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Abstract RNA-binding proteins (RBPs) modulate all aspects of RNA metabolism, but a comprehensive picture of RBP expression across tissues is lacking. Here, we describe our development of the method we call HARD-AP that robustly retrieves RBPs and tightly associated RNA regulatory complexes from cultured cells and fresh tissues. We successfully use HARD-AP to establish a comprehensive atlas of RBPs across mouse primary organs. We then systematically map RNA-binding sites of these RBPs using machine learning-based modeling. Notably, the modeling reveals that the LIM domain as an RNA-binding domain in many RBPs. We validate the LIM-domain-only protein Csrp1 as a tissue-dependent RNA binding protein. Taken together, HARD-AP is a powerful approach that can be used to identify RBPomes from any type of sample, allowing comprehensive and physiologically relevant networks of RNA-protein interactions.

Background/Purpose: Congenital nephrotic syndrome (CNS) is one of the important causes of end-stage kidney disease in children. Studies on the genotype, phenotype, and clinical outcome in infants with CNS caused by genetic mutations are scarce. Methods: We analyzed the genetic background, clinical manifestations, treatment response, and prognosis of pediatric patients with CNS in Taiwan. Results: Fifteen infants with CNS were enrolled, and 11 patients of median age 21 (interquartile range 3∼44) days caused by genetic mutations from 10 unrelated families were included in the study. Of the eleven patients, 9 had extra-renal manifestations including microcephaly, facial dysmorphism, and skeletal anomalies. More than two-thirds of the patients had disease onset before 1 month of age. Diffuse meningeal sclerosis was the most common histological characteristic. Whole exome sequencing followed by direct Sanger sequence revealed mutations in OSGEP (R247Q), WT1 (R366H and R467Q), LAMB2 (Q1209∗ and c. 5432–5451 19 bp deletion), NUP93 (D302V), and LAGE3 (c.188+1G > A). Three of the variants were novel. Corticosteroids and/or immunosuppressants were administered in 2 patients, but both were refractory to treatment. During the mean 3.5 years of follow-up, all but two died of uremia and sepsis. The two survivors reached end-stage kidney disease and required peritoneal dialysis, and one of them underwent uneventful renal transplantation. Conclusion: The majority of patients with CNS in Taiwan were caused by OSGEP followed by WT1 mutation. R247Q is the hotspot mutation of OSGEP in Taiwan. CNS patients in Taiwan suffer from significant morbidity and mortality.

Malachite green (MG), a triphenylmethane dye, is used in the aquaculture industry as a disinfectant and insect repellent due to its potent bactericidal and pesticidal properties. However, its use poses potential environmental and health risks. This study analyzed and designed two haptens using computer simulation. Serum data confirmed the feasibility of introducing an arm at the dimethylamine group. Subsequently, a highly selective monoclonal antibody strain was successfully prepared based on the hapten. After optimizing the working conditions of indirect competitive enzyme-linked immunosorbent assay (IC-ELISA), the IC50 value was 0.83 ng/mL, with a detection limit (IC10) of 0.08 ng/mL and a linear range of 0.19–3.52 ng/mL. The developed monoclonal antibody exhibited a crossover rate of less than 0.1% with other similar structures and can be used to establish an immunoassay.

Abstract Direct conversion of inexpensive biomass into value-added chemicals via furanic platform molecules is highly attractive. In this work, we present a straightforward chemobiocatalytic route for glucose valorization into 2,5-bis(hydroxymethyl)furan (BHMF) in one pot, with no purification of the intermediate 5-hydroxymethylfurfural (HMF). Six candidate alcohol dehydrogenase (ADH) genes were located from Meyerozyma guilliermondii SC1103, based on comparative transcriptome analysis and real-time quantitative polymerase chain reaction. An ADH (MgADH1) was identified upon evaluation of catalytic performances of recombinant Saccharomyces cerevisiae harboring candidate ADHs in HMF reduction. Soluble expression of the enzyme in S. cerevisiae was greatly enhanced by its codon optimization, leading to improved HMF tolerance (up to 400 mM). In a fed-batch process, the desired product of approximately 473 mM (60.5 g/L) was produced within 30 h by recombinant S. cerevisiae_MgADH1. A chemobiocatalytic route toward BHMF was constructed by merging CaCl2-mediated isomerization and dehydration with biocatalytic reduction with an overall yield of approximately 42%, starting from glucose. This work may pave the way for green manufacture of valuable biobased chemicals.

