Distinct Processing of lncRNAs Contributes to Non-conserved Functions in Stem Cells.

Long noncoding RNAs (lncRNAs) evolve more rapidly than mRNAs. Whether conserved lncRNAs undergo conserved processing, localization, and function remains unexplored. We report differing subcellular localization of lncRNAs in human and mouse embryonic stem cells (ESCs). A significantly higher fraction of lncRNAs is localized in the cytoplasm of hESCs than in mESCs. This turns out to be important for hESC pluripotency. FAST is a positionally conserved lncRNA but is not conserved in its processing and localization. In hESCs, cytoplasm-localized hFAST binds to the WD40 domain of the E3 ubiquitin ligase ß-TrCP and blocks its interaction with phosphorylated ß-catenin to prevent degradation, leading to activated WNT signaling, required for pluripotency. In contrast, mFast is nuclear retained in mESCs, and its processing is suppressed by the splicing factor PPIE, which is highly expressed in mESCs but not hESCs. These findings reveal that lncRNA processing and localization are previously under-appreciated contributors to the rapid evolution of function.

The lysine deacetylase activity of histone deacetylases 1 and 2 is required to safeguard zygotic genome activation in mice and cattle.

The genome is transcriptionally inert at fertilization and must be activated through a remarkable developmental process called zygotic genome activation (ZGA). Epigenetic reprogramming contributes significantly to the dynamic gene expression during ZGA; however, the mechanism has yet to be resolved. Here, we find histone deacetylases 1 and 2 (HDAC1/2) can regulate ZGA through lysine deacetylase activity. Notably, in mouse embryos, overexpression of a HDAC1/2 dominant-negative mutant leads to developmental arrest at the two-cell stage. RNA-seq reveals that 64% of downregulated genes are ZGA genes and 49% of upregulated genes are developmental genes. Inhibition of the deacetylase activity of HDAC1/2 causes a failure of histone deacetylation at multiple sites, including H4K5, H4K16, H3K14, H3K18 and H3K27. ChIP-seq analysis exhibits an increase and decrease of H3K27ac enrichment at promoters of up- and downregulated genes, respectively. Moreover, HDAC1 mutants prohibit the removal of H3K4me3 by impeding expression of Kdm5 genes. Importantly, the developmental block can be greatly rescued by Kdm5b injection and by partially correcting the expression of the majority of dysregulated genes. Similar functional significance of HDAC1/2 is conserved in bovine embryos. Overall, we propose that HDAC1/2 are indispensable for ZGA by creating correct transcriptional repressive and active states in mouse and bovine embryos.

Single-cell RNA-Seq reveals the earliest lineage specification and X chromosome dosage compensation in bovine preimplantation embryos.

Lineage specification and X chromosome dosage compensation are two crucial biological processes that occur during preimplantation embryonic development. Although extensively studied in mice, the timing and regulation of these processes remain elusive in other species, including humans. Previous studies have suggested conserved principles of human and bovine early development. This study aims to provide fundamental insights into these programs and the regulation using a bovine embryo model by employing single-cell transcriptomics and genome editing approaches. The study analyzes the transcriptomes of 286 individual cells and reveals that bovine trophectoderm/inner cell mass transcriptomes diverge at the early blastocyst stage, after cavitation but before blastocyst expansion. The study also identifies transcriptomic markers and provides the timing of lineage specification events in the bovine embryo. Importantly, we find that SOX2 is required for the first cell decision program in bovine embryos. Moreover, the study shows the occurrence of X chromosome dosage compensation from morula to late blastocyst and reveals that this compensation results from downregulation of X-linked genes in female embryonic cells. The transcriptional atlas generated by this study is expected to be widely useful in improving our understanding of mammalian early embryo development.

Base editing in bovine embryos reveals a species-specific role of SOX2 in regulation of pluripotency.

