We used the Exploris 240 mass spectrometer for non-targeted metabolomics on Saccharomyces cerevisiae strain BY4741 and tested AcquireX software for increasing the number of detectable compounds and ...Compound Discoverer 3.3 software for identifying compounds by MS
spectral library matching. AcquireX increased the number of potentially identifiable compounds by 50% through six iterations of MS
acquisition. On the basis of high-scoring MS
matches made by Compound Discoverer, there were 483 compounds putatively identified from nearly 8000 candidate spectra. Comparisons to 20 amino acid standards, however, revealed instances whereby compound matches could be incorrect despite strong scores. Situations included the candidate with the top score not being the correct compound, matching the same compound at two different chromatographic peaks, assigning the highest score to a library compound much heavier than the mass for the parent ion, and grouping MS
isomers to a single parent ion. Because the software does not calculate false positive and false discovery rates at these multiple levels where such errors can propagate, we conclude that manual examination of findings will be required post software analysis. These results will interest scientists who may use this platform for metabolomics research in diverse disciplines including medical science, environmental science, and agriculture.
An enhanced NADH/NAD
ratio, termed reductive stress, is associated with many diseases. However, whether a downstream sensing pathway exists to mediate pathogenic outcomes remains unclear. Here, we ...generate a soluble pyridine nucleotide transhydrogenase from Escherichia coli (EcSTH), which can elevate the NADH/NAD
ratio and meantime reduce the NADPH/NADP
ratio. Additionally, we fuse EcSTH with previously described LbNOX (a water-forming NADH oxidase from Lactobacillus brevis) to resume the NADH/NAD
ratio. With these tools and by using genome-wide CRISPR/Cas9 library screens and metabolic profiling in mammalian cells, we find that accumulated NADH deregulates PRPS2 (Ribose-phosphate pyrophosphokinase 2)-mediated downstream purine biosynthesis to provoke massive energy consumption, and therefore, the induction of energy stress. Blocking purine biosynthesis prevents NADH accumulation-associated cell death in vitro and tissue injury in vivo. These results underscore the pathophysiological role of deregulated purine biosynthesis in NADH accumulation-associated disorders and demonstrate the utility of EcSTH in manipulating NADH/NAD
and NADPH/NADP
.
The Chinese state has recently implemented the COVID-19 Vaccine Communication Campaign (CVCC) to counter vaccine hesitancy. Nonetheless, the extant literature that examines COVID-19 vaccine ...acceptance has less represented COVID-19 vaccine communication efforts.
To address this lacuna, we qualitatively explored how CVCCs were organized in Chinese communities by investigating 54 Chinese stakeholders.
This study indicates that the CVCC was sustained by top-down political pressure. CVCCs' components involve ideological education among politically affiliated health workers, expanding health worker networks, training health workers, implementing media promotion, communicating with residents using persuasive and explanatory techniques, encouraging multistakeholder partnerships, and using public opinion-steered and coercive approaches. While CVCCs significantly enhanced COVID-19 vaccine acceptance, lacking open communication, stigmatizing vaccine refusers, insufficient stakeholder collaboration, and low trust in the COVID-19 vaccination program (CVP) eroded the validity of CVCCs.
To promote the continuity of CVCCs in China, CVCC performers are expected to conduct open and inclusive communication with residents. Furthermore, CVP planers should create robust partnerships among health workers by ensuring their agreements on strategies for implementing CVCCs and optimize COVID-19 immunization service provision to depoliticize CVPs. Our study will not only deepen global audiences' understanding of CVCCs in authoritarian China but also offer potential neighborhood-level solutions for implementing local and global public health communication efforts.
A root-knot nematode gene (MiIDL1) produces a peptide that mimics the Arabidopsis IDA signaling peptide that regulates cell separation events including abscission and lateral root emergence.
Abstract
...INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) is a signaling peptide that regulates cell separation in Arabidopsis including floral organ abscission and lateral root emergence. IDA is highly conserved in dicotyledonous flowering plant genomes. IDA-like sequences were also found in the genomic sequences of root-knot nematodes, Meloidogyne spp., which are globally deleterious pathogens of agriculturally important plants, but the role of these genes is unknown. Exogenous treatment of the Arabidopsis ida mutant with synthetic peptide identical to the M. incognita IDA-like 1 (MiIDL1) protein sequence minus its N-terminal signal peptide recovered both the abscission and root architecture defects. Constitutive expression of the full-length MiIDL1 open reading frame in the ida mutant substantially recovered the delayed floral organ abscission phenotype whereas transformants expressing a construct missing the MiIDL1 signal peptide retained the delayed abscission phenotype. Importantly, wild-type Arabidopsis plants harboring an MiIDL1-RNAi construct and infected with nematodes had approximately 40% fewer galls per root than control plants. Thus, the MiIDL1 gene produces a functional IDA mimic that appears to play a role in successful gall development on Arabidopsis roots.
