SARS-CoV-2 RNA stabilizes host mRNAs to elicit immunopathogenesis.

SARS-CoV-2 RNA interacts with host factors to suppress interferon responses and simultaneously induces cytokine release to drive the development of severe coronavirus disease 2019 (COVID-19). However, how SARS-CoV-2 hijacks host RNAs to elicit such imbalanced immune responses remains elusive. Here, we analyzed SARS-CoV-2 RNA in situ structures and interactions in infected cells and patient lung samples using RIC-seq. We discovered that SARS-CoV-2 RNA forms 2,095 potential duplexes with the 3' UTRs of 205 host mRNAs to increase their stability by recruiting RNA-binding protein YBX3 in A549 cells. Disrupting the SARS-CoV-2-to-host RNA duplex or knocking down YBX3 decreased host mRNA stability and reduced viral replication. Among SARS-CoV-2-stabilized host targets, NFKBIZ was crucial for promoting cytokine production and reducing interferon responses, probably contributing to cytokine storm induction. Our study uncovers the crucial roles of RNA-RNA interactions in the immunopathogenesis of RNA viruses such as SARS-CoV-2 and provides valuable host targets for drug development.

RNA-binding protein complex AMG-1/SLRP-1 mediates germline development and spermatogenesis by maintaining mitochondrial homeostasis in Caenorhabditis elegans.

The mechanisms of RNA-binding proteins (RBPs)-mediated post-transcriptional regulation of pre-existing mRNAs, which is essential for spermatogenesis, remain poorly understood. In this study, we identify that a germline-specific mitochondrial RBP AMG-1(abnormal mitochondria in germline 1), a homolog of mammalian leucine-rich PPR motif-containing protein (LRPPRC), is required for spermatogenesis in Caenorhabditis elegans. The amg-1 mutation hinders germline development without affecting somatic development and leads to the aberrant mitochondrial morphology and structure associated with mitochondrial dysfunctions specifically in the germline. We demonstrate that AMG-1 is most frequently bound to mtDNA-encoded 12S and 16S ribosomal RNA, the essential components of mitochondrial ribosomes, and that 12S rRNA expression mediated by AMG-1 is crucial for germline mitochondrial protein homeostasis. Furthermore, steroid receptor RNA activator (SRA) stem loop interacting RNA binding protein (SLRP-1), a homolog of mammalian SRA stem loop interacting RNA binding protein (SLIRP) in C. elegans, interacts with AMG-1 genetically to regulate germline development and reproductive success in C. elegans. Overall, these findings reveal the novel function of mtRBP, specifically in regulating germline development.

LSM14B is an Oocyte-Specific RNA-Binding Protein Indispensable for Maternal mRNA Metabolism and Oocyte Development in Mice.

Mammalian oogenesis features reliance on the mRNAs produced and stored during early growth phase. These are essential for producing an oocyte competent to undergo meiotic maturation and embryogenesis later when oocytes are transcriptionally silent. The fate of maternal mRNAs hence ensures the success of oogenesis and the quality of the resulting eggs. Nevertheless, how the fate of maternal mRNAs is determined remains largely elusive. RNA-binding proteins (RBPs) are crucial regulators of oogenesis, yet the identity of the full complement of RBPs expressed in oocytes is unknown. Here, a global view of oocyte-expressed RBPs is presented: mRNA-interactome capture identifies 1396 RBPs in mouse oocytes. An analysis of one of these RBPs, LSM family member 14 (LSM14B), demonstrates that this RBP is specific to oocytes and associated with many networks essential for oogenesis. Deletion of Lsm14b results in female-specific infertility and a phenotype characterized by oocytes incompetent to complete meiosis and early embryogenesis. LSM14B serves as an interaction hub for proteins and mRNAs throughout oocyte development and regulates translation of a subset of its bound mRNAs. Therefore, RNP complexes tethered by LSM14B are found exclusively in oocytes and are essential for the control of maternal mRNA fate and oocyte development.

Selective Translation of Maternal mRNA by eIF4E1B Controls Oocyte to Embryo Transition.

Maternal messenger ribonucleic acids (mRNAs) are driven by a highly orchestrated scheme of recruitment to polysomes and translational activation. However, selecting and regulating individual mRNAs for the translation from a competitive pool of mRNAs are little-known processes. This research shows that the maternal eukaryotic translation initiation factor 4e1b (Eif4e1b) expresses during the oocyte-to-embryo transition (OET), and maternal deletion of Eif4e1b leads to multiple defects concerning oogenesis and embryonic developmental competence during OET. The linear amplification of complementary deoxyribonucleic acid (cDNA) ends, and sequencing (LACE-seq) is used to identify the distinct subset of mRNA and its CG-rich binding sites within the 5' untranslated region (UTR) targeted by eIF4E1B. The proteomics analyses indicate that eIF4E1B-specific bound genes show stronger downregulation at the protein level, which further verify a group of proteins that plays a crucial role in oocyte maturation and embryonic developmental competence is insufficiently synthesized in Eif4e1b-cKO oocytes during OET. Moreover, the biochemical results in vitro are combined to further confirm the maternal-specific translation activation model assembled by eIF4E1B and 3'UTR-associated mRNA binding proteins. The findings demonstrate the indispensability of eIF4E1B for selective translation activation in mammalian oocytes and provide a potential network regulated by eIF4E1B in OET.

Migration and Deposition Law of Pollutants in Urban Sewage Confluence Pipe Network from the Perspective of Ecology.