The emergence of the first three lineages during development is orchestrated by a network of transcription factors, which are best characterized in mice. However, the role and regulation of these factors are not completely conserved in other mammals, including human and cattle. Here, we establish a gene inactivation system with a robust efficiency by introducing premature codon with cytosine base editors in bovine early embryos. By using this approach, we have determined the functional consequences of three critical lineage-specific genes (SOX2, OCT4 and CDX2) in bovine embryos. In particular, SOX2 knockout results in a failure of the establishment of pluripotency in blastocysts. Indeed, OCT4 level is significantly reduced and NANOG barely detectable. Furthermore, the formation of primitive endoderm is compromised with few SOX17 positive cells. RNA-seq analysis of single blastocysts (day 7.5) reveals dysregulation of 2074 genes, among which 90% are up-regulated in SOX2-null blastocysts. Intriguingly, more than a dozen lineage-specific genes, including OCT4 and NANOG, are down-regulated. Moreover, SOX2 level is sustained in the trophectoderm in absence of CDX2. However, OCT4 knockout does not affect the expression of SOX2. Overall, we propose that SOX2 is indispensable for OCT4 and NANOG expression and CDX2 represses the expression of SOX2 in the trophectoderm in cattle, which are all in sharp contrast with results in mice.

Coxsackievirus A6 2C protein antagonizes IFN-ß production through MDA5 and RIG-I depletion.

IMPORTANCE: Coxsackievirus A6 (CV-A6) is a major emerging pathogen associated with atypical hand, foot, and mouth disease and can cause serious complications such as encephalitis, acute flaccid paralysis, and neurorespiratory syndrome. Therefore, revealing the associated pathogenic mechanisms could benefit the control of CV-A6 infections. In this study, we demonstrate that the nonstructural 2CCV-A6 suppresses IFN-ß production, which supports CV-A6 infection. This is achieved by depleting RNA sensors such as melanoma differentiation-associated gene 5 and retinoic acid-inducible gene I (RIG-I) through the lysosomal pathway. Such a function is shared by 2CEV-A71 and 2CCV-B3 but not 2CCV-A16, suggesting the latter might have an alternative way to promote viral replication. This study broadens our understanding of enterovirus 2C protein regulation of the RIG-I-like receptor signaling pathway and reveals a novel mechanism by which CV-A6 and other enteroviruses evade the host innate immune response. These findings on 2C may provide new therapeutic targets for the development of effective inhibitors against CV-A6 and other enterovirus infections.

S-locus F-Box Protein as Pollen S determinant Targets Non-self S-RNase underlying Self-Incompatibility in Citrus.

Self-incompatibility (SI) is a crucial mechanism that prevents self-fertilization and inbreeding in flowering plants. Citrus exhibits SI regulated by a polymorphic S-locus containing an S-RNase gene and multiple S-locus F-box (SLF) genes. It has been documented that S-RNase functions as the pistil S determinant, but there is no direct evidence that the SLFs closely linked with S-RNase function as pollen S determinants in Citrus. This study assembled the genomes of two pummelo (Citrus grandis) plants and obtained three novel complete and well-annotated S-haplotypes and isolated 36 SLF or SLF-like alleles on the S-loci. Phylogenetic analysis of 138 SLFs revealed that the SLFs were classified into 12 types, including six types with divergent or missing alleles. Furthermore, transformation experiments verified that the conserved S6-SLF7a protein can lead the transition of SI to self-compatibility (SC) by recognizing non-self S8-RNase in 'Mini-Citrus' plants (S7S8 and S8S29, Fortunella hindsii), a model plant for citrus gene function studies. In vitro assays demonstrated interactions between SLFs of different S haplotypes and the Skp1-Cullin1-F-box subunit CgSSK1 protein. This study provides direct evidence that SLF controls the pollen function in Citrus, demonstrating its role in the 'non-self-recognition' SI system.

TEAD4 regulates KRT8 and YAP in preimplantation embryos in mice but not in cattle.

