Nanopore strand-specific mismatch enables de novo detection of bacterial DNA modifications.

DNA modifications in bacteria present diverse types and distributions, playing crucial functional roles. Current methods for detecting bacterial DNA modifications via nanopore sequencing typically involve comparing raw current signals to a methylation-free control. In this study, we found that bacterial DNA modification induces errors in nanopore reads. And these errors are found only in one strand but not the other, showing a strand-specific bias. Leveraging this discovery, we developed Hammerhead, a pioneering pipeline designed for de novo methylation discovery that circumvents the necessity of raw signal inference and a methylation-free control. The majority (14 out of 16) of the identified motifs can be validated by raw signal comparison methods or by identifying corresponding methyltransferases in bacteria. Additionally, we included a novel polishing strategy employing duplex reads to correct modification-induced errors in bacterial genome assemblies, achieving a reduction of over 85% in such errors. In summary, Hammerhead enables users to effectively locate bacterial DNA methylation sites from nanopore FASTQ/FASTA reads, thus holds promise as a routine pipeline for a wide range of nanopore sequencing applications, such as genome assembly, metagenomic binning, decontaminating eukaryotic genome assembly, and functional analysis for DNA modifications.

Analysis of bacterial transcriptome and epitranscriptome using nanopore direct RNA sequencing.

Bacterial gene expression is a complex process involving extensive regulatory mechanisms. Along with growing interests in this field, Nanopore Direct RNA Sequencing (DRS) provides a promising platform for rapid and comprehensive characterization of bacterial RNA biology. However, the DRS of bacterial RNA is currently deficient in the yield of mRNA-mapping reads and has yet to be exploited for transcriptome-wide RNA modification mapping. Here, we showed that pre-processing of bacterial total RNA (size selection followed by ribosomal RNA depletion and polyadenylation) guaranteed high throughputs of sequencing data and considerably increased the amount of mRNA reads. This way, complex transcriptome architectures were reconstructed for Escherichia coli and Staphylococcus aureus and extended the boundaries of 225 known E. coli operons and 89 defined S. aureus operons. Utilizing unmodified in vitro-transcribed (IVT) RNA libraries as a negative control, several Nanopore-based computational tools globally detected putative modification sites in the E. coli and S. aureus transcriptomes. Combined with Next-Generation Sequencing-based N6-methyladenosine (m6A) detection methods, 75 high-confidence m6A candidates were identified in the E. coli protein-coding transcripts, while none were detected in S. aureus. Altogether, we demonstrated the potential of Nanopore DRS in systematic and convenient transcriptome and epitranscriptome analysis.

Primerdiffer: a python command-line module for large-scale primer design in haplotype genotyping.

MOTIVATION: Primer design is a routine practice for modern molecular biology labs. Bioinformatics tools like primer3 and primer-blast have standardized the primer design for a specific region. However, large-scale primer design, especially for genome-wide screening, is still a labor-intensive job for most wet-lab researchers using these pipelines. RESULTS: Here, we present the primerdiffer pipeline, which can be used to batch design primers that differentiate haplotypes on a large scale with precise false priming checking. This command-line interface (CLI) pipeline includes greedy primer search, local and global in silico PCR-based false priming checking, and automated best primer selection. The local CLI application provides flexibility to design primers with the user's own genome sequences and specific parameters. Some species-specific primers designed to genotype the hybrid introgression strains from Caenorhabditis briggsae and Caenorhabditis nigoni have been validated using single-worm PCR. This pipeline provides the first CLI-based large-scale primer design tool to differentiate haplotypes in any targeted region. AVAILABILITY AND IMPLEMENTATION: The open-source python modules are available at github (https://github.com/runsheng/primerdiffer, https://github.com/runsheng/primervcf) and Python package index (https://pypi.org/project/primerdiffer/, https://pypi.org/project/primervcf/).

Chromosome level genome assembly and transcriptome analysis of E11 cells infected by tilapia lake virus.