Aiming at the problem of pollutant migration and deposition in urban sewage confluence pipe, an experimental simulation system of sewage confluence pipe was established. The confluence conditions of three flow patterns (velocity ratio Vaccess/Vtrunk = 0.1/0.2, Vaccess/Vtrunk = 0.1/0.3, and Vaccess/Vtrunk = 0.2/0.3) were simulated. The changes of sediment thickness, carbon pollutants, nitrogen pollutants, and phosphorus pollutants in different confluence areas were analyzed, and the migration and deposition laws of various pollutants in urban sewage confluence pipe network under different flow patterns were revealed. The results show that when the flow velocity of trunk and branch roads changes, the deposition of various pollutants and the carrying capacity of water flow in the pipeline change, resulting in the change of sediment layer thickness and pollutant content. With the increase of trunk velocity, the sediment thickness in the area before and after confluence decreases, while the increase of branch velocity only reduces the sediment thickness in the area at the back of confluence. Under any flow pattern, the sediment thickness in the retention area (G3 and G4) shows an increasing trend, which is the key area of pollution removal. Under the three flow patterns, the content of carbon pollutants reaches the peak at the TCOD and SCOD values of G4 monitoring point. Increasing the trunk velocity can effectively reduce the content of carbon pollutants. The content of nitrogen pollutants in each flow pattern also reaches the maximum at G4 point, which are 213.6 mg/g, 205.2 mg/g, and 212.8 mg/g, respectively. Increasing the trunk velocity can effectively reduce the nitrogen content at points G1-G4, while increasing the flow velocity of the branch road can reduce the nitrogen content at points G5-G7. The distribution of phosphorus pollutants is complex, and the flow pattern needs to be adjusted according to different monitoring points.

Dynamic Evaluation of Intensive Land Use Based on Objective Empowerment by Entropy Method and Neural Network Algorithm.

In the past, the extreme value standardization of indicators, the traditional weighting method, and the multifactor comprehensive model of land intensive use inevitably linearly correlate the evaluation indicators with the evaluation objects, ignoring the direction differences of different indicators in different intervals. At the same time, these methods are also difficult to meet the change of evaluation index weight value with land use type, and cannot adapt to the actual situation of land use environment level and dynamic change. Considering the objectivity of nonlinear correlation moderate index and weight assignment, based on the standardization of quadratic function index and entropy assignment, this paper studies the intensive and dynamic use of land in development zones by different regions to improve the realistic fit of the evaluation model. The results show that the overall level of land intensive use in Chongqing center district and western Chongqing is better than that in northeast Chongqing and southeast Chongqing, roughly showing the state of "high in west and low in east."

Global profiling of RNA-binding protein target sites by LACE-seq.

RNA-binding proteins (RBPs) have essential functions during germline and early embryo development. However, current methods are unable to identify the in vivo targets of a RBP in these low-abundance cells. Here, by coupling RBP-mediated reverse transcription termination with linear amplification of complementary DNA ends and sequencing, we present the LACE-seq method for identifying RBP-regulated RNA networks at or near the single-oocyte level. We determined the binding sites and regulatory mechanisms for several RBPs, including Argonaute 2 (Ago2), Mili, Ddx4 and Ptbp1, in mature mouse oocytes. Unexpectedly, transcriptomics and proteomics analysis of Ago2-/- oocytes revealed that Ago2 interacts with endogenous small interfering RNAs (endo-siRNAs) to repress mRNA translation globally. Furthermore, the Ago2 and endo-siRNA complexes fine-tune the transcriptome by slicing long terminal repeat retrotransposon-derived chimeric transcripts. The precise mapping of RBP-binding sites in low-input cells opens the door to studying the roles of RBPs in embryonic development and reproductive diseases.

Global in situ profiling of RNA-RNA spatial interactions with RIC-seq.

Emerging evidence has demonstrated that RNA-RNA interactions are vital in controlling diverse biological processes, including transcription, RNA splicing and protein translation. RNA in situ conformation sequencing (RIC-seq) is a technique for capturing protein-mediated RNA-RNA proximal interactions globally in living cells at single-base resolution. Cells are first treated with formaldehyde to fix all the protein-mediated RNA-RNA interactions in situ. After cell permeabilization and micrococcal nuclease digestion, the proximally interacting RNAs are 3' end-labeled with pCp-biotin and subsequently ligated using T4 RNA ligase. The chimeric RNAs are then enriched and converted into libraries for paired-end sequencing. After deep sequencing, computational analysis yields interaction strength scores for every base on proximally interacting RNAs in the starting populations. The whole experimental procedure is designed to be completed within 6 d, followed by an additional 8 d for computational analysis. RIC-seq technology can unbiasedly detect intra- and intermolecular RNA-RNA interactions, thereby rendering it useful for reconstructing RNA higher-order structures and identifying direct noncoding RNA targets.

Automated hearing screening for children: A pilot study in China.

OBJECTIVE: To establish the reliability and validity of an automated hearing screening test system for children. DESIGN: Cross-sectional within a comparative study of subjects. STUDY SAMPLE: Subjects were 325 first-grade and second-grade children (6-10 years old) from primary schools in Shenzhen, China. RESULTS: Using the conventional pure-tone screening test with the pass/refer criterion set as 25 dB HL, as the 'gold standard", the sensitivity and specificity of the automated hearing screening test was 0.63 and 0.82, respectively. No specific pattern in the failure rates was observed to relate to the students' grade. There was no statistically significant age effect or gender effect. CONCLUSIONS: The results suggest that with further improvement in terms of its sensitivity and specificity, it may be feasible to use the automated hearing screening test system to conduct routine school hearing screenings.