achieved remarkable electrochemical performances with high areal specific capacitance (1255.24F/g at 1A/g) and superior cycling stability (91.3% of the original specific capacitance after 3000 cycles at 1A/g). The all-solid-state asymmetric supercapacitor composed of 3D-graphene/CoMoO4 and activated carbon (AC) exhibited a specific capacitance of 109F/g at 0.2A/g and an excellent cycling stability with only 12.1% of the initial specific capacitance off after 3000 cycles at 2A/g. The effects of temperature and charge-discharge current densities on the charge storage capacity of the supercapacitor were also investigated in detail for practical applications.

High carrier concentration ZnO nanowire arrays for binder-free conductive support of supercapacitors electrodes by Al doping.

Doping semiconductor nanowires (NWs) for altering their electrical and optical properties is a critical strategy for tailoring the performance of nanodevices. Here, we prepared in situ Al-doped ZnO nanowire arrays by using continuous flow injection (CFI) hydrothermal method to promote the conductivity. This reasonable method offers highly stable precursor concentration for doping that effectively avoid the appearance of the low conductivity ZnO nanosheets. Benefit from this, three orders of magnitude rise of the carrier concentration from 1016cm-3 to 1019cm-3 can be achieved compared with the common hydrothermal (CH) mothed in Mott-Schottky measurement. Possible effect of Al-doping was discussed by first-principle theory. On this basis, Al-doped ZnO nanowire arrays was developed as a binder-free conductive support for supercapacitor electrodes and high capacitance was triggered. It is owing to the dramatically decreased transfer resistance induced by the growing free-moving electrons and holes. Our results have a profound significance not merely in the controlled synthesis of other doping nanomaterials by co-precipitation method but also in the application of binder-free energy materials or other materials.