Brassinosteroids biosynthetic gene MdBR6OX2 regulates salt stress tolerance in both apple and Arabidopsis.

Salt stress is a critical limiting factor for fruit yield and quality of apples. Brassinosteroids (BRs) play an important role in response to abiotic stresses. In the present study, application of 2,4- Epicastasterone on seedlings of Malus 'M9T337' and Malus domestica 'Gala3' alleviated the physiological effects, such as growth inhibition and leaf yellowing, induced by salt stress. Further analysis revealed that treatment with NaCl induced expression of genes involved in BR biosynthesis in 'M9T337' and 'Gala3'. Among which, the expression of BR biosynthetic gene MdBR6OX2 showed a three-fold upregulation upon salt treatment, suggesting its potential role in response to salt stress in apple. MdBR6OX2, belonging to the CYP450 family, contains a signal peptide region and a P450 domain. Expression patterns analysis showed that the expression of MdBR6OX2 can be significantly induced by different abiotic stresses. Overexpressing MdBR6OX2 enhanced the tolerance of apple callis to salt stress, and the contents of endogenous BR-related compounds, such as Typhastero (TY), Castasterone (CS) and Brassinolide (BL) were significantly increased in transgenic calli compared with that of wild-type. Extopic expression of MdBR6OX2 enhanced tolerance to salt stress in Arabidopsis. Genes associated with salt stress were significantly up-regulated, and the contents of BR-related compounds were significantly elevated under salt stress. Our data revealed that BR-biosynthetic gene MdBR6OX2 positively regulates salt stress tolerance in both apple calli and Arabidopsis.

PROSCA: an online platform for humanized scaffold mining facilitating rational protein engineering.

Protein scaffolds with small size, high stability and low immunogenicity show important applications in the field of protein engineering and design. However, no relevant computational platform has been reported yet to mining such scaffolds with the desired properties from massive protein structures in human body. Here, we developed PROSCA, a structure-based online platform dedicated to explore the space of the entire human proteome, and to discovery new privileged protein scaffolds with potential engineering value that have never been noticed. PROSCA accepts structure of protein as an input, which can be subsequently aligned with a certain class of protein structures (e.g. the human proteome either from experientially resolved or AlphaFold2 predicted structures, and the human proteins belonging to specific families or domains), and outputs humanized protein scaffolds which are structurally similar with the input protein as well as other related important information such as families, sequences, structures and expression level in human tissues. Through PROSCA, the user can also get excellent experience in visualizations of protein structures and expression overviews, and download the figures and tables of results which can be customized according to the user's needs. Along with the advanced protein engineering and selection technologies, PROSCA will facilitate the rational design of new functional proteins with privileged scaffolds. PROSCA is freely available at https://idrblab.org/prosca/.

Exploring the maximum nitrite production rate through the granular sludge-type reactor to match the needs of anammox process realizing efficient nitrogen removal.

The reliable and efficient nitrite production rate (NPR) through nitritation process is the prerequisite for the efficient running of subsequent processes, like the anammox process and the nitrite shunt. However, there has been scant research on stable and productive nitritation process in recent years. In this study, at a stable hydraulic retention time of 12.0 h and with precise and strict DO control, the upper limit of the NPR was initially investigated using a continuous-flow granular sludge reactor. The NPR of 1.69 kg/m3/d with a nitrite production efficiency of 81.97% was finally achieved, which set a record until now in similar research. The median sludge particle size of 270.0 µm confirmed the development of clearly defined granular sludge. The genus Nitrosomonas was the major ammonium oxidizing bacteria. In conclusion, this study provides valuable insights for the practical application of the effective nitritation process driving subsequent nitrogen removal processes.

Brønsted acid catalyzed Ficini [2 + 2] cycloaddition of ynamides with enones.

Herein, we describe a novel metal-free Brønsted acid-catalyzed Ficini [2 + 2] cycloaddition of ynamides with enones under mild reaction conditions, leading to the formation of various cyclobutenamides in generally good to excellent yields within short reaction times. This work represents the first example of ynamides involved in a nonmetal-catalyzed [2 + 2] cycloaddition with enones.

TMSOTf-Catalyzed [4 + 2] Annulation of Ynamides and ß-(2-Aminophenyl)-α,ß-ynones for the Synthesis 2-Aminoquinolines.