The methyltransferase like 3 (METTL3) has been generally recognized as a nuclear protein bearing oncogenic properties. We find predominantly cytoplasmic METTL3 expression inversely correlates with ...node metastasis in human cancers. It remains unclear if nuclear METTL3 is functionally distinct from cytosolic METTL3 in driving tumorigenesis and, if any, how tumor cells sense oncogenic insults to coordinate METTL3 functions within these intracellular compartments. Here, we report an acetylation-dependent regulation of METTL3 localization that impacts on metastatic dissemination. We identify an IL-6-dependent positive feedback axis to facilitate nuclear METTL3 functions, eliciting breast cancer metastasis. IL-6, whose mRNA transcript is subjected to METTL3-mediated m
A modification, promotes METTL3 deacetylation and nuclear translocation, thereby inducing global m
A abundance. This deacetylation-mediated nuclear shift of METTL3 can be counterbalanced by SIRT1 inhibition, a process that is further enforced by aspirin treatment, leading to ablated lung metastasis via impaired m
A methylation. Intriguingly, acetylation-mimetic METTL3 mutant reconstitution results in enhanced translation and compromised metastatic potential. Our study identifies an acetylation-dependent regulatory mechanism determining the subcellular localization of METTL3, which may provide mechanistic clues for developing therapeutic strategies to combat breast cancer metastasis.
A series of vaccine incidents have stimulated vaccine hesitance in China over the last decade. Many scholars have studied the institutional management of these incidents, but a qualitative study of ...stakeholders' perspectives on vaccine hesitancy in China is missing. To address this lacuna, we conducted in-depth interviews and collected online data to explore diverse stakeholders' narratives on vaccine hesitance. Our analysis shows the different perspectives of medical experts, journalists, parents, and self-defined vaccination victims on vaccination and vaccination hesitance. Medical experts generally consider vaccines, despite some flaws, as safe, and they consider most vaccine safety incidents to be related to coupling symptoms, not to vaccinations. Some parents agree with medical experts, but most do not trust vaccine safety and do not want to put their children at risk. Media professionals, online medical experts, and doctors who do not need to align with the political goal of maintaining a high vaccination rate are less positive about vaccination and consider vaccine hesitance a failure of expert-lay communication in China. Our analysis exhibits the tensions of medical expert and lay perspectives on vaccine hesitance, and suggests that vaccination experts 'see like a state', which is a finding consistent with other studies that have identified the over-politicization of expert-lay communication in Chinese public discourse. Chinese parents need space to express their concerns so that vaccination programs can attune to them.
Abscission zone (AZ) development and the progression of abscission (detachment of plant organs) have been roughly separated into four stages: first, AZ differentiation; second, competence to respond ...to abscission signals; third, activation of abscission; and fourth, formation of a protective layer and post-abscission trans-differentiation. Stage three, activation of abscission, is when changes in the cell wall and extracellular matrix occur to support successful organ separation. Most abscission research has focused on gene expression for enzymes that disassemble the cell wall within the AZ and changes in phytohormones and other signaling events that regulate their expression. Here, transcriptome data for soybean, tomato and Arabidopsis were examined and compared with a focus not only on genes associated with disassembly of the cell wall but also on gene expression linked to the biosynthesis of a new extracellular matrix. AZ-specific up-regulation of genes associated with cell wall disassembly including cellulases (beta-1,4-endoglucanases, CELs), polygalacturonases (PGs), and expansins (EXPs) were much as expected; however, curiously, changes in expression of xyloglucan endotransglucosylase/hydrolases (XTHs) were not AZ-specific in soybean. Unexpectedly, we identified an early increase in the expression of genes underlying the synthesis of a waxy-like cuticle. Based on the expression data, we propose that the early up-regulation of an abundance of small pathogenesis-related (PR) genes is more closely linked to structural changes in the extracellular matrix of separating cells than an enzymatic role in pathogen resistance. Furthermore, these observations led us to propose that, in addition to cell wall loosening enzymes, abscission requires (or is enhanced by) biosynthesis and secretion of small proteins (15-25 kDa) and waxes that form an extensible extracellular matrix and boundary layer on the surface of separating cells. The synthesis of the boundary layer precedes what is typically associated with the post-abscission synthesis of a protective scar over the fracture plane. This modification in the abscission model is discussed in regard to how it influences our interpretation of the role of multiple abscission signals.
