Nucleoporin Nup98 is an essential factor for ipo4 dependent protein import.

Nucleocytoplasmic transport of macromolecules is essential in eukaryotic cells. In this process, the karyopherins play a central role when they transport cargoes across the nuclear pore complex. Importin 4 belongs to the karyopherin ß family. Many studies have focused on finding substrates for importin 4, but no direct mechanism studies of its precise transport function have been reported. Therefore, this paper mainly aimed to study the mechanism of nucleoporins in mediating nuclear import and export of importin 4. To address this question, we constructed shRNAs targeting Nup358, Nup153, Nup98, and Nup50. We found that depletion of Nup98 resulted in a shift in the subcellular localization of importin 4 from the cytoplasm to the nucleus. Mutational analysis demonstrated that Nup98 physically and functionally interacts with importin 4 through its N-terminal phenylalanine-glycine (FG) repeat region. Mutation of nine of these FG motifs to SG motifs significantly attenuated the binding of Nup98 to importin 4, and we further confirmed the essential role of the six FG motifs in amino acids 121-360 of Nup98 in binding with importin 4. In vitro transport assay also confirmed that VDR, the substrate of importin 4, could not be transported into the nucleus after Nup98 knockdown. Overall, our results showed that Nup98 is required for efficient importin 4-mediated transport. This is the first study to reveal the mechanism of importin 4 in transporting substrates into the nucleus.

Metabolomic insights into pulmonary fibrosis: a mendelian randomization study.

BACKGROUND: This study leverages a two-sample Mendelian Randomization (MR) approach to explore the causal relationships between 1,400 metabolites and pulmonary fibrosis, using genetic variation as instrumental variables. By adhering to stringent criteria for instrumental variable selection, the research aims to uncover metabolic pathways that may influence the risk and progression of pulmonary fibrosis, providing insights into potential therapeutic targets. METHODS: Utilizing data from the OpenGWAS project, which includes a significant European cohort, and metabolite GWAS data from the Canadian Longitudinal Aging Study (CLSA), the study employs advanced statistical methods. These include inverse variance weighting (IVW), weighted median estimations, and comprehensive sensitivity analyses conducted using the R software environment to ensure the robustness of the causal inferences. RESULTS: The study identified 62 metabolites with significant causal relationships with pulmonary fibrosis, highlighting both risk-enhancing and protective metabolic factors. This extensive list of metabolites presents a broad spectrum of potential therapeutic targets and biomarkers for early detection, underscoring the metabolic complexity underlying pulmonary fibrosis. CONCLUSIONS: The findings from this MR study significantly advance our understanding of the metabolic underpinnings of pulmonary fibrosis, suggesting that alterations in specific metabolites could influence the risk and progression of the disease. These insights pave the way for the development of novel diagnostic and therapeutic strategies, emphasizing the potential of metabolic modulation in managing pulmonary fibrosis.

Associations of toxic metals and their mixture with hyperuricemia in Chinese rural older adults.

Previous studies have related single toxic metals (TMs) to hyperuricemia (HUA) among the general population, however, the association of the TM mixture with HUA, especially in older adults, remains poorly understood. We aimed to examine the relationships between individual TMs and their mixture and HUA in Chinese rural older adults. This study consisted of 2075 rural older adults aged 60 years or over. Blood concentrations of aluminum (Al), arsenic (As), barium (Ba), cadmium (Cd), cesium (Cs), gallium (Ga), mercury (Hg), lead (Pb), thallium (Tl), and uranium (U) were detected using inductively coupled plasma mass spectrometry. The associations of single TMs with HUA were assessed using logistic regression and restricted cubic spline (RCS) models, and the association of TM mixture with HUA was explored using the elastic net with environmental risk score (ENET-ERS), quantile g-computation (QGC), and Bayesian kernel machine regression (BKMR) models, respectively. Adjusted logistic regression model showed that Cs (OR = 1.65, 95% CI 1.37-1.99) and Pb (OR = 1.46, 95% CI 1.28-1.67) were positively related to HUA, and RCS model exhibited a positive linear association of Cs and Pb with HUA. ENET-ERS and QGC models quantified a positive correlation between the TM mixture and the odds of HUA, with estimated ORs of 1.15 (95% CI 1.11-1.19) and 1.84 (95% CI 1.37-2.47), respectively, and Cs and Pb had the most weight. BKMR model demonstrated a significant linear association between the TM mixture and increased odds of HUA, with the posterior inclusion probabilities (PIPs) of both Cs and Pb being 1.00. Moreover, we observed a positive interaction between Cs and Pb on HUA. The TM mixture is associated with increased odds of HUA in rural older adults, which may mainly be driven by Cs and Pb. Subsequent studies are warranted to confirm these findings and clarify the mechanisms linking multiple TMs with HUA.