In brief: Lineage specification plays a vital role in preimplantation development. TEAD4 is an essential transcription factor for trophectoderm lineage specification in mice but not in cattle. Abstract: Tead4, a critical transcription factor expressed during preimplantation development, is essential for the expression of trophectoderm-specific genes in mice. However, the functional mechanism of TEAD4 in mouse preimplantation development and its conservation across mammals remain unclear. Here, we report that Tead4 is a crucial transcription factor necessary for blastocyst formation in mice. Disruption of Tead4 through base editing results in developmental arrest at the morula stage. Additionally, RNA-seq analysis reveals dysregulation of 670 genes in Tead4 knockout embryos. As anticipated, Tead4 knockout led to a decrease in trophectoderm genes Cdx2 and Gata3. Intriguingly, we observed a reduction in Krt8, suggesting that Tead4 influences the integrity of the trophectoderm epithelium in mice. More importantly, we noted a dramatic decrease in nuclear Yap in outside cells for Tead4-deficient morula, indicating that Tead4 directly regulates Hippo signaling. In contrast, bovine embryos with TEAD4 depletion could still develop to blastocysts with normal expression of CDX2, GATA3, and SOX2, albeit with a decrease in total cell number and ICM cell number. In conclusion, we propose that Tead4 regulates mouse blastocyst formation via Krt8 and Yap, both of which are critical regulators of mouse preimplantation development.

Microglial SCAP deficiency protects against diabetes-associated cognitive impairment through inhibiting NLRP3 inflammasome-mediated neuroinflammation.

Hyperglycemia-induced pathological microglial responses and subsequent neuronal damage are notable characteristics of diabetes-associated cognitive impairment (DACI). Cholesterol accumulation in the brain is a prevalent consequence of diabetes mellitus (DM), exacerbating pathological microglial responses. Regarding disordered glucose and lipid metabolism, the Sterol Regulatory Element-Binding Protein (SREBP) cleavage-activating protein (SCAP), a cholesterol sensor, exhibits increased expression and abnormal translocation from the endoplasmic reticulum to the Golgi, amplifying the inflammatory response. Therefore, we hypothesized that overexpression of microglia-SCAP and cholesterol accumulation in DM mice could induce pathological microglial responses associated with DACI. Our type 2 DM mice model presented an abnormal increase in microglial SCAP expression. The functional loss of microglia-specific SCAP in DM mice improved cognitive impairment, neuronal synaptic plasticity deficits, and abnormal microglial responses. Mechanistically, the accumulated SCAP directly bound to and enhanced the activation of the microglial-specific inflammatory amplifier, NLRP3 inflammasome, in Golgi, thereby increasing pathological microglial responses and promoting neuronal damage. These findings indicate an important regulatory axis of microglial responses from SCAP to the NLRP3 inflammasome pathway in microglia. These underscore the crosstalk between cholesterol disorders and pathological microglial responses, offering a promising avenue for pharmaceutical interventions in DACI.

Squalene epoxidase promotes breast cancer progression by regulating CCNB1 protein stability.

BACKGROUND: Breast cancer (BC) is a prevalent malignancy affecting women, characterized by a substantial occurrence rate. Squalene epoxidase (SQLE) is a crucial regulator of ferroptosis and has been associated with promoting cell growth and invasion in different types of human cancers. This study aimed to investigate the functional significance of SQLE in BC and elucidate the underlying molecular mechanisms involved. METHODS: SQLE expression levels in BC tissues were evaluated using quantitative real-time polymerase chain reaction, western blotting, and immunohistochemistry. Cell viability, invasion, migration, and cell cycle distribution were assessed using a combination of assays, including the Cell Counting Kit-8, EdU, colony formation, Transwell, and wound healing assays and flow cytometry analysis. Measurement of intracellular reactive oxygen species (ROS), malondialdehyde assay, and glutathione assay were utilized to investigate ferroptosis. Furthermore, co-immunoprecipitation and immunofluorescence assays were conducted to explore the correlation between SQLE and CCNB1. The in vivo tumor growth was evaluated by conducting a xenograft tumorigenicity assay to investigate the impact of SQLE. RESULTS: SQLE expression was significantly increased in BC, and higher SQLE expression levels were significantly associated with an unfavorable prognosis. In vitro functional assays revealed that the overexpression of SQLE markedly enhanced the proliferation, migration, and invasion capacities of BC cells. Furthermore, SQLE overexpression facilitated tumor growth in nude mice. Mechanistically, SQLE alleviated the ubiquitination modification of CCNB1, leading to enhanced stability of the CCNB1 protein and decreased intracellular ROS levels. Ultimately, this inhibited ferroptosis and facilitated the progression of BC. Our findings have provided insights into a crucial pathway by which elevated SQLE expression confers protection to BC cells against ferroptosis, thus promoting cancer progression. SQLE may serve as a novel oncological marker and a potential therapeutic target for BC progression. CONCLUSIONS: In conclusion, this study provides evidence that SQLE plays a regulatory role in BC progression by modulating CCNB1 and ferroptosis. These findings offer valuable insights into the role of SQLE in the pathogenesis of BC and demonstrate its potential as a therapeutic target for treating BC.

Different biomarker ratios in peripheral blood have limited value in diagnosing periprosthetic joint infection after total joint arthroplasty: a single-center, retrospective study.

BACKGROUND: Periprosthetic joint infection (PJI) is a severe complication that can occur after total joint arthroplasty (TJA). The timely and accurate diagnosis of PJI is the key to treatment. This study investigated the diagnostic value of platelet to lymphocyte ratio (PLR), platelet count to mean platelet volume ratio (PVR), neutrophil to lymphocyte ratio (NLR) and monocyte to lymphocyte ratio (MLR) in PJI after total knee arthroplasty (TKA) and total hip arthroplasty (THA). METHODS: We performed a retrospective analysis of the patients who underwent revision hip or knee arthroplasty at our Institute between June 2015 and June 2020. Of the 187 patients reviewed, 168 were included in the study. According to the diagnostic criteria of the Musculoskeletal Infection Society (MSIS), 58 patients were in the PJI group, and 110 patients were in the aseptic loosening (AL) group. We recorded and compared the preoperative peripheral blood white blood cell (WBC) count, platelet count (PLT), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), PLR, PVR, NLR, and MLR in both groups. The diagnostic performance of the WBC, PLT, PLR, PVR, NLR, and MLR individually and in combination with the ESR and CRP for PJI diagnosis was evaluated by receiver operating characteristic (ROC) curves, and the sensitivity, specificity, positive predictive value, and negative predictive value were calculated. RESULTS: Compared to those in the AL group, the mean WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR in the peripheral blood of the PJI group were significantly greater (P < 0.05). The analysis of the ROC curve revealed that the ESR, CRP, PLR, PVR, NLR, and MLR in peripheral blood had moderate effectiveness in diagnosing PJI, with area under the curve (AUC) values of 0.760 (95% CI: 0.688-0.823), 0.758 (95% CI: 0.687-0.821), 0.714 (95% CI: 0.639-0.781), 0.709 (95% CI: 0.634-0.777), 0.723 (95% CI: 0.649-0.789), and 0.728 (95% CI: 0.654-0.793), respectively. Conversely, the WBC and PLT counts demonstrated poor diagnostic value for PJI, with AUC values of 0.578 (95% CI: 0.499-0.653) and 0.694 (95% CI: 0.619-0.763), respectively. The results of the prediction model calculations revealed that the combined AUC of the WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR was the highest at 0.853 (95% CI, 0.790-0.909), indicating good value in the diagnosis of PJI, with a sensitivity of 82.8% and a specificity of 72.7%. Moreover, the novel composite of parameters improved the accuracy and reliability in diagnosing PJI compared to the traditional biomarkers ESR and CRP (P = 0.015). CONCLUSION: Our study suggested that the diagnostic value of the peripheral blood biomarkers PLR, PVR, NLR, and MLR for diagnosing PJI is limited and not superior to that of the ESR or CRP. However, when the WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR are combined, the diagnostic performance of PJI in TJA patients can be improved.

Incidence of cardiovascular mortality among head and neck cancer patients.

