TCF7L2 acts as a molecular switch in midbrain to control mammal vocalization through a transcriptional repression mechanism

Vocalization is an essential medium for sexual and social signaling in birds and mammals. Periaqueductal gray (PAG) a conserved midbrain structure is believed to be responsible for innate vocalizations, but its molecular regulation remains largely unknown. Here, through a mouse forward genetic screening we identified one of the key Wnt/β-catenin effectors TCF7L2/TCF4 controls ultrasonic vocalization (USV) production and syllable complexity during maternal deprivation and sexual encounter. Expression of TCF7L2 in PAG excitatory neurons is necessary for the complex trait, while TCF7L2 loss reduces neuronal gene expressions and synaptic transmission in PAG. TCF7L2-mediated vocal control is independent of its β-catenin-binding domain but dependent of its DNA binding ability. Patient mutations associated with severe speech delay disrupt the transcriptional repression effect of TCF7L2, while mice carrying those mutations display severe USV impairments. Therefore, we conclude that TCF7L2 orchestrates gene expression in midbrain to control vocal production through a transcriptional repression mechanism.

DNA binding by the antimalarial compound artemisinin

Artemisinin (ART) is a vital medicinal compound that is used alone or as part of a combination therapy against malaria. ART is thought to function by attaching to heme covalently and alkylating a range of proteins. Using a combination of biophysical methods, we demonstrate that ART is bound by three-way junction and duplex containing DNA molecules. Binding of ART by DNA is first shown for the cocaine-binding DNA aptamer and extensively studied using this DNA molecule. Isothermal titration calorimetry methods show that the binding of ART is both entropically and enthalpically driven at physiological NaCl concentration. Native mass spectrometry methods confirm DNA binding and show that a non-covalent complex is formed. Nuclear magnetic resonance spectroscopy shows that ART binds at the three-way junction of the cocaine-binding aptamer, and that binding results in the folding of the structure-switching variant of this aptamer. This structure-switching ability was exploited using the photochrome aptamer switch assay to demonstrate that ART can be detected using this biosensing assay. This study is the first to demonstrate the DNA binding ability of ART and should lay the foundation for further work to study implications of DNA binding for the antimalarial activity of ART.

Bacterial infections exacerbate myeloma bone disease

Multiple myeloma is characterized by osteolytic lesions caused by reduced bone formation and activated bone resorption. An important feature of myeloma is a failure of bone healing after successful treatment. In this work, clinical studies indicated a highly positive correlation between bone marrow bacteria abundance and bone lesion numbers of myeloma patients in complete remission. Coculture experiments demonstrated that marrow Escherichia coli (E. coli) promotes osteoclast differentiation and inhibits osteoblast differentiation. Mechanism studies showed that E. coli lipopolysaccharides (LPS) activated NF-κB p65 signaling and reduced phosphorylated smad1/5/9 binding ability with RUNX2 promoter, leading to decreased RUNX2 expression in osteoblast progenitors. Additionally, LPS enhanced phosphorylated NF-κB p65 binding ability with NFATc1 promoter, leading to increased NFATc1 expression in osteoclast progenitors. In vivo studies revealed E. coli contributes to osteolytic bone lesion, and elimination of E. coli infection assists healing of bone lesion in mouse model of myeloma in complete remission. These findings establish a heretofore unrecognized effect for E. coli in the genesis of myeloma bone disease and suggest a new treatment strategy.

The Optimization of Catfish Smart Flavor Production by Biduri and Papain Enzymatic Hydrolysis

The consumption of Monosodium Glutamate with a large amount can lead to nerve cell damage to the brain so that natural ingredients substitute MSG is needed. In this research, we produced smart flavors from catfish through enzymatic hydrolysis by combining papain and biduri enzymes. The purpose of the study was to identify the influence of enzyme concentration and length of hydrolysis on the smart flavor characteristics and determine the best treatment to produce smart flavors. The parameters identified were color, yield, moisture content, dissolved proteins, degrees of hydrolysis, antioxidants, water binding ability, and emulsion stability. The results show the highest brightness are biduri and papain combination by 60:40 with one-hour hydrolysis. The highest dissolved protein is 50:50 combination with three-hour hydrolysis. In addition, antioxidant activity is marked in a combination of 50:50 with one-hour hydrolysis.