Integrative analysis of omics summary data reveals putative mechanisms linked to different cell populations in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a complex disease involving many interactions at the molecular level, the details of which remain unclear. Here, we demonstrated an analytical paradigm of prioritizing genes and regulatory elements based on GWAS loci at the single-cell levels. Our initial step was to apply TWMR to identify causal genes and causal methylation sites in SLE. Based on the eQTL, LD and mQTL, we calculated the correlation between these genes and methylation sites. Next, we separately used gene expression and DNAm as exposure variables and outcome variables to analyze the regulatory mechanisms. We identified two mediating modes for SLE: 1) transcription mediation model and 2) epigenetic mediation model. Further, using single-cell RNA sequencing data, we revealed the cell subclusters associated with these mechanisms. Our identification of the mechanisms of SLE in different cell populations is of great significance for understanding the heterogeneity of disease in different cell populations.

Spin-dependent directional emission from a quantum dot ensemble embedded in an asymmetric waveguide.

In this study, we examine a photonic wire waveguide embedded with an ensemble of quantum dots (QDs) that directionally emits into the waveguide depending on the spin state of the ensemble. The directional emission is facilitated by the spin-orbit interaction of light. The waveguide has a two-step stair-like cross section and QDs are embedded only in the upper step, such that the circular polarization of emission from the spin-polarized QDs controls the direction of the radiation. We numerically verify that more than 70% of the radiation from the ensemble emitter is toward a specific direction in the waveguide. We also examine a microdisk resonator with a stair-like edge, which supports selective coupling of the QD ensemble radiation into a whispering gallery mode that rotates unidirectionally. Our study provides a foundation for spin-dependent optoelectronic devices.

PKA-site phosphorylation of importin13 regulates its subcellular localization and nuclear transport function.

Importin 13 (IPO13) is a key member of the importin ß superfamily, which can transport cargoes both into and out of the nucleus to contribute to a variety of important cellular processes. IPO13 is known to undergo phosphorylation, but the impact of this on function has not been investigated. Here, we show for the first time that IPO13 is phosphorylated by cAMP-dependent protein kinase A specifically at serine 193. Results from fluorescence recovery after photobleaching and fluorescence loss in photobleaching approaches establish that negative charge at serine 193 through phosphorylation or point mutation both reduces IPO13 nuclear import and increases its nuclear export. Importantly, phosphorylation also appears to enhance cargo interaction on the part of IPO13, with significant impact on localization, as shown for the Pax6 homeobox-containing transcription partner. This is the first report that IPO13 can be phosphorylated at Ser193 and that this modification regulates IPO13 subcellular localization and nucleocytoplasmic transport function, with important implications for IPO13's role in development and other processes.

STK24 expression is modulated by DNA copy number/methylation in lung adenocarcinoma and predicts poor survival.