A metal-free TMSOTf-catalyzed [4 + 2] annulation of ynamides with ß-(2-aminophenyl)-α,ß-ynones enables the regiospecific and facile assembly of 2-aminoquinoline frameworks. The catalyst TMSOTf presented a remarkable advancement compared to previously reported transition-metal catalysts. A wide range of 3-aryl/alkyl-substituted 2-aminoquinolines were generated in moderate to excellent yields due to the mild conditions.

Design, synthesis, and evaluation of novel quindoline derivatives with fork-shaped side chains as RNA G-quadruplex stabilizers for repressing oncogene NRAS translation.

NRAS mutation is the second most common oncogenic factor in cutaneous melanoma. Inhibiting NRAS translation by stabilizing the G-quadruplex (G4) structure with small molecules seems to be a potential strategy for cancer therapy due to the NRAS protein's lack of a druggable pocket. To enhance the effects of previously reported G4 stabilizers quindoline derivatives, we designed and synthesized a novel series of quindoline derivatives with fork-shaped side chains by introducing (alkylamino)alkoxy side chains. Panels of experimental results showed that introducing a fork-shaped (alkylamino)alkoxy side chain could enhance the stabilizing abilities of the ligands against NRAS RNA G-quadruplexes and their anti-melanoma activities. One of them, 10b, exhibited good antitumor activity in the NRAS-mutant melanoma xenograft mouse model, showing the therapeutic potential of this kind of compounds.

Knittable Electrochemical Yarn Muscle for Morphing Textiles.

Morphing textiles, crafted using electrochemical artificial muscle yarns, boast features such as adaptive structural flexibility, programmable control, low operating voltage, and minimal thermal effect. However, the progression of these textiles is still impeded by the challenges in the continuous production of these yarn muscles and the necessity for proper structure designs that bypass operation in extensive electrolyte environments. Herein, a meters-long sheath-core structured carbon nanotube (CNT)/nylon composite yarn muscle is continuously prepared. The nylon core not only reduces the consumption of CNTs but also amplifies the surface area for interaction between the CNT yarn and the electrolyte, leading to an enhanced effective actuation volume. When driven electrochemically, the CNT@nylon yarn muscle demonstrates a maximum contractile stroke of 26.4%, a maximum contractile rate of 15.8% s-1, and a maximum power density of 0.37 W g-1, surpassing pure CNT yarn muscles by 1.59, 1.82, and 5.5 times, respectively. By knitting the electrochemical CNT@nylon artificial muscle yarns into a soft fabric that serves as both a soft scaffold and an electrolyte container, we achieved a morphing textile is achieved. This textile can perform programmable multiple motion modes in air such as contraction and sectional bending.

Correction: Teng et al. Efficacy Assessment of Phage Therapy in Treating Staphylococcus aureus-Induced Mastitis in Mice. Viruses 2022, 14, 620.

In the original publication [...].

miR164-MhNAC1 regulates apple root nitrogen uptake under low nitrogen stress.

Nitrogen is an essential nutrient for plant growth and serves as a signaling molecule to regulate gene expression inducing physiological, growth and developmental responses. An excess or deficiency of nitrogen may have adverse effects on plants. Studying nitrogen uptake will help us understand the molecular mechanisms of utilization for targeted molecular breeding. Here, we identified and functionally validated an NAC (NAM-ATAF1/2-CUC2) transcription factor based on the transcriptomes of two apple rootstocks with different nitrogen uptake efficiency. NAC1, a target gene of miR164, directly regulates the expression of the high-affinity nitrate transporter (MhNRT2.4) and citric acid transporter (MhMATE), affecting root nitrogen uptake. To examine the role of MhNAC1 in nitrogen uptake, we produced transgenic lines that overexpressed or silenced MhNAC1. Silencing MhNAC1 promoted nitrogen uptake and citric acid secretion in roots, and enhanced plant tolerance to low nitrogen conditions, while overexpression of MhNAC1 or silencing miR164 had the opposite effect. This study not only revealed the role of the miR164-MhNAC1 module in nitrogen uptake in apple rootstocks but also confirmed that citric acid secretion in roots affected nitrogen uptake, which provides a research basis for efficient nitrogen utilization and molecular breeding in apple.

Dense Multibutterfly Spots-Enhanced Miniaturized Optical Fiber Photoacoustic Gas Sensor.