OligoTRAFTACs: A Generalizable Method for Transcription Factor Degradation

Dysregulated transcription factors (TFs) that rewire gene expression circuitry are frequently identified as key players in disease. Although several TFs have been drugged with small molecules, the majority of oncogenic TFs are not currently pharmaceutically tractable due to their paucity of ligandable pockets. The first generation of transcription factor targeting chimeras (TRAFTACs) was developed to target TFs for proteasomal degradation by exploiting their DNA binding ability. In the current study, we have developed the second generation TRAFTACs (oligoTRAFTACs) comprised of a TF-binding oligonucleotide and an E3 ligase-recruiting ligand. Herein, we demonstrate the development of oligoTRAFTACs to induce the degradation of two oncogenic TFs, c-Myc and brachyury. In addition, we show that brachyury can be successfully degraded by oligoTRAFTACs in chordoma cell lines. Furthermore, zebrafish experiments demonstrate in vivo oligoTRAFTAC activity. Overall, our data demonstrate oligoTRAFTACs as a generalizable platform towards difficult-to-drug TFs and their degradability via the proteasomal pathway.

Structural and Functional Changes of Reconstituted High-Density Lipoprotein (HDL) by Incorporation of α-synuclein: A Potent Antioxidant and Anti-Glycation Activity of α-synuclein and apoA-I in HDL at High Molar Ratio of α-synuclein

α-synuclein (α-syn) is a major culprit of Parkinson’s disease (PD), although lipoprotein metabolism is very important in the pathogenesis of PD. α-syn was expressed and purified using the pET30a expression vector from an E. coli expression system to elucidate the physiological effects of α-syn on lipoprotein metabolism. The human α-syn protein (140 amino acids) with His-tag (8 amino acids) was expressed and purified to at least 95% purity. Isoelectric focusing gel electrophoresis showed that the isoelectric point (pI) of α-syn and apoA-I were pI = 4.5 and pI = 6.4, respectively. The lipid-free α-syn showed almost no phospholipid-binding ability, while apoA-I showed rapid binding ability with a half-time (T1/2) = 8 ± 0.7 min. The α-syn and apoA-I could be incorporated into the reconstituted HDL (rHDL, molar ratio 95:5:1:1, palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC):cholesterol:apoA-I:α-syn with the production of larger particles (92 Å) than apoA-I-rHDL (86 and 78 Å) and α-syn-rHDL (65 Å). An rHDL containing both apoA-I and α-syn showed lower α-helicity around 45% with a red shift of the Trp wavelength maximum fluorescence (WMF) from 339 nm, while apoA-I-HDL showed 76% α-helicity and 337 nm of WMF. The denaturation by urea addition showed that the incorporation of α-syn in rHDL caused a larger increase in the WMF than apoA-I-rHDL, suggesting that the destabilization of the secondary structure of apoA-I by the addition of α-syn. On the other hand, the addition of α-syn induced two-times higher resistance to rHDL glycation at apoA-I:α-syn molar ratios of 1:1 and 1:2. Interestingly, low α-syn in rHDL concentrations, molar ratio of 1:0.5 (apoA-I:α-syn), did not prevent glycation with more multimerization of apoA-I. In the lipid-free and lipid-bound state, α-syn showed more potent antioxidant activity than apoA-I against cupric ion-mediated LDL oxidation. On the other hand, microinjection of α-syn (final 2 μM) resulted in 10% less survival of zebrafish embryos than apoA-I. A subcutaneous injection of α-syn (final 34 μM) resulted in less tail fin regeneration than apoA-I. Interestingly, incorporation of α-syn at a low molar ratio (apoA-I:α-syn, 1:0.5) in rHDL resulted destabilization of the secondary structure and impairment of apoA-I functionality via more oxidation and glycation. However, at a higher molar ratio of α-syn in rHDL (apoA-I:α-syn = 1:1 or 1:2) exhibited potent antioxidant and anti-glycation activity without aggregation. In conclusion, there might be a critical concentration of α-syn and apoA-I in HDL-like complex to prevent the aggregation of apoA-I via structural and functional enhancement.