Abstract RNA-binding proteins (RBPs)-RNA networks have contributed to cancer development. Circular RNAs (circRNAs) are considered as protein recruiters; nevertheless, the patterns of circRNA-protein interactions in colorectal cancer (CRC) are still lacking. Processing bodies (PBs) formed through liquid-liquid phase separation (LLPS) are membrane-less organelles (MLOs) consisting of RBPs and RNA. Previous evidence suggests a connection between PBs dynamics and cancer progression. Despite the increasingly acknowledged crucial role of RBPs and RNA in the accumulation and maintenance of MLOs, there remains a lack of specific research on the interactions between PBs-related RBPs and circRNAs in CRC. Herein, we identify that MEX-3 RNA binding family member A (MEX3A), frequently upregulated in CRC tissues, predicts poorer patient survival. Elevated MEX3A accelerates malignance and inhibits autophagy of CRC cells. Importantly, MEX3A undergoes intrinsically disordered regions (IDRs)-dependent LLPS in the cytoplasm. Specifically, circMPP6 acts as a scaffold to facilitate the interaction between MEX3A and PBs proteins. The MEX3A/circMPP6 complex modulates PBs dynamic and promotes UPF-mediated phosphodiesterase 5A (PDE5A) mRNA degradation, consequently leading to the aggressive properties of CRC cells. Clinically, CRC patients exhibiting high MEX3A expression and low PDE5A expression have the poorest overall survival. Our findings reveal a collaboration between MEX3A and circMPP6 in the regulation of mRNA decay through triggering the PBs aggregation, which provides prognostic markers and/or therapeutic targets for CRC.

Abstract Improving drug sensitivity is an important strategy in chemotherapy of cancer and accumulating evidence indicates that miRNAs are involved in the regulation of drug sensitivity, but the specific mechanism is still unclear. Our previous study has found that miR-296-5p was significantly downregulated in nasopharyngeal carcinoma (NPC). Here, we aim to explore whether miR-296-5p is involved in regulating cisplatin sensitivity in NPC by regulating STAT3/KLF4 signaling axis. The cell proliferation and clonogenic capacity of NPC cells were evaluated by CCK8 Assay and plate colony assay, respectively. The Annexin V-FITC staining kit was used to determine and quantify the apoptotic cells using flow cytometry. The drug efflux ability of NPC cells were determined by Rhodamine 123 efflux experiment. The expression of miR-296-5p, apoptosis-related genes and protein in NPC cell lines were detected by qPCR and Western blot, respectively. Animal study was used to evaluate the sensitivity of NPC cells to DDP treatment in vivo. Our results showed that elevated miR-296-5p expression obviously promoted the sensitivity of NPC cells to DDP by inhibiting cell proliferation and clonogenic capacity, and inducing apoptosis. In addition, we found that miR-296-5p inhibited the expression of STAT3 and KLF4 in NPC cells, while overexpression of exogenous STAT3 reversed miR-296-5p-mediated enhancement in cell death of DDP-treated NPC cells. In vivo studies further confirmed that miR-296-5p promotes the sensitivity of NPC cells to DDP treatment. miRNA-296-5p enhances the drug sensitivity of nasopharyngeal carcinoma cells to cisplatin via STAT3/KLF4 signaling pathway.

BackgroundViral tracers are important tools for mapping brain connectomes. The feature of predominant anterograde transneuronal transmission offers herpes simplex virus-1 (HSV-1) strain H129 (HSV1-H129) as a promising candidate to be developed as anterograde viral tracers. In our earlier studies, we developed H129-derived anterograde polysynaptic tracers and TK deficient (H129-dTK) monosynaptic tracers. However, their broad application is limited by some intrinsic drawbacks of the H129-dTK tracers, such as low labeling intensity due to TK deficiency and potential retrograde labeling caused by axon terminal invasion. The glycoprotein K (gK) of HSV-1 plays important roles in virus entry, egress, and virus-induced cell fusion. Its deficiency severely disables virus egress and spread, while only slightly limits viral genome replication and expression of viral proteins. Therefore, we created a novel H129-derived anterograde monosynaptic tracer (H129-dgK) by targeting gK, which overcomes the limitations of H129-dTK.MethodsUsing our established platform and pipeline for developing viral tracers, we generated a novel tracer by deleting the gK gene from the H129-G4. The gK-deleted virus (H129-dgK-G4) was reconstituted and propagated in the Vero cell expressing wildtype H129 gK (gKwt) or the mutant gK (gKmut, A40V, C82S, M223I, L224V, V309M), respectively. Then the obtained viral tracers of gKmut pseudotyped and gKwt coated H129-dgK-G4 were tested in vitro and in vivo to characterize their tracing properties.ResultsH129-dgK-G4 expresses high levels of fluorescent proteins, eliminating the requirement of immunostaining for imaging detection. Compared to the TK deficient monosynaptic tracer H129-dTK-G4, H129-dgK-G4 labeled neurons with 1.76-fold stronger fluorescence intensity, and visualized 2.00-fold more postsynaptic neurons in the downstream brain regions. gKmut pseudotyping leads to a 77% decrease in retrograde labeling by reducing axon terminal invasion, and thus dramatically improves the anterograde-specific tracing of H129-dgK-G4. In addition, assisted by the AAV helper trans-complementarily expressing gKwt, H129-dgK-G4 allows for mapping monosynaptic connections and quantifying the circuit connectivity difference in the Alzheimer's disease and control mouse brains.ConclusionsgKmut pseudotyped H129-dgK-G4, a novel anterograde monosynaptic tracer, overcomes the limitations of H129-dTK tracers, and demonstrates desirable features of strong labeling intensity, high tracing efficiency, and improved anterograde specificity.