A miniaturized optical fiber photoacoustic gas sensor enhanced by dense multibutterfly spots is reported for the first time. The principle of space light transmission of neglecting paraxial approximation is theoretically analyzed for designing a dense multibutterfly spots-based miniature multipass cell. In a multipass photoacoustic tube with a diameter of 16 mm, the light beam is reflected about a hundred times. The light spots on the mirror surfaces at both ends of the photoacoustic tube form a dense multibutterfly distribution. The volume of the micro multipass gas chamber is only 5.3 mL. An optical fiber cantilever based on F-P interference is utilized as a photoacoustic pressure detector. Compared with that of the single-pass structure, the gas detection ability of the photoacoustic system with dense multibutterfly spots is improved by about 50 times. The proposed miniaturized sensor realizes a detection limit of 3.4 ppb for C2H2 gas with an averaging time of 100 s. The recognized coefficients of minimum detectable absorption (αmin) and normalized noise equivalent absorption are 1.9 × 10-8 cm-1 and 8.4 × 10-10 W cm-1 Hz-1/2, respectively.

Association between weight change, waist circumference change, and the risk of nonalcoholic fatty liver disease in individuals with metabolically healthy overweight or obesity and metabolically unhealthy overweight or obesity.

BACKGROUND: This study aimed to explore and compare the effect of weight change, and waist circumference (WC) change, on the risk of nonalcoholic fatty liver disease (NAFLD) in individuals with metabolically healthy overweight or obesity (MHOW/O) and metabolically unhealthy overweight or obesity (MUOW/O) in a health check-up cohort in China. METHODS: 5625 adults with overweight or obesity, and free from NAFLD at baseline were included. Metabolically healthy was defined as not having any components of metabolic syndrome. Weight/WC changes were calculated as the relative difference between the first and second visits of check-up. NAFLD was assessed based on abdominal ultrasound. RESULTS: During a median follow-up of 2.1 (IQR: 1.1-4.3) years, 1849 participants developed NAFLD. In MHOW/O participants, the multivariable adjusted HRs (95 % CIs) for NAFLD in weight change ≤ -5.0 %, and - 4.9-- 1.0 % were 0.36 (0.23-0.59), 0.59 (0.43-0.80), respectively, compared to the weight stable group (-0.9% to 0.9 %). The corresponding HRs (95 % CIs) for the association between WC change (≤ 6.0 %, - 5.9 to -3.0 %) and NAFLD in MHOW/O participants were 0.41 (0.27-0.62), and 0.74 (0.54-1.01), respectively, compared to the WC stable group (-2.9-2.9 %). Similar patterns were observed in MUOW/O participants. A more marked gradient of cumulative incidence of NAFLD across weight/WC change categories was observed in MHOW/O than in MUOW/O individuals. CONCLUSIONS: A more evident association between weight/WC loss and risk of NAFLD was observed in MHOW/O than in MUOW/O individuals. Our findings indicate the practical significance of encouraging all individuals with overweight and obesity to achieve a clinically relevant level of weight/WC loss to prevent NAFLD, even among metabolic healthy groups.

Assessing immunogenicity of CRISPR-NCas9 engineered strain against porcine epidemic diarrhea virus.

Porcine epidemic diarrhea (PED) caused by porcine epidemic diarrhea virus (PEDV), is an acute and highly infectious disease, resulting in substantial economic losses in the pig industry. Given that PEDV primarily infects the mucosal surfaces of the intestinal tract, it is crucial to improve the mucosal immunity to prevent viral invasion. Lactic acid bacteria (LAB) oral vaccines offer unique advantages and potential applications in combatting mucosal infectious diseases, making them an ideal approach for controlling PED outbreaks. However, traditional LAB oral vaccines use plasmids for exogenous protein expression and antibiotic genes as selection markers. Antibiotic genes can be diffused through transposition, transfer, or homologous recombination, resulting in the generation of drug-resistant strains. To overcome these issues, genome-editing technology has been developed to achieve gene expression in LAB genomes. In this study, we used the CRISPR-NCas9 system to integrate the PEDV S1 gene into the genome of alanine racemase-deficient Lactobacillus paracasei △Alr HLJ-27 (L. paracasei △Alr HLJ-27) at the thymidylate synthase (thyA) site, generating a strain, S1/△Alr HLJ-27. We conducted immunization assays in mice and piglets to evaluate the level of immune response and evaluated its protective effect against PEDV through challenge tests in piglets. Oral administration of the strain S1/△Alr HLJ-27 in mice and piglets elicited mucosal, humoral, and cellular immune responses. The strain also exhibited a certain level of resistance against PEDV infection in piglets. These results demonstrate the potential of S1/△Alr HLJ-27 as an oral vaccine candidate for PEDV control. KEY POINTS: • A strain S1/△Alr HLJ-27 was constructed as the candidate for an oral vaccine. • Immunogenicity response and challenge test was carried out to analyze the ability of the strain. • The strain S1/△Alr HLJ-27 could provide protection for piglets to a certain extent.

SIRT2 promotes base excision repair by transcriptionally activating OGG1 in an ATM/ATR-dependent manner.

Sirtuin 2 (SIRT2) regulates the maintenance of genome integrity by targeting pathways of DNA damage response and homologous recombination repair. However, whether and how SIRT2 promotes base excision repair (BER) remain to be determined. Here, we found that independent of its catalytic activity SIRT2 interacted with the critical glycosylase OGG1 to promote OGG1 recruitment to its own promoter upon oxidative stress, thereby enhancing OGG1 promoter activity and increasing BER efficiency. Further studies revealed that SIRT2 was phosphorylated on S46 and S53 by ATM/ATR upon oxidative stress, and SIRT2 phosphorylation enhanced the SIRT2-OGG1 interaction and mediated the stimulatory effect of SIRT2 on OGG1 promoter activity. We also characterized 37 cancer-derived SIRT2 mutants and found that 5 exhibited the loss of the stimulatory effects on OGG1 transcription. Together, our data reveal that SIRT2 acts as a tumor suppressor by promoting OGG1 transcription and increasing BER efficiency in an ATM/ATR-dependent manner.

Epstein-Barr virus-positive iris diffuse large B-cell lymphoma detected by metagenomic next-generation sequencing.

PURPOSE: Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma (DLBCL) is a relatively rare subtype of DLBCL. Herein, we report a case of a patient with EBV-positive iris DLBCL after undergoing penetrating keratoplasty and discuss its possible pathogenesis. METHODS: A 72-year-old male patient presented to our hospital with progressive blurring of vision in the left eye for the past 4 months. Small white nodular lesions were observed on the iris and retinal surface of the left eye, with a white cloud-like opacity in the vitreous cavity. RESULTS: The patient was eventually diagnosed with EBV-positive iris DLBCL after undergoing pathological and metagenomic tests. After injecting methotrexate in the left vitreous cavity and administering systemic and local antiviral treatments, the ocular lesions disappeared. CONCLUSION: EBV infection, drug immunosuppression, and aging-related immune deterioration may play significant roles in the pathogenesis of EBV-positive iris DLBCL. SYNOPSIS: Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma (DLBCL) is a new subtype of DLBCL, which rarely occurs. Herein, we report a case of a patient with EBV-positive iris DLBCL after undergoing penetrating keratoplasty and discuss its possible pathogenesis.

Tempol alleviates acute lung injury by affecting glutathione synthesis through Nrf2 and inhibiting ferroptosis in lung epithelial cells.

As a life-threatening disease, acute lung injury (ALI) may progress to chronic pulmonary fibrosis. For the treatment of lung injury, Tempol is a superoxide dismutase mimetic and intracellular redox agent that can be a potential drug. This study investigated the regulatory mechanism of Tempol in the treatment of ALI. A mouse model of ALI was established, and HE staining was used to examine histomorphology. The CCK-8 assay was used to measure cell viability, and oxidative stress was assessed by corresponding kits. Flow cytometry and dichlorodihydrofluorescein diacetate staining assays were used to detect reactive oxygen species (ROS) levels. Protein expression levels were measured by Western blot analysis and ELISA. Pulmonary vascular permeability was used to measure the lung wet/dry weight ratio. The level of oxidative stress was increased in ALI mice, and the level of ferroptosis was upregulated. Tempol inhibited this effect and alleviated ALI. The administration of Tempol alleviated the pathological changes in ALI, inhibited pulmonary vascular permeability, and improved lung injury in ALI mice. The upregulation of genes essential for glutathione (GSH) metabolism induced by lipopolysaccharide (LPS) was inhibited by Tempol. In addition, nuclear factor-related factor 2 (Nrf2) is activated by Tempol therapy to regulate the de novo synthesis pathway of GSH, thereby alleviating LPS-induced lung epithelial cell damage. The results showed that Tempol alleviated ALI by activating the Nrf2 pathway to inhibit oxidative stress and ferroptosis in lung epithelial cells. In conclusion, this study demonstrates that Tempol alleviates ALI by inhibiting ferroptosis in lung epithelial cells through the effect of Nrf2 on GSH synthesis.

Annulations of Ynamides with 1,2-Benzisothiazoles to Construct 1,4-Benzothiazepines and 3-Aminoisoquinolines.