The E11 cell line, derived from striped snakehead fish (Channa striata), possesses a distinctive feature: it is persistently infected with a C-type retrovirus. Notably, it exhibits high permissiveness to piscine nodavirus and the emerging tilapia lake virus (TiLV). Despite its popularity in TiLV research, the absence of genome assembly for the E11 cell line and Channa striata has constrained research on host-virus interactions. This study aimed to fill this gap by sequencing, assembling, and annotating the E11 cell line genome. Our efforts yielded a 600.5 Mb genome including 24 chromosomes with a BUSCO score of 98.8%. In addition, the complete proviral DNA sequence of snakehead retrovirus (SnRV) was identified in the E11 cell genome. Comparative genomic analysis between the E11 cell line and another snakehead species Channa argus revealed the loss of many immune-related gene families in the E11 cell genome, indicating a compromised immune response. We also conducted transcriptome analysis of mock- and TiLV-infected E11 cells, unveiling new perspectives on virus-virus and host-virus interactions. The TiLV infection suppressed the high expression of SnRV in E11 cells, and activated some other endogenous retroviruses. The protein-coding gene comparison revealed a pronounced up-regulation of genes involved in immune response, alongside a down-regulation of genes associated with specific metabolic processes. In summary, the genome assembly and annotation of the E11 cell line provide valuable resources to understand the SnRV and facilitate further studies on nodavirus and TiLV. The RNA-seq profiles shed light on the cellular mechanisms employed by fish cells in response to viral challenges, potentially guiding the development of therapeutic strategies against TiLV in aquaculture. This study also provides the first insights into the viral transcriptome profiles of endogenous SnRV and evading TiLV, enhancing our understanding of host-virus interactions in fish.

A comparative analysis of alternative splicing patterns in Atlantic salmon (Salmo salar) in response to Moritella viscosa and sea lice (Lepeophtheirus salmonis) infection.

Moritella viscosa (M. viscosa) and sea lice (Lepeophtheirus salmonis) are severe pathogens that primarily infect the skin of Atlantic salmon (Salmo salar), which cause significant economic losses in the farming industry. However, the pathogenesis and molecular mechanisms underlying the host's immune defence at the post-transcriptional level remain unclear. Alternative splicing (AS) is an evolutionarily conserved post-transcriptional mechanism that can greatly increase the richness of the transcriptome and proteome. In this study, transcriptomic data derived from skin tissues of Atlantic salmon after M. viscosa and sea lice infections were used to examine the AS profiles and their differential expression patterns. In total, we identified 33,044 AS events (involving 13,718 genes) in the control (CON) group, 35,147 AS events (involving 14,340 genes) in the M. viscosa infection (MV) group, and 30,364 AS events (involving 13,142 genes) in the sea lice infection (LC) group, respectively. Among the five types of AS identified in our study (i.e., SE, A5SS, A3SS, MXE, and RI), SE was the most prevalent type in all three groups (i.e., CON, MV, and LC groups). Decreased percent-spliced-in (PSI) levels were observed in SE events under both MV- and LC-infected conditions, suggesting that MV or LC infection elevated exon-skipping isoforms and promoted the selection of shorter transcripts in numerous DAS genes. In addition, most of the differential AS genes were found to be associated with pathways related to mRNA regulation, epithelial or muscle development, and immune response. These findings provide novel insights into the role of AS in host-pathogen interactions and represent the first comparative analysis of AS in response to bacterial and parasitic infections in fish.

Direct full-length RNA sequencing reveals unexpected transcriptome complexity during Caenorhabditis elegans development.