The Nuts and Bolts of SARS-CoV-2 Spike Receptor-Binding Domain Heterologous Expression

COVID-19 is a highly infectious disease caused by a newly emerged coronavirus (SARS-CoV-2) that has rapidly progressed into a pandemic. This unprecedent emergency has stressed the significance of developing effective therapeutics to fight the current and future outbreaks. The receptor-binding domain (RBD) of the SARS-CoV-2 surface Spike protein is the main target for vaccines and represents a helpful “tool” to produce neutralizing antibodies or diagnostic kits. In this work, we provide a detailed characterization of the native RBD produced in three major model systems: Escherichia coli, insect and HEK-293 cells. Circular dichroism, gel filtration chromatography and thermal denaturation experiments indicated that recombinant SARS-CoV-2 RBD proteins are stable and correctly folded. In addition, their functionality and receptor-binding ability were further evaluated through ELISA, flow cytometry assays and bio-layer interferometry.

Research of Vertical Migration and Occurrence of Cd and As in Acid Paddy Soil under Simulated Irrigation

According to the research of vertical migration and occurrence of Cd and As in acid paddy soil, the conclusions are as follows. The longitudinal migration of Cd in TZkb1, TZ1, TZ2, TZ3 and TZ5 found that Cd in the upper and lower layers of soil columns all lost to varying degrees with the increase of water injection contents (irrigation years). With the longitudinal leaching of irrigation water, when the adsorption capacity of the soil was greater than the water injection and rinsing capacity, the trend of accumulation (TZ4-TZ6) further appeared. The vertical migration results of As in TZkb1, TZ1, TZ2, TZ3 and TZ5 found that As mainly accumulated in the upper layer of soil columns. Without considering other output ways, it can be seen that the binding ability of As to acid paddy soil was stronger than Cd, and its cumulative efficiency in 20 years was higher than Cd. However Cd cumulative efficiency of TZ3 and TZ5 in acid paddy soil may be greater than As with time. In general, As was less affected by irrigation water on vertical leaching of soil columns compared with Cd.

Preparation of a Molecularly Imprinted Film on Quartz Crystal Microbalance Chip for Determination of Furanic Compounds

The structural preferences of furanic compounds were studied using a combination of a molecularly imprinted film (MIF) on a piezoelectric-quartz chip. The furanic compounds and their derivatives were used as the templates. Owing to their similar heterocyclic structures, it is difficult to verify the structural differences between the templates. Therefore, a new cross-linker (Methacr-l-Cys-NHBn)2, was employed to generate a platform on a quartz crystal microbalance (QCM) chip. The cross-linker self-assembled to link the surface of the chip to copolymerize with other functional monomers. A layered film with chiral hydrophobicity and rigidity was thus fabricated. Subsequently, Acr-l-Ser-NHBn was utilized as a chiral monomer to construct MIF on a QCM chip. Forcomparison, we synthesized a more hydrophobic monomer, Methacr-l-Ser-NHBn, to enhance the binding ability of the MIF. The QCM flow injection system was handled in an organic solvent system. The proportion of the monomers was adjusted to optimize the recognition ability of these films. As the binding ability of the MIF toward model templates and structurally-related furanic compounds was improved, a MIF derived from 2-furaldehyde (FUL) achieved a lower detection limit (10 ng/mL). The binding properties of MIFs prepared against furanic compounds exhibited strong similarities to the binding properties of other compounds with heterocyclic ring structures. For example, 2-furaldehyde is very similar to 2-formylthiazole, 2-acetylfuran is similar to 2-acetylthiazole, and 2-furfuryl alcohol is similar to imidazole-2-methanol. Such recognition ability can help distinguish between the structural counterparts of other small heterocyclic compounds.