Two distinct TMSOTf-catalyzed annulation reactions between ynamides and 1,2-benzisothiazoles have been developed. The direction of the reaction could be easily switched by applying different thermodynamic-controlled conditions. At room temperature, ynamides undergo a [5 + 2] annulation reaction with 1,2-benzisothiazoles to afford 1,4-benzothiazepines, whereas under heating conditions a desulfurizative annulation reaction proceeds well to access 3-aminoisoquinolines. These two protocols provide biologically important 1,4-benzothiazepines and 3-aminoisoquinolines with high efficiency with broad substrate scopes under mild reaction conditions.

Chronic but not acute nicotine treatment ameliorates acute inflammation-induced working memory impairment by increasing CRTC1 and HCN2 in adult male mice.

BACKGROUND: Systemic inflammation in which lipopolysaccharide (LPS) is released into circulation can cause cognitive dysfunction and we have previously shown that LPS impaired working memory (WM) which refers to the ability to guide incoming behavior by retrieving recently acquired information. However, the mechanism is not very clear, and currently, there is no approved strategy to improve inflammation-induced WM deficit. Notably, epidemiological studies have demonstrated a lower occurrence rate of inflammatory-related diseases in smoking patients, suggesting that inflammation-induced WM impairment may be improved by nicotine treatment. Here, our object is to investigate the effect and potential mechanisms of acute and chronic nicotine treatment on LPS-produced WM deficiency. METHODS: Delayed alternation T-maze task (DAT) was applied for evaluating WM which includes both the short-term information storage and the ability to correct errors in adult male mice. Immunofluorescence staining and immunoblotting were used for assessing the levels and distribution of CREB-regulated transcription coactivator 1 (CRTC1) and hyperpolarization-activated cation channels 2 (HCN2) in the medial prefrontal cortex (mPFC) and hippocampus. Quantitative PCR and ELISA were employed for analyzing the mRNA and protein levels of TNF-α and IL-1ß. RESULTS: Our results revealed that administration of LPS (i.p.) at a dose of 0.5 mg/kg significantly produced WM impairment in the DAT task accompanied by an increase in IL-1ß and TNF-α expression in the mPFC. Moreover, intra-mPFC infusion of IL-1Ra, an IL-1 antagonist, markedly alleviated LPS-induced WM deficiency. More important, chronic (2 weeks) but not acute nicotine (0.2 mg/kg, subcutaneous) treatment significantly alleviated LPS-induced WM deficiency by upregulating CRTC1 and HCN2. Of note, intra-mPFC infusion of HCN blocker ZD7288 produced significant WM deficiency. CONCLUSIONS: In summary, in this study, we show that chronic nicotine treatment ameliorates acute inflammation-induced working memory deficiency by increasing CRTC1 and HCN2 in adult male mice.

Restoration of nNOS Expression Rescues Autistic-Like Phenotypes Through Normalization of AMPA Receptor-Mediated Neurotransmission.

Autism spectrum disorder (ASD) is associated with a range of abnormalities characterized by deficits in socialization, communication, repetitive behaviors, and restricted interests. We have recently shown that neuronal nitric oxide synthase (nNOS) expression was decreased in the basolateral amygdala (BLA) of mice after postnatal valproic acid exposure. Neuronal activity-regulated pentraxin (Narp) could contribute to the regulation of the GluA4 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl) propanoic acid (AMPA) subunits which are predominantly expressed in interneurons. However, the specific role of nNOS re-expression on excitatory neurotransmitter with relevance to ASD core symptoms in VPA-treated animals remains to be elucidated. Herein, nNOS overexpression using a lentiviral vector and L-arginine-activating PI3K-Akt-mTOR signaling can restore nNOS expression in the BLA induced by VPA. Restoration of nNOS expression in these mice was sufficient to reduce the severity of ASD-like behavioral patterns such that animals exhibited decreases in abnormal social interactions and communication, stereotyped/repetitive behaviors, and anxiety-like traits. Most strikingly, re-expression of nNOS upregulated surface expression of Narp and GluA4 in nNOS-positive interneuron as shown by immunoprecipitation and Western blotting. Whole-cell patch-clamp recordings demonstrated that restoration of nNOS had a significant enhancing effect on AMPA receptor-mediated excitatory glutamatergic synaptic neurotransmission, which was inhibited by disturbing the interaction between Narp and GluA4 in acutely dissociated BLA slices. Overall, these data offer a scientific basis for the additional study of nNOS re-expression as a promising therapeutic target by correcting AMPA receptor-mediated synaptic function in ASD and related neurodevelopmental disorders.