Massively parallel sequencing of the polyadenylated RNAs has played a key role in delineating transcriptome complexity, including alternative use of an exon, promoter, 5' or 3' splice site or polyadenylation site, and RNA modification. However, reads derived from the current RNA-seq technologies are usually short and deprived of information on modification, compromising their potential in defining transcriptome complexity. Here, we applied a direct RNA sequencing method with ultralong reads using Oxford Nanopore Technologies to study the transcriptome complexity in Caenorhabditis elegans We generated approximately six million reads using native poly(A)-tailed mRNAs from three developmental stages, with average read lengths ranging from 900 to 1100 nt. Around half of the reads represent full-length transcripts. To utilize the full-length transcripts in defining transcriptome complexity, we devised a method to classify the long reads as the same as existing transcripts or as a novel transcript using sequence mapping tracks rather than existing intron/exon structures, which allowed us to identify roughly 57,000 novel isoforms and recover at least 26,000 out of the 33,500 existing isoforms. The sets of genes with differential expression versus differential isoform usage over development are largely different, implying a fine-tuned regulation at isoform level. We also observed an unexpected increase in putative RNA modification in all bases in the coding region relative to the UTR, suggesting their possible roles in translation. The RNA reads and the method for read classification are expected to deliver new insights into RNA processing and modification and their underlying biology in the future.

Formation of artificial chromosomes in Caenorhabditis elegans and analyses of their segregation in mitosis, DNA sequence composition and holocentromere organization.

To investigate how exogenous DNA concatemerizes to form episomal artificial chromosomes (ACs), acquire equal segregation ability and maintain stable holocentromeres, we injected DNA sequences with different features, including sequences that are repetitive or complex, and sequences with different AT-contents, into the gonad of Caenorhabditis elegans to form ACs in embryos, and monitored AC mitotic segregation. We demonstrated that AT-poor sequences (26% AT-content) delayed the acquisition of segregation competency of newly formed ACs. We also co-injected fragmented Saccharomyces cerevisiae genomic DNA, differentially expressed fluorescent markers and ubiquitously expressed selectable marker to construct a less repetitive, more complex AC. We sequenced the whole genome of a strain which propagates this AC through multiple generations, and de novo assembled the AC sequences. We discovered CENP-AHCP-3 domains/peaks are distributed along the AC, as in endogenous chromosomes, suggesting a holocentric architecture. We found that CENP-AHCP-3 binds to the unexpressed marker genes and many fragmented yeast sequences, but is excluded in the yeast extremely high-AT-content centromeric and mitochondrial DNA (> 83% AT-content) on the AC. We identified A-rich motifs in CENP-AHCP-3 domains/peaks on the AC and on endogenous chromosomes, which have some similarity with each other and similarity to some non-germline transcription factor binding sites.

Genomic architecture of 5S rDNA cluster and its variations within and between species.

BACKGROUND: Ribosomal DNAs (rDNAs) are arranged in purely tandem repeats, preventing them from being reliably assembled onto chromosomes during generation of genome assembly. The uncertainty of rDNA genomic structure presents a significant barrier for studying their function and evolution. RESULTS: Here we generate ultra-long Oxford Nanopore Technologies (ONT) and short NGS reads to delineate the architecture and variation of the 5S rDNA cluster in the different strains of C. elegans and C. briggsae. We classify the individual rDNA's repeating units into 25 types based on the unique sequence variations in each unit of C. elegans (N2). We next perform assembly of the cluster by taking advantage of the long reads that carry these units, which led to an assembly of 5S rDNA cluster consisting of up to 167 consecutive 5S rDNA units in the N2 strain. The ordering and copy number of various rDNA units are consistent with the separation time between strains. Surprisingly, we observed a drastically reduced level of variation in the unit composition in the 5S rDNA cluster in the C. elegans CB4856 and C. briggsae AF16 strains than in the C. elegans N2 strain, suggesting that N2, a widely used reference strain, is likely to be defective in maintaining the 5S rDNA cluster stability compared with other wild isolates of C. elegans or C. briggsae. CONCLUSIONS: The results demonstrate that Nanopore DNA sequencing reads are capable of generating assembly of highly repetitive sequences, and rDNA units are highly dynamic both within and between population(s) of the same species in terms of sequence and copy number. The detailed structure and variation of the 5S rDNA units within the rDNA cluster pave the way for functional and evolutionary studies.