AIM: To explore the independent prognostic value of STK24 expression in terms of overall survival and recurrence-free survival and the potential mechanisms of its dysregulation in non-small-cell lung adenocarcinoma. PATIENTS & METHODS: Data were from the Cancer Genome Atlas-lung adenocarcinoma. RESULTS: Increased STK24 expression was an independent prognostic indicator of unfavorable overall survival (Hazard ratio: 1.478; 95% CI: 1.149-1.901; p < 0.002) and recurrence-free survival (Hazard ratio: 1.855; 95% CI: 1.399-2.458; p < 0.001). DNA amplification was associated with significantly upregulated STK24 expression. There was a weak negative correlation between STK24 expression and its DNA methylation (Pearson's r = -0.32). CONCLUSION: Aberrant STK24 expression was an independent prognostic indicator in lung adenocarcinoma patients. Its dysregulation was associated with its DNA copy number alteration and methylation.

DNA methylation of a non-CpG island promoter represses NQO1 expression in rat arsenic-transformed lung epithelial cells.

NAD(P)H:quinone oxidoreductase 1 (NQO1), a phase II flavoenzyme that catalyzes reduction reactions to protect cells against electrophiles and oxidants, is involved in tumorigenesis. Altered methylation of the NQO1 gene has been observed and is speculated to result in aberrant NQO1 expression in rat cells undergoing chemical carcinogenesis, although this has not been proven experimentally. In this study, we first investigated the potential epigenetic mechanisms underlying the phenomenon of NQO1 differential expression in individual subclones of rat arsenic-transformed lung epithelial cells (TLECs). NQO1 expression of TLEC subclones with or without 5-aza-2'-deoxycytidine (5-Aza-CdR) treatment was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR), western blot analysis, and real-time PCR. Methylation status of the NQO1 promoter in TLEC subclones was analyzed by bisulfite sequencing. Transcriptional activity of NQO1 promoter in vitro methylated was determined by luciferase assay using a CpG-free luciferase reporter driven by the NQO1 promoter region (-435 to +229). We found that non-CpG island (non-CpGI) within the NQO1 promoter was hyper- or hypo-methylated in TLEC subclones and corresponded to low and high gene expressions, respectively. Following the treatment with 5-Aza-CdR, transcription of the NQO1 gene in the hypermethylated subclones was restored, accompanied by demethylation of the NQO1 promoter. In vitro promoter methylation almost completely silenced reporter activity in TLECs. These results indicate that DNA methylation of the non-CpGI promoter contributes to epigenetic silencing of NQO1 in rat TLECs.

Nuclear import of Nkx2-2 is mediated by multiple pathways.

Nkx2-2 homeoprotein is essential for the development of the central nervous system and pancreas. Although the nuclear localization signals of Nkx2-2 have been identified, the responsible transport receptor is still unknown. Here, we demonstrate that imp α1 not only interacts with Nkx2-2 but also transports it into the nucleus in vitro by acting together with imp ß1. However, the nuclear import of Nkx2-2 in cells was not inhibited in response to knockdown expression of endogenous imp ß1 or over-expression of Bimax2. Furthermore, imp ß1 and imp 13, but not imp 4, directly interact with Nkx2-2 and are capable of transporting Nkx2-2 in an in vitro import assay. By GST pull-down assay, we demonstrate that mutation of NLS1 or NLS2 has no effect on interaction with imp α1 or imp 13, but significantly reduced binding to imp ß1. Thus, the nuclear import of Nkx2-2 is mediated not only by the classical import pathway but also directly by imp ß1 or imp 13.

Near-infrared light-actuated on-demand botanicals release and hyperthermia by an antibiotic-free polysaccharide-based hydrogel dressing for the synergistic treatment of wound infections.

Bacterial infection is a key factor affecting wound healing. Conventional treatments might lead to the widespread emergence of drug-resistant bacteria due to the long-term and excessive use of antibiotics. It is necessary to develop an antibiotic-free method for effective treatment of bacterial wound infections. In this work, we constructed an antibiotic-free polysaccharide-based hydrogel dressing (ATB) with near-infrared light-actuated on-demand botanicals release and hyperthermia for the synergistic treatment of wound infections. The ATB hydrogel dressing was made up of agarose as a support matrix, berberine hydrochloride as the active botanicals and TA-Fe(III) nanoparticles as NIR laser-activated photothermal reagents. The ATB hydrogel dressing showed spatiotemporal botanicals release and excellent photothermal properties with NIR irradiation. With the results of in vitro and in vivo antibacterial experiments, the antibiotic-free ATB hydrogel could synergistically eliminate bacteria and accelerate wound healing. Overall, the near-infrared light-responsive ATB hydrogel could provide a promising antibiotic-free strategy for the treatment of bacterial wound infections.

Human dental pulp stem cells have comparable abilities to umbilical cord mesenchymal stem/stromal cells in regulating inflammation and ameliorating hepatic fibrosis.

Hepatic fibrosis, also called cirrhosis, have wide prevalence worldwide for long yeas. Recently, many treatments for liver cirrhosis made marked progress, especially the umbilical cord-derived mesenchymal stromal cells (UCMSC) therapy. However, limited recourses and potential immune-related issues become the obstacles on UCMSC popularization in clinic. Therefore, we took dental pulp stem cells (DPSCs) into the consideration, since autologous DPSCs can be easily obtained without any ethnic or immune-related issues that heterogenous UCMSCs could encounter. We systematically compared the effects of both cell types and found that DPSCs had similar results to UCMSCs in regulating inflammation and reversing hepatic fibrosis. In our study, co-culturing T cells and PBMSCs showed that DPSCs have the ability to inhibit the proliferation of inflammatory cells and downregulate relevant inflammatory factors. In vitro and in vivo sterility tests confirmed the bio-safety of DPSCs. Moreover, the 1 year-aged mouse model demonstrated that DPSCs successfully reversed hepatic fibrosis. Overall, DPSCs demonstrated comparable effectiveness to UCMSCs in regulating inflammation and reversing hepatic fibrosis, particularly in the aged mouse model that represents middle-aged and elderly humans. Since autologous DPSCs avoid potential immune-related issues that heterogenous UCMSCs could encounter, they may be a better choice for stem cell-related therapies.

Lymphatic-localized Treg-mregDC crosstalk limits antigen trafficking and restrains anti-tumor immunity.

The tumor microenvironment (TME) has a significant impact on tumor growth and immunotherapy efficacies. However, the precise cellular interactions and spatial organizations within the TME that drive these effects remain elusive. Using advanced multiplex imaging techniques, we have discovered that regulatory T cells (Tregs) accumulate around lymphatic vessels in the peripheral tumor stroma. This localized accumulation is facilitated by mature dendritic cells enriched in immunoregulatory molecules (mregDCs), which promote chemotaxis of Tregs, establishing a peri-lymphatic Treg-mregDC niche. Within this niche, mregDCs facilitate Treg activation, which in turn restrains the trafficking of tumor antigens to the draining mesenteric lymph nodes, thereby impeding the initiation of anti-tumor adaptive immune responses. Disrupting Treg recruitment to mregDCs inhibits tumor progression. Our study provides valuable insights into the organization of TME and how local crosstalk between lymphoid and myeloid cells suppresses anti-tumor immune responses.

Nuclear import of aristaless-related homeobox protein via its NLS1 regulates its transcriptional function.

Nucleocytoplasmic transport of transcription factors is essential in eukaryotes. We previously reported the presence of two functional NLSs in the homeodomain protein, aristaless-related homeobox (Arx) protein, which is a key transcriptional repressor of LMO1, SHOX2, and PAX4 during development. NLS2, that overlaps the homeodomain, is recognized directly by multiple importin ßs, but not by importin αs. In this study, we found that the N-terminal NLS1 of Arx is targeted by multiple importin α proteins, including importin α3 and α5. Both in vivo and in vitro assays demonstrated that nuclear import of Arx via NLS1 is mediated by the importin α/ß pathway. Mutagenesis analysis indicated that two basic amino acids, (84)K and (87)R, are essential to the function of NLS1, and that their mutation prevents interactions of Arx with importin αs. Interestingly, inhibition of nuclear import of Arx via NLS1 clearly attenuates its ability of transcriptional repression, suggesting that nuclear import of Arx via NLS1 contributes to its transcriptional function.

Recent advances in emerging application of functional materials in sample pretreatment methods for liquid chromatography-mass spectrometry analysis of plant growth regulators: A mini-review.

Plant growth regulators (PGRs) are a class of small molecular compounds, which can remarkably affect the physiological process of plants. The complex plant matrix along with a wide polarity range and unstable chemical properties of PGRs hinder their trace analysis. In order to obtain a reliable and accurate result, a sample pretreatment process must be carried out, including eliminating the interference of the matrix effect and pre-concentrating the analytes. In recent years, the research of functional materials in sample pretreatment has experienced rapid growth. This review comprehensively overviews recent development in functional materials covering one-dimensional materials, two-dimensional materials, and three-dimensional materials applied in the pretreatment of PGRs before liquid chromatography-mass spectrometry (LC-MS) analysis. Besides, the advantages and limitations of the above functionalized enrichment materials are discussed, and their future trends have been prospected. The work could be helpful to bring new insights for researchers engaged in functional materials in sample pretreatment of PGRs based on LC-MS.

Increased Cordycepin Production in Yarrowia lipolytica Using Combinatorial Metabolic Engineering Strategies.

As the first nucleoside antibiotic discovered in fungi, cordycepin, with its various biological activities, has wide applications. At present, cordycepin is mainly obtained from the natural fruiting bodies of Cordyceps militaris. However, due to long production periods, low yields, and low extraction efficiency, harvesting cordycepin from natural C. militaris is not ideal, making it difficult to meet market demands. In this study, an engineered Yarrowia lipolytica YlCor-18 strain, constructed by combining metabolic engineering strategies, achieved efficient de novo cordycepin production from glucose. First, the cordycepin biosynthetic pathway derived from C. militaris was introduced into Y. lipolytica. Furthermore, metabolic engineering strategies including promoter, protein, adenosine triphosphate, and precursor engineering were combined to enhance the synthetic ability of engineered strains of cordycepin. Fermentation conditions were also optimized, after which, the production titer and yields of cordycepin in the engineered strain YlCor-18 under fed-batch fermentation were improved to 4362.54 mg/L and 213.85 mg/g, respectively, after 168 h. This study demonstrates the potential of Y. lipolytica as a cell factory for cordycepin synthesis, which will serve as the model for the green biomanufacturing of other nucleoside antibiotics using artificial cell factories.

Karyopherins in nuclear transport of homeodomain proteins during development.

Homeodomain proteins are crucial transcription factors for cell differentiation, cell proliferation and organ development. Interestingly, their homeodomain signature structure is important for both their DNA-binding and their nucleocytoplasmic trafficking. The accurate nucleocytoplasmic distribution of these proteins is essential for their functions. We summarize information on (a) the roles of karyopherins for import and export of homeoproteins, (b) the regulation of their nuclear transport during development, and (c) the corresponding complexity of homeoprotein nucleocytoplasmic transport signals. This article is part of a Special Issue entitled: Regulation of Signaling and Cellular Fate through Modulation of Nuclear Protein Import.

High-level de novo biosynthesis of cordycepin by systems metabolic engineering in Yarrowia lipolytica.

Cordycepin is a nucleoside antibiotic with various biological activities, which has wide applications in the area of cosmetic and medicine industries. However, the current production of cordycepin is costly and time-consuming. To construct the promising cell factory for high-level cordycepin production, firstly, the design and construction of cordycepin biosynthetic pathway were performed in Yarrowia lipolytica. Secondly, the adaptivity between cordycepin biosynthetic pathway and Y. lipolytica was enhanced by enzyme fusion and integration site engineering. Then, the production of cordycepin was improved by the enhancement of adenosine supply. Furthermore, through modular engineering, the production of cordycepin was achieved at 3588.59 mg/L from glucose. Finally, 3249.58 mg/L cordycepin with a yield of 76.46 mg/g total sugar was produced by the engineered strain from the mixtures of glucose and molasses. This research is the first report on the de novo high-level production of cordycepin in the engineered Y. lipolytica.

Early acute fat embolism syndrome caused by femoral fracture: A case report.