BACKGROUND: While treatment advancements have prolonged the lives of patients with head and neck cancer, the subgroups of these patients at higher risk for cardiovascular disease (CVD) mortality remain unclear. METHODS: We analyzed data from the Surveillance, Epidemiology, and End Results (SEER) database for patients diagnosed with head and neck cancer from 2000 to 2019. We compared their CVD mortality against the general US population using standardized mortality ratios (SMRs). RESULTS: Our analysis included 474,366 patients, identifying that 14% of deaths were due to CVD, with an SMR of 1.19. Notably, patients under the age of 39 had a CVD SMR increase of over 100-fold. Those with distant tumor stages showed the highest CVD SMR of 1.52 (95% CI 1.50-1.54). An upward trend in SMR to 2.53 (95% CI 2.51-2.56) was observed from 2011 to 2019. Within the initial 5-year post-diagnosis, the SMR for CVD was 3.17 (95% CI 3.14-3.20), which exceeded the general population's rates but declined in the 5-20-year range after diagnosis. Patients who did not any therapy had the greatest CVD SMR of 2.26 (95% CI 2.24-2.28). Hypopharyngeal cancer patients exhibited the highest CVD SMR of 1.54 (95% CI 1.52-1.56). CONCLUSIONS: The study highlights that head and neck cancer patients, especially younger individuals and those with advanced disease stages, face substantial CVD mortality risks. The CVD SMR peaks within 5 years following diagnosis. Patients abstaining from treatment bear the highest risk of CVD mortality. Cardioprotective measures should be considered critical for this patient population.

Manipulating the Spin State of Spinel Octahedral Sites via a π-π Type Orbital Coupling to Boost Water Oxidation.

Spin state is often regarded as the crucial valve to release the reactivity of energy-related catalysts, yet it is also challenging to precisely manipulate, especially for the active center ions occupied at the specific geometric sites. Herein, a π-π type orbital coupling of 3d (Co)-2p (O)-4f (Ce) was employed to regulate the spin state of octahedral cobalt sites (CoOh) in the composite of Co3O4/CeO2. More specifically, the equivalent high-spin ratio of CoOh can reach to 54.7% via tuning the CeO2 content, thereby triggering the average eg filling (1.094) close to the theoretical optimum value. The corresponding catalyst exhibits a superior water oxidation performance with an overpotential of 251 mV at 10 mA cm-2, rivaling most cobalt-based oxides state-of-the-art. The π-π type coupling corroborated by the matched energy levels between Ce t1u/t2u-O and CoOh t2g-O π type bond in the calculated crystal orbital Hamilton population and partial density of states profiles, stimulates a π-donation between O 2p and π-symmetric Ce 4fyz2 orbital, consequently facilitating the electrons hopping from t2g to eg orbital of CoOh. This work offers an in-depth insight into understanding the 4f and 3d orbital coupling for spin state optimization in composite oxides.

Enantiocontrolled Total Synthesis of (-)-Retigeranic Acid A.

The asymmetric total synthesis of (-)-retigeranic acid A has been realized. The key features of the current synthesis include (1) a Pt-catalyzed Conia-ene 5-exo-dig cyclization of enolyne to establish the key quaternary stereochemical center of C-10 (D/E ring), (2) an intramolecular diastereoselective Prins cyclization to construct the trans-hydrindane backbone (A/B ring), and (3) a late-stage intramolecular Fe-mediated hydrogen atom transfer (HAT) Baldwin-disfavored 5-endo-trig radical cyclization to rapidly assemble vicinal quaternary centers and the core structure of (-)-retigeranic acid A (C ring).

Systemic inflammation indicators and risk of incident arrhythmias in 478,524 individuals: evidence from the UK Biobank cohort.

BACKGROUND: The role of systemic inflammation in promoting cardiovascular diseases has attracted attention, but its correlation with various arrhythmias remains to be clarified. We aimed to comprehensively assess the association between various indicators of systemic inflammation and atrial fibrillation/flutter (AF), ventricular arrhythmia (VA), and bradyarrhythmia in the UK Biobank cohort. METHODS: After excluding ineligible participants, a total of 478,524 eligible individuals (46.75% male, aged 40-69 years) were enrolled in the study to assess the association between systemic inflammatory indicators and each type of arrhythmia. RESULTS: After covariates were fully adjusted, CRP levels were found to have an essentially linear positive correlation with the risk of various arrhythmias; neutrophil count, monocyte count, and NLR showed a non-linear positive correlation; and lymphocyte count, SII, PLR, and LMR showed a U-shaped association. VA showed the strongest association with systemic inflammation indicators, and it was followed sequentially by AF and bradyarrhythmia. CONCLUSIONS: Multiple systemic inflammatory indicators showed strong associations with the onset of AF, VA, and bradyarrhythmia, of which the latter two have been rarely studied. Active systemic inflammation management might have favorable effects in reducing the arrhythmia burden and further randomized controlled studies are needed.