Epithelial-to-mesenchymal transition (EMT) activation is important in cancer progression and metastasis. Evidence indicates that nc886 is a representative Pol III gene that processes microRNA ...products via Dicer and further downregulates its target gene transforming growth factor- β1 (TGF-β1), which is the most prominent inducer of EMT in prostate cancer (PC). Consistent with the previous literature, we found that nc886 downregulation was strongly associated with metastatic behavior and showed worse outcomes in PC patients. However, little is known about the association between nc886 and the EMT signaling pathway. We developed a PC cell model with stable overexpression of nc886 and found that nc886 changed cellular morphology and drove MET. The underlying mechanism may be related to its promotion of SNAIL protein degradation via ubiquitination, but not to its neighboring genes, TGFβ-induced protein (TGFBI) and SMAD5, which are Pol II-transcribed. TGF-β1 also override nc886 promotion of MET via transient suppression the transcription of nc886, promotion of TGFBI or increase in SMAD5 phosphorylation. Both nc886 inhibition and TGFBI activation occur regardless of their methylation status. The literature suggests that MYC inhibition by TGF-β1 is attributed to nc886 downregulation. We incidentally identified MYC-associated zinc finger protein (MAZ) as a suppressive transcription factor of TGFBI, which is controlled by TGF-β1. We elucidate a new mechanism of TGF-β1 differential control of Pol II and the transcription of its neighboring Pol III gene and identify a new EMT unit consisting of nc886 and its neighboring genes.
Multiple myeloma (MM) remains incurable by conventional chemotherapy. Sphingosine-1-phosphate (S1P) receptor-mediated signaling has been recently demonstrated to have critical roles in cell survival ...and drug resistance in a number of hematological malignancies. To dissect the roles of S1P receptor pathway in MM, we systematically examined cell viability and protein expression associated with cell survival and drug resistance in MM cell lines upon treatment with either pathway activator (S1P) or inhibitor (FTY720). Our results reveal that FTY720 inhibits cell proliferation by downregulating expression of target genes, while S1P has an opposite effect. Knocking down of S1P receptor S1P5R results in a reduction of cell survival-related gene expression; however, it does not have impacts on expression of drug resistance genes. These results suggest that S1P signaling plays a role in cell proliferation and drug resistance in MM, and targeting this pathway will provide a new therapeutic direction for MM management.
B-cell antigen receptor (BCR) signaling is required to maintain the physiological functions of normal B cells and plays an important pathogenic role in B-cell malignancies. Bruton tyrosine kinase ...(BTK), a critical mediator of BCR signaling, is an attractive target for the treatment of B-cell malignancies. This study aimed to identify a highly potent and selective BTK inhibitor.
Homogeneous time-resolved fluorescence assays were used to screen BTK inhibitors. Typhoon fluorescence imaging and Western blot analysis were used to confirm the effects of SY-1530 on the BCR signaling pathway. Additionally, the anti-tumor activities of SY-1530 were evaluated in TMD8 xenografts and spontaneous canine B-cell lymphoma.
We found a novel irreversible and non-competitive inhibitor of BTK, SY-1530, which provided dose-dependent and time-dependent inhibition. SY-1530 selectively bound to BTK rather than inducible T-cell kinase; consequently, it did not significantly affect T-cell receptor signaling and caused limited off-target effects. SY-1530 blocked the BCR signaling pathway through down-regulation of BTK activity, thus leading to impaired phosphorylation of BTK and its downstream kinases. Moreover, SY-1530 induced apoptosis in a caspase-dependent manner and efficaciously inhibited tumor growth in mouse xenograft models of B-cell malignancy (
< 0.001). SY-1530 also induced positive clinical responses in spontaneous canine B-cell lymphoma.
SY-1530 is an irreversible and selective BTK inhibitor that shows inhibitory effects on B-cell malignancies by blocking the BCR signaling pathway. Therefore, it may be a promising therapeutic approach for the treatment of B-cell malignancies.
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