Transcriptomic profiling of adding cobalt chloride to improve dendrobine-type total alkaloid production.

Trichoderma longibrachiatum UN32 is known for its efficient production of dendrobine-type total alkaloids (DTTAs). This study aimed to determine the optimal medium composition for the UN32 strain using response surface methodology. Key factors, including glucose, beef extract, and CoCl2, were selected through the Plackett-Burman design. Subsequently, a factorial optimization approach was employed using the steepest ascent design, and 17 trial sets were completed via the Box-Behnken design. The optimal medium composition was found to consist of 29.4 g/L of glucose, 17.3 g/L of beef extract, and 0.28 mmol/L of CoCl2. This optimized medium resulted in an impressive 80.8% increase in mycelial dry weight (reaching 12.303 g/L) and a substantial 76.4% boost in DTTA production (reaching 541.63 ± 46.95 µg). Furthermore, the fermentation process was scaled up to a 5-L bioreactor, leading to a DTTA production approximately 1.95 times than the control. Transcriptome analysis of strain UN32 in response to CoCl2 supplementation revealed significant changes in the expression of critical genes associated with the TCA cycle and L-valine, L-leucine, and L-isoleucine biosynthesis changed. These alterations resulted in a heightened influx of acetyl-CoA into DTTA production. Additionally, the expression of genes related to antioxidant enzymes was modified to maintain homeostasis of reactive oxygen species (ROS). A potential mechanism for the accumulation of DTTAs based on ROS as a signal transduction was proposed. These findings provide valuable insights into the regulatory mechanisms of DTTA biosynthesis, potentially offering a method to enhance the production of secondary metabolites in the UN32 strain. KEY POINTS: • After the RSM optimization, there is a substantial increase of 80.8% in biomass production and a significant 76.4% rise in DTTA production. • Transcriptome analysis revealed that the inclusion of CoCl2 supplements resulted in an enhanced influx of acetyl-CoA. • Proposed a mechanism for the accumulation of DTTAs for the role of ROS as a signal transduction pathway.

Atomic Force Microscopy for Tumor Research at Cell and Molecule Levels.

Tumors have posed a serious threat to human life and health. Researchers can determine whether or not cells are cancerous, whether the cancer cells are invasive or metastatic, and what the effects of drugs are on cancer cells by the physical properties such as hardness, adhesion, and Young's modulus. The atomic force microscope (AFM) has emerged as a key important tool for biomechanics research on tumor cells due to its ability to image and collect force spectroscopy information of biological samples with nano-level spatial resolution and under near-physiological conditions. This article reviews the existing results of the study of cancer cells with AFM. The main foci are the operating principle of AFM and research advances in mechanical property measurement, ultra-microtopography, and molecular recognition of tumor cells, which allows us to outline what we do know it in a systematic way and to summarize and to discuss future directions.

Solvent Effect on Product Distribution in the Aerobic Autoxidation of 2-Ethylhexanal: Critical Role of Polarity.

In the aerobic oxidation of aldehydes to acids, how the solvent affect the reaction remains unclear. Herein, the solvent effect in the oxidation of 2-ethylhexanal (2-ETH) to 2-ethylhexanoic acid (2-ETA) was systematically investigated. The vastly different product distributions were observed which could be ascribed to the dominant intermolecular forces. Though strong intermolecular forces in protic solvents limit the oxidation, the optimal 2-ETA yield (96%) was obtained in ipropanol via gradually evaporating the solvent to remove the interactions. Theoretical calculations further revealed that the hydrogen bonds between reactant and protic solvent increase the C-H bond energy (-CHO in 2-ETH). Meanwhile, the hydrogen bonds may improve 2-ETA selectivity by promoting H transfer in the oxidation rearrangement step. Our work discloses the critical role of polarity in determining the reactivity and selectivity of 2-ETH oxidation, and could guide the rational design of more desirable reaction processes with solvent effect.

Comparative Transcriptome Analysis of Genes Involved in Sesquiterpene Alkaloid Biosynthesis in Trichoderma longibrachiatum MD33 and UN32.

Trichoderma longibrachiatum MD33, a sesquiterpene alkaloid-producing endophyte isolated from Dendrobium nobile, shows potential medical and industrial applications. To understand the molecular mechanisms of sesquiterpene alkaloids production, a comparative transcriptome analysis was performed on strain MD33 and its positive mutant UN32, which was created using Ultraviolet (UV) mutagenesis and nitrogen ion (N+) implantation. The alkaloid production of UN32 was 2.62 times more than that of MD33. One thousand twenty-four differentially expressed genes (DEGs), including 519 up-regulated and 505 down-regulated genes, were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed 139 GO terms and 87 biosynthesis pathways. Dendrobine, arguably the main sesquiterpene alkaloid the strain MD33 produced, might start synthesis through the mevalonate (MVA) pathway. Several MVA pathway enzyme-coding genes (hydroxy-methylglutaryl-CoA synthase, mevalonate kinase, and farnesyl diphosphate synthase) were found to be differentially expressed, suggesting that physical mutagenesis can disrupt genome integrity and gene expression. Some backbone post-modification enzymes and transcript factors were either discovered, suggesting the sesquiterpene alkaloid metabolism in T. longibrachiatum is a complex genetic network. Our findings help to shed light on the underlying molecular regulatory mechanism of sesquiterpene alkaloids production in T. longibrachiatum.