Interfacial Engineering of Fluorinated TiO2 Nanosheets with Abundant Oxygen Vacancies for Boosting the Hydrogen Storage Performance of MgH2.

The interaction between fluorinated surface in the partially reduced nano-crystallite titanium dioxide (TiO2-x (F)) and MgH2 is studied for the first time. Compared with pristine MgH2 (416 °C), the onset desorption temperature of MgH2 +5 wt.% TiO2-x (F) composite can be dramatically lowered to 189 °C. In addition, the composite exhibits remarkable dehydrogenation kinetics, which can release 6.0 wt.% hydrogen thoroughly within 6 min at 250 °C. The apparent activation energy for dehydriding is decreased from 268.42 to 119.96 kJ mol-1 . Structural characterization and theoretical calculations indicate that the synergistic effect between multivalent Ti species, and the in situ formed MgF2 and MgF2-x Hx is beneficial for improving the hydrogen storage performance of MgH2 . Moreover, oxygen vacancies can accelerate the electron transportation and facilitate hydrogen diffusion. The study provides a novel perspective on the modification of MgH2 by fluorinated transition metal oxide catalyst.

Nef Suppresses LINE-1 Retrotransposition through Two Distinct Mechanisms.

Long interspersed element type 1 (LINE-1) is the only known type of retroelement that can replicate autonomously, and its retrotransposition activity can trigger interferon (IFN) production. IFN production suppresses the infectivity of exogenous viruses, such as human immunodeficiency virus (HIV). As a counteraction, HIV has been reported to use multiple proteins and mechanisms to suppress LINE-1 replication. However, the mechanisms of HIV-mediated LINE-1 regulation are not fully understood. In this study, we discovered that Nef protein, which is expressed by HIV and is important for HIV pathogenesis, inhibits LINE-1 retrotransposition. Two distinct mechanisms have been uncovered for Nef-induced LINE-1 suppression. Without direct interaction with LINE-1 DNA, Nef potently inhibits the promoter activity of the LINE-1 5'-untranslated region (5'-UTR) and reduces the expression levels of LINE-1 RNA and proteins. Alternatively, although Nef does not bind to the LINE-1 open reading frame 1 protein (ORF1p) or LINE-1 RNA, it significantly compromises the ORF1p-LINE-1 RNA interaction, which is essential for LINE-1 retrotransposition. Both mechanisms can be suppressed by the G2A mutation, which abolishes myristoylation of Nef, suggesting that membrane attachment is essential for Nef to suppress LINE-1. Consequently, through LINE-1 inhibition, Nef downregulates IFN production in host cells. Therefore, our data revealed that Nef is a potent LINE-1 suppressor and an effective innate immune regulator, which not only provides new information on the intricate interaction between HIV, LINE-1, and IFN signaling systems but also strengthens the importance of Nef in HIV infection and highlights the potential of designing novel Nef-targeting anti-HIV drugs. IMPORTANCE Human immunodeficiency viruses are pathogens of AIDS that were first discovered almost 40 years ago and continue to threaten human lives to date. While currently used anti-HIV drugs are sufficient to suppress viral loads in HIV-infected patients, both drug-resistant HIV strains and adverse side effects triggered by the long-term use of these drugs highlight the need to develop novel anti-HIV drugs targeting different viral proteins and/or different steps in viral replication. To achieve this, more information is required regarding HIV pathogenesis and especially its impact on cellular activities in host cells. In this study, we discovered that the Nef protein expressed by HIV potently inhibits LINE-1 retrotransposition. During our attempt to determine the mechanism of Nef-mediated LINE-1 suppression, two additional functions of Nef were uncovered. Nef effectively repressed the promoter activity of LINE-1 5'-UTR and destabilized the interaction between ORF1p and LINE-1 RNA. Consequently, Nef not only compromises LINE-1 replication but also reduces LINE-1-triggered IFN production. The reduction in IFN production, in theory, promotes HIV infectivity. Together with its previously known functions, these findings indicate that Nef is a potential target for the development of novel anti-HIV drugs. Notably, the G2 residue, which has been reported to be essential for most Nef functions, was found to be critical in the regulation of innate immune activation by Nef, suggesting that compromising myristoylation or membrane attachment of Nef may be a good strategy for the inhibition of HIV infection.