BACKGROUND: Fat embolism syndrome (FES) is a rare complication caused by the presence of fat particles in the microcirculation, which usually occurs within 12-72 h after trauma. At present, there have been few cases of fat embolism presenting within 3 h after trauma. Here, we report a case of femoral fracture complicated with an acute fat embolism caused by a car accident. CASE SUMMARY: A 29-year-old woman with pain, swelling and limited movement of her left lower limb after a car accident was taken by ambulance to our hospital. X-ray examination showed fracture of the middle and lower part of the left femur and fracture of the base of the left fifth metatarsal bone. She was hospitalized and admitted to the orthopedic ward. After the attending doctor performed tibial tubercle bone traction, the patient became confused, followed by respiratory distress. Finally, she was transferred to the intensive care unit. After nearly a month of treatment in the intensive care unit, the patient's cognitive function gradually recovered over 6 mo. CONCLUSION: For patients with early traumatic fractures, young emergency physicians and orthopedics should be aware of the possibility of FES.

Identification and validation of the phosphorylation sites on Aristaless-related homeobox protein.

The Aristaless-related homeobox protein (ARX) is a transcription factor expressed in the developing forebrain, skeletal muscle, pancreas, testis, and a variety of other tissues. It is known to have context-dependent transcriptional activator and repressor activity, although how it can achieve these opposing functions remains poorly understood. We hypothesized phosphorylation status might play a role in pivoting ARX between functioning as an activator or repressor. To gain further mechanistic insight as to how ARX functions, we identified multiple phosphorylation sites on ARX. We further established PKA as the kinase that phosphorylates ARX at least at Ser266 in mice. Two other kinases, CK2α and CDK4/cyclin D1, were also identified as kinases that phosphorylate ARX in vitro. Unexpectedly, phosphorylation status did not change either the nuclear localization or transcriptional function of ARX.

Application of Zero-profile Spacer in the Treatment of Three-level Cervical Spondylotic Myelopathy: 5-year Follow-up Results.

STUDY DESIGN: A retrospective study. OBJECTIVE: To assess the long-term results of zero-profile spacer for 3-level anterior cervical discectomy and fusion (ACDF). SUMMARY OF BACKGROUND DATA: Although widely used, there are still controversies about the long-term results of zero-profile spacer, especially in multilevel cases. METHODS: Cases received 3-level ACDF for cervical spondylotic myelopathy (CSM) using either zero-profile spacer (n = 27) (ZP Group), or plate and cages (n = 34) (PC Group), and with 5-year follow-up were reviewed. Neurological function and life quality were assessed by modified Japanese Orthopaedic Association (mJOA) score, Neck Disability Index (NDI), and Short-Form 36 (SF-36) score. Disc height, cervical lordosis, fusion rate, and surgical complications were observed. RESULTS: Neurological recovery and life quality improvement were similar in both groups. Disc height and cervical lordosis (C2-7 Cobb angle) were well restored after operations, but lost in both groups during follow-up. Loss of correction (LOC) in disc height was larger in ZP Group (11.38% vs 5.71%, P < 0.05) at 5-year follow-up. LOC of cervical lordosis in ZP group constantly grew from 11.28% to 48.13% during 5-year follow-up, significantly higher than that in the PC group (from 7.43% to 14.01%) (P < 0.05). The rate of postoperative dysphagia was no statistical difference between the two groups, and symptoms were all disappeared within 1 year. There were 10 levels of adjacent segment degeneration (1 in ZP Group, and 10 in PC Group, P = 0.02). Cage subsidence (11 of 81 levels, 13.58%) and screw migration (2 of 81 levels, 2.47%) were only observed in the ZP Group. The migrated screws in one case were surgically removed. Fusion was achieved in all cases. CONCLUSIONS: In long-term follow-up of 3-level ACDF for CSM, zero-profile spacer has the similar clinical results, but loss of correction of disc height and cervical alignment were significantly higher, compared with anterior plate and cages. LEVEL OF EVIDENCE: 3.