Host-Endosymbiont Genome Integration in a Deep-Sea Chemosymbiotic Clam.

Endosymbiosis with chemosynthetic bacteria has enabled many deep-sea invertebrates to thrive at hydrothermal vents and cold seeps, but most previous studies on this mutualism have focused on the bacteria only. Vesicomyid clams dominate global deep-sea chemosynthesis-based ecosystems. They differ from most deep-sea symbiotic animals in passing their symbionts from parent to offspring, enabling intricate coevolution between the host and the symbiont. Here, we sequenced the genomes of the clam Archivesica marissinica (Bivalvia: Vesicomyidae) and its bacterial symbiont to understand the genomic/metabolic integration behind this symbiosis. At 1.52 Gb, the clam genome encodes 28 genes horizontally transferred from bacteria, a large number of pseudogenes and transposable elements whose massive expansion corresponded to the timing of the rise and subsequent divergence of symbiont-bearing vesicomyids. The genome exhibits gene family expansion in cellular processes that likely facilitate chemoautotrophy, including gas delivery to support energy and carbon production, metabolite exchange with the symbiont, and regulation of the bacteriocyte population. Contraction in cellulase genes is likely adaptive to the shift from phytoplankton-derived to bacteria-based food. It also shows contraction in bacterial recognition gene families, indicative of suppressed immune response to the endosymbiont. The gammaproteobacterium endosymbiont has a reduced genome of 1.03 Mb but retains complete pathways for sulfur oxidation, carbon fixation, and biosynthesis of 20 common amino acids, indicating the host's high dependence on the symbiont for nutrition. Overall, the host-symbiont genomes show not only tight metabolic complementarity but also distinct signatures of coevolution allowing the vesicomyids to thrive in chemosynthesis-based ecosystems.

New insights into Arabidopsis transcriptome complexity revealed by direct sequencing of native RNAs.

Arabidopsis thaliana transcriptomes have been extensively studied and characterized under different conditions. However, most of the current 'RNA-sequencing' technologies produce a relatively short read length and demand a reverse-transcription step, preventing effective characterization of transcriptome complexity. Here, we performed Direct RNA Sequencing (DRS) using the latest Oxford Nanopore Technology (ONT) with exceptional read length. We demonstrate that the complexity of the A. thaliana transcriptomes has been substantially under-estimated. The ONT direct RNA sequencing identified novel transcript isoforms at both the vegetative (14-day old seedlings, stage 1.04) and reproductive stages (stage 6.00-6.10) of development. Using in-house software called TrackCluster, we determined alternative transcription initiation (ATI), alternative polyadenylation (APA), alternative splicing (AS), and fusion transcripts. More than 38 500 novel transcript isoforms were identified, including six categories of fusion-transcripts that may result from differential RNA processing mechanisms. Aided by the Tombo algorithm, we found an enrichment of m5C modifications in the mobile mRNAs, consistent with a recent finding that m5C modification in mRNAs is crucial for their long-distance movement. In summary, ONT DRS offers an advantage in the identification and functional characterization of novel RNA isoforms and RNA base modifications, significantly improving annotation of the A. thaliana genome.

Flurochloridone induced abnormal spermatogenesis by damaging testicular Sertoli cells in mice.