BST2 Suppresses LINE-1 Retrotransposition by Reducing the Promoter Activity of LINE-1 5' UTR.

Endogenous retrotransposons are considered the "molecular fossils" of ancient retroviral insertions. Several studies have indicated that host factors restrict both retroviruses and retrotransposons through different mechanisms. Type 1 long interspersed elements (LINE-1 or L1) are the only active retroelements that can replicate autonomously in the human genome. A recent study reported that LINE-1 retrotransposition is potently suppressed by BST2, a host restriction factor that prevents viral release mainly by physically tethering enveloped virions (such as HIV) to the surface of producer cells. However, no endoplasmic membrane structure has been associated with LINE-1 replication, suggesting that BST2 may utilize a distinct mechanism to suppress LINE-1. In this study, we showed that BST2 is a potent LINE-1 suppressor. Further investigations suggested that BST2 reduces the promoter activity of LINE-1 5' untranslated region (UTR) and lowers the levels of LINE-1 RNA, proteins, and events during LINE-1 retrotransposition. Surprisingly, although BST2 apparently uses different mechanisms against HIV and LINE-1, two membrane-associated domains that are essential for BST2-mediated HIV tethering also proved important for BST2-induced inhibition of LINE-1 5' UTR. Additionally, by suppressing LINE-1, BST2 prevented LINE-1-induced genomic DNA damage and innate immune activation. Taken together, our data uncovered the mechanism of BST2-mediated LINE-1 suppression and revealed new roles of BST2 as a promoter regulator, genome stabilizer, and innate immune suppressor. IMPORTANCE BST2 is a potent antiviral protein that suppresses the release of several enveloped viruses, mainly by tethering the envelope of newly synthesized virions and restraining them on the surface of producer cells. In mammalian cells, there are numerous DNA elements replicating through reverse transcription, among which LINE-1 is the only retroelement that can replicate autonomously. Although LINE-1 retrotransposition does not involve the participation of a membrane structure, BST2 has been reported as an efficient LINE-1 suppressor, suggesting a different mechanism for BST2-mediated LINE-1 inhibition and a new function for BST2 itself. We found that BST2 specifically represses the promoter activity of LINE-1 5' UTR, resulting in decreased levels of LINE-1 transcription, translation, and subsequent retrotransposition. Additionally, by suppressing LINE-1 activity, BST2 maintains genome stability and regulates innate immune activation. These findings expand our understanding of BST2 and its biological significance.

RNA Sequencing of Idiopathic Subglottic Stenosis Tissues Uncovers Putative Profibrotic Mechanisms and Identifies a Prognostic Biomarker.

Idiopathic subglottic stenosis (iSGS) is a localized airway disease that almost exclusively affects females. Understanding the molecular mechanisms involved may provide insights leading to therapeutic interventions. Next-generation sequencing was performed on tissue sections from patients with iSGS (n = 22), antineutrophil cytoplasmic antibody-associated vasculitis (AAV; n = 5), and matched controls (n = 9) to explore candidate genes and mechanisms of disease. Gene expression changes were validated, and selected markers were identified by immunofluorescence staining. Epithelial-mesenchymal transition (EMT) and leukocyte extravasation pathways were the biological mechanisms most relevant to iSGS pathogenesis. Alternatively activated macrophages (M2) were abundant in the subepithelium and perisubmucosal glands of the airway in iSGS and AAV. Increased expression of the mesenchymal marker S100A4 and decreased expression of the epithelial marker epithelial cell adhesion molecule (EPCAM) further supported a role for EMT, but to different extents, in iSGS and antineutrophil cytoplasmic antibody-associated subglottic stenosis. In patients with iSGS, high expression of prostate transmembrane protein, androgen induced 1 (PMEPA1), an EMT regulator, was associated with a shorter recurrence interval (25 versus 116 months: hazard ratio = 4.16; P = 0.041; 95% CI, 1.056-15.60). Thus, EMT is a key pathogenetic mechanism of subglottic stenosis in iSGS and AAV. M2 macrophages contribute to the pathogenesis of both diseases, suggesting a shared profibrotic mechanism, and PMEPA1 may be a biomarker for predicting disease recurrence in iSGS.

Functional role of GATA3 and CDX2 in lineage specification during bovine early embryonic development.

In brief: The lineage specification during early embryonic development in cattle remains largely elusive. The present study determines the effects of trophectoderm-associated factors GATA3 and CDX2 on lineage specification during bovine early embryonic development. Abstract: Current understandings of the initiation of the trophectoderm (TE) program during mammalian embryonic development lack evidence of how TE-associated factors such as GATA3 and CDX2 participate in bovine lineage specification. In this study, we describe the effects of TE-associated factors on the expression of lineage specification marker genes such as SOX2, OCT4, NANOG, GATA6, and SOX17, by using cytosine base editor system. We successfully knockout GATA3 or CDX2 in bovine embryos with a robust efficiency. However, GATA3 or CDX2 deletion does not affect the developmental potential of embryos to reach the blastocyst stage. Interestingly, GATA3 deletion downregulates the NANOG expression in bovine blastocysts. Further analysis of the mosaic embryos shows that GATA3 is required for NANOG in the TE of bovine blastocysts. Single blastocyst RNA-seq analysis reveals that GATA3 deletion disrupts the transcriptome in bovine blastocysts. Altogether, we propose that GATA3 plays an important role in maintaining TE lineage program in bovine embryos and the functional role of GATA3 is species-specific.

Canonical and noncanonical regulatory roles for JAK2 in the pathogenesis of rheumatoid arthritis-associated interstitial lung disease and idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) and rheumatoid arthritis-associated interstitial lung disease (RA-ILD) are two fibrotic interstitial lung diseases that share the usual interstitial pneumonia (UIP) injury pattern. Here, we report that RNA sequencing of lung biopsies from patients with RA-ILD and IPF revealed shared and distinct disease-causing pathways. Analysis of transcriptomic data identified a JAK2 related JAK/STAT signaling pathway gene signature that distinguishes RA-UIP from idiopathic UIP. This was further confirmed by immunohistostaining, which identified JAK2 phosphorylation with two distinct forms of activation: a cytoplasmic form of JAK2 activation in most IPF cases (13/20) and a nuclear form of p-JAK2 in RA-UIP (5/5) and a minority of IPF (6/20) cases. Further immunohistostaining identified STAT5A&B as the downstream transcriptional activator for JAK2-mediated canonical signal transduction and phosphorylation of Tyr41 on histone H3 (H3Y41ph) as the downstream epigenetic regulation site for JAK2-mediated noncanonical signal transduction. Gene Set Enrichment Analysis (GSEA) of the RNA-Seq data further supported this shared pathogenic mechanism for the two diseases with the enrichment of STAT5A&B target gene sets as well as the JAK2 regulated H3Y41ph target gene set. This regulatory role of JAK2 in the pathogenesis of pulmonary fibrosis was further demonstrated by the attenuation of bleomycin-induced murine pulmonary fibrosis using a JAK2-selective pharmacological inhibitor CEP33779. In vitro studies with normal and IPF derived lung fibroblasts revealed a central role for JAK2 as an essential intermediary molecule in TGF-ß-mediated myofibroblast trans-differentiation, proliferation, and extracellular matrix protein production. These observations support a crucial role for JAK2 as an intermediary molecule in fibrotic lung disease development.