IntroductionThis study uses survey data from 504 Chinese teachers to investigate the relationship between social media use for work during non-work hours (SMUNW) and turnover intention, focusing on the mediating role of burnout and the moderating role of resilience in this relationship.MethodsIn November 2023, online survey links were sent to 529 teachers from Shandong Province, China, asking them to report their perceptions of SMUNW, burnout, resilience, and turnover intention. A sample of 504 valid responses was obtained. The analysis was performed using SPSS 26.0 and Hayes’ PROCESS MACRO for SPSS for testing the hypotheses.ResultsThe results revealed that burnout acts as a mediator in the relationship between SMUNW and turnover intention. In addition, resilience moderated the relationship between SMUNW and burnout such that when resilience was higher, the relationship between SMUNW and burnout was alleviated. Moreover, resilience moderated the indirect relationship between SMUNW and turnover intention through burnout such that when resilience was higher, the indirect relationship was alleviated.DiscussionThe results of this study indicate that SMUNW is related to turnover intention through the mediating role of burnout. Furthermore, resilience moderates the influence of SMUNW on burnout and thus weakens the influence of SMUNW on turnover intention through burnout. This study expands our knowledge of the nuanced influence mechanisms of social media use in the context of increasing technostress among public employees. Practically, it suggests that managers should pay due attention to the impairments brought about by social media use, especially from the perspectives of preventing burnout and fostering resilience.

Considering the limited efficacy of current therapies in lung, colorectal, and pancreatic cancers, innovative combination treatments with diverse mechanisms of action are needed to improve patients’ outcomes. Chitinase-3 like-1 protein (CHI3L1) emerges as a versatile factor with significant implications in various diseases, particularly cancers, fostering an immunosuppressive tumor microenvironment for cancer progression. Therefore, pre-clinical validation is imperative to fully realize its potential in cancer treatment. We developed phage display-derived fully human monoclonal CHI3L1 neutralizing antibodies (nAbs) and verified the nAbs-antigen binding affinity and specificity in lung, pancreatic and colorectal cancer cell lines. Tumor growth signals, proliferation and migration ability were all reduced by CHI3L1 nAbs in vitro. Orthotopic or subcutaneous tumor mice model and humanized mouse model were established for characterizing the anti-tumor properties of two CHI3L1 nAb leads. Importantly, CHI3L1 nAbs not only inhibited tumor growth but also mitigated fibrosis, angiogenesis, and restored immunostimulatory functions of immune cells in pancreatic, lung, and colorectal tumor mice models. Mechanistically, CHI3L1 nAbs directly suppressed the activation of pancreatic stellate cells and the transformation of macrophages into myofibroblasts, thereby attenuating fibrosis. These findings strongly support the therapeutic potential of CHI3L1 nAbs in overcoming clinical challenges, including the failure of gemcitabine in pancreatic cancer.

Energy recovery from low-strength wastewater through anaerobic methanogenesis is constrained by limited substrate availability. The development of efficient methanogenic communities is critical but challenging. Here we develop a strategy to acclimate methanogenic communities using conductive carrier (CC), electrical stress (ES), and Acid Orange 7 (AO7) in a modified biofilter. The synergistic integration of CC, ES, and AO7 precipitated a remarkable 72-fold surge in methane production rate compared to the baseline. This increase was attributed to an altered methanogenic community function, independent of the continuous presence of AO7 and ES. AO7 acted as an external electron acceptor, accelerating acetogenesis from fermentation intermediates, restructuring the bacterial community, and enriching electroactive bacteria (EAB). Meanwhile, CC and ES orchestrated the assembly of the archaeal community and promoted electrotrophic methanogens, enhancing acetotrophic methanogenesis electron flow via a mechanism distinct from direct electrochemical interactions. The collective application of CC, ES, and AO7 effectively mitigated electron flow impediments in low-strength wastewater methanogenesis, achieving an additional 34% electron recovery from the substrate. This study proposes a new method of amending anaerobic digestion systems with conductive materials to advance wastewater treatment, sustainability, and energy self-sufficiency.

Augmented Reality (AR) is increasingly recognized as a transformative tool for creating restorative environments within museums. It has the potential to provide psychological benefits for visitors, including attention restoration, stress reduction, and anxiety alleviation. This study explores how AR can foster these benefits within museum spaces. By adopting AR technology, museums can go beyond their traditional roles of knowledge dissemination. The immersive, adaptive, and interactive features of AR can enhance the museum experience, transforming it into an innovative therapeutic space. By combining real exhibits with virtual elements, AR can restore visitors’ psychological energy within museum settings. This integration of digital innovation into restorative contexts surpasses the traditional functions of visual service. Through empirical investigation of multiple dimensions of restorative environments, AR museum experiences offer comprehensive attention restoration. In this study, a survey was conducted with 279 participants to assess the impact of AR museum experiences on visitors’ psychology. The results revealed that such experiences contribute to heightened attention restoration levels, stress reduction, and anxiety relief. With the latest advancements in generative artificial intelligence, AR technology is empowered to integrate within museums. This integration will merge individuals with customized technology, expanding human perceptual experiences and highlighting AR's significant influence within the museum environment.

Objective: This study aimed to describe real-world patient and physician characteristics, rearranged during transfection (RET) mutation testing and results, treatment patterns and patient-reported outcomes (PROs) in advanced or metastatic medullary thyroid cancer (aMTC) across five populous European countries. Methods: Cross-sectional physician and patient surveys were used to collect quantitative and qualitative data in France, Germany, Italy, Spain and the UK from July to December 2020, prior to the introduction of selective RET inhibitors in Europe. Physicians completed patient record forms and a survey about their specialty and practice site. Patients were asked to provide PRO data using four validated instruments, including the EuroQol 5 Dimension (EQ-5D) questionnaire. Results: The physician-reported sample included 275 patients with aMTC, including 79 patients with RET mutation-positive disease; median age was 60 and 56 years, respectively. Overall, 75% were tested for RET mutation (35% germline only, 21% somatic only and 44% both). Common physician-cited barriers to RET mutation testing included high cost, difficulty accessing the latest tests and time delay for results. First-line systemic therapy (most commonly vandetanib or cabozantinib) was prescribed for 69% of patients overall and 82% of the RET mutation-positive subgroup. Second-line therapy was prescribed for 12% of patients who received first-line therapy; most patients remained on first-line therapy at data capture. PROs revealed a substantial disease/treatment burden. Conclusion: Patients with aMTC report a substantial disease/treatment burden. Outcomes could be improved by identifying patients eligible for treatment with selective RET inhibitors through more optimal RET mutation testing.

With the better understanding of human genome and genetic contributions to human complex diseases, more and more clinical genetic tests have been developed for the prevention, diagnosis, and treatment of human diseases. This has raised new challenges to classroom teaching of medical genetics (MG) in medical education of undergraduates, who are relatively naive about the theory and technology of MG. A clear understanding of genotype-phenotype relationships is critical in MG learning. In the current study, we incorporated a practical approach of classroom MG teaching by using alcohol tolerance, a phenotype that is widely experienced by students and closely related to genetic polymorphism. To verify this teaching approach, which we named as student experience-based teaching (SEBT), undergraduates of two classes were recruited, with one accepting SEBT and the other accepting conventional lecture-based teaching (LBT). Final examination of the MG course indicated that while the two classes showed similar results to multiple-choice questions, the SEBT class showed significantly better performance to short-answer questions about gene polymorphism and phenotype, suggesting that SEBT might improve understanding. Thus, using student experience-based phenotypic instances and explaining the phenotype by practical genetic tests could improve understanding of genotype-phenotype relationships in classroom MG teaching.

Alport syndrome (AS) is a progressive hereditary kidney disease characterized by hematuria, proteinuria, and progressive kidney dysfunction accompanied by sensorineural hearing loss and ocular abnormalities. Pathogenic COL4A3-5 variants can result in different AS spectra. Further, kidney cysts have been reported in adults with AS. However, the relationship between kidney cysts and AS remains unclear. Here, we report 3 cases of AS in children that occurred with kidney cysts. The patient in case 1 was initially diagnosed with IgA nephropathy at the age of 8 years but later developed bilateral multiple kidney cysts at the age of 17 years, suggesting autosomal-dominant polycystic kidney disease. Whole-exome sequencing identified a pathogenic COL4A5 variant and confirmed the AS diagnosis. The patients in cases 2 and 3 had already been diagnosed with X-linked AS using kidney biopsy and genetic analysis. Initial kidney ultrasonography showed nephromegaly; however, kidney cyst formation was observed during their annual follow-up. Our study supports the association between AS and kidney cysts. Kidney cysts in adolescents with suspected AS should not discourage clinicians from testing for pathogenic COL4A3-COL4A5 variants. Early detection of kidney cysts is critical because it may indicate kidney disease progression.

Kaposi's sarcoma-associated herpesvirus (KSHV) is a double-stranded DNA virus etiologically associated with multiple malignancies. Both latency and sporadic lytic reactivation contribute to KSHV-associated malignancies, however, the specific roles of many KSHV lytic gene products in KSHV replication remain elusive. In this study, we report that ablation of ORF55, a late gene encoding a tegument protein, does not impact KSHV lytic reactivation but significantly reduces the production of progeny virions. We found that cysteine 10 and 11 (C10 and C11) of pORF55 are palmitoylated, and the palmytoilation is essential for its Golgi localization and secondary envelope formation. Palmitoylation-defective pORF55 mutants are unstable and undergo proteasomal degradation. Notably, introduction of a putative Golgi localization sequence to these palmitoylation-defective pORF55 mutants restores Golgi localization and fully reinstates KSHV progeny virion production. Together, our study provides new insight into the critical role of pORF55 palmitoylation in KSHV progeny virion production and offers potential therapeutic targets for the treatment of related malignancies.

Monosynaptic viral tracers are essential tools for dissecting neuronal connectomes and for targeted delivery of molecular sensors and effectors. Viral toxicity and complex multi-injection protocols are major limiting application barriers. To overcome these barriers, we developed an anterograde monosynaptic H129Amp tracer system based on HSV-1 strain H129. The H129Amp tracer system consists of two components: an H129-dTK-T2-pacFlox helper which assists H129Amp tracer's propagation and transneuronal monosynaptic transmission. The shared viral features of tracer/helper allow for simultaneous single-injection and subsequent high expression efficiency from multiple-copy of expression cassettes in H129Amp tracer. These improvements of H129Amp tracer system shorten experiment duration from 28-day to 5-day for fast-bright monosynaptic tracing. The lack of toxic viral genes in the H129Amp tracer minimizes toxicity in postsynaptic neurons, thus offering the potential for functional anterograde mapping and long-term tracer delivery of genetic payloads. The H129Amp tracer system is a powerful tracing tool for revealing neuronal connectomes.

The volatile organic compounds of six spices, including black pepper, dried ginger, cinnamon, fennel, clove, and zanthoxylum, were analyzed by gas chromatography–ion mobility spectrometry (HS-GC-IMS) combined with principal component analysis (PCA) and Euclidean distance. In further analyses, the effects of volatile oils in six spices on ulcerative colitis were assayed in a zebrafish model induced by 3-nitrobenzenesulfonic acid. A total of 120 kinds of volatile organic compounds were detected and 80 among them were identified, which included 10 common components and 3 to 24 characteristic components belonging to different spices. The major VOCs in six spices were estimated to be terpenes with the contents of 45.02%, 56.87%, 36.68%, 58.19%, 68.68%, and 30.62%, respectively. Meanwhile, the volatile components of fennel, dried ginger, black pepper, and cinnamon are quite similar, but differ from clove and zanthoxylum. The volatile oils in six spices presented efficient activity to improve ulcerative colitis which can decrease the number of neutrophils, restore the structure of intestinal epithelial and the morphology of the epithelial cells. Our study achieved rapid analysis of the volatile organic compounds and flavors in six spices and further revealed the potential health benefits of their volatile oils on ulcerative colitis, especially for clove and zanthoxylum. This study is expected to provide certain data support for the quality evaluation and the potential use in functional foods of six spices.