BACKGROUND: Flurochloridone (FLC), a selective herbicide used on a global scale, has been reported to have male reproductive toxicity whose evidence is limited, but its mechanism remains unclear. The present study was conducted to systematically explore the male reproductive toxicity of FLC, including sperm quality, spermatogenesis, toxicity targets, and potential mechanisms. METHODS: Male C57BL/6 mice aged 6-7 weeks received gavage administration of FLC (365/730 mg/kg/day) for 28 consecutive days. Then, the tissue and sperm of mice were collected for analysis. We measured the gonadosomatic index and analyzed sperm concentration, motility, malformation rate, and mitochondrial membrane potential (MMP). Spermatocyte immunofluorescence staining was performed to analyze meiosis. We also performed pathological staining on the testis and epididymis tissue and TUNEL staining, immunohistochemical analysis, and ultrastructural observation on the testicular tissue. RESULTS: Results showed that FLC caused testicular weight reduction, dysfunction, and architectural damage in mice, but no significant adverse effect was found in the epididymis. The exposure interfered with spermatogonial proliferation and meiosis, affecting sperm concentration, motility, kinematic parameters, morphology, and MMP, decreasing sperm quality. Furthermore, mitochondrial damage and apoptosis of testicular Sertoli cells were observed in mice treated with FLC. CONCLUSION: We found that FLC has significant adverse effects on spermatogonial proliferation and meiosis. Meanwhile, apoptosis and mitochondrial damage may be the potential mechanism of Sertoli cell damage. Our study demonstrated that FLC could induce testicular Sertoli cell damage, leading to abnormal spermatogenesis, which decreased sperm quality. The data provided references for the toxicity risk and research methods of FLC application in the environment.

Melatonin improves bisphenol A-induced cell apoptosis, oxidative stress and autophagy impairment via inhibition of the p38 MAPK signaling pathway in FLK-BLV cells.

The aim of this study was to assess the protective effect and potential mechanism of melatonin against bisphenol A (BPA)-induced apoptosis and oxidative damage in FLK-BLV cells. The results showed that BPA reduced cell viability in a dose- and time-dependent manner, caused cell shrinkage and induced oxidative stress and apoptosis in FLK-BLV cells, which were effectively reversed by melatonin. In addition, BPA caused autophagy flux impairment, which was confirmed by the increased of LC3-II and p62 levels, whereas melatonin treatment effectively reduced p62 levels under BPA treatment, and reversed apoptosis-related protein expression patterns caused by BPA. However, inhibition of autophagy by CQ partially abolished the protective effect of melatonin on apoptosis, suggesting that melatonin against BPA-induced oxidative injury and apoptosis by activating autophagy pathway. Moreover, we found that melatonin inhibited BPA-induced the activation of p38 MAPK, which was comparable to SB203580 pretreatment, and companied by the activation of autophagy and decreases of apoptosis when compared to BPA alone, indicating that melatonin protected against BPA-induced apoptosis partially through the p38 MAPK-autophagy pathway. In conclusion, these results suggest that melatonin may prevent BPA-induced FLK-BLV cell damage by inhibiting p38/MAPK signaling pathway and activating autophagy, and it could be a potential therapeutic compound in preventing BPA-induced cell damage.

Serum amyloid A3 is required for caerulein-induced acute pancreatitis through induction of RIP3-dependent necroptosis.

Serum amyloid A (SAA) is an early and sensitive biomarker of inflammatory diseases, but its role in acute pancreatitis (AP) is still unclear. Here, we used a caerulein-induced mouse model to investigate the role of SAA in AP and other related inflammatory responses. In our study, we found that the expression of a specific SAA isoform, SAA3, was significantly elevated in a caerulein-induced AP animal model. In addition, SAA3-knockout (Saa3-/- ) mice showed lower serum levels of amylase and lipase, tissue damage and proinflammatory cytokine production in the pancreas compared with those of wild-type mice in response to caerulein administration. AP-associated acute lung injury was also significantly attenuated in Saa3-/- mice. In our in vitro experiments, treatment with cholecystokinin and recombinant SAA3 significantly induced necroptosis and cytokine production. Moreover, we found that the regulatory effect of SAA3 on acinar cell necroptosis was through a receptor-interacting protein 3 (RIP3)-dependent manner. Collectively, our findings indicate that SAA3 is required for AP by inducing an RIP3-dependent necroptosis pathway in acinar cells and is a potential drug target for AP.

A novel posttranslational modification of histone, H3 S-sulfhydration, is down-regulated in asthenozoospermic sperm.

Oxidative stress is one of the major causes leading to male infertility including asthenozoospermia. Hydrogen sulfide (H2S) has been widely recognized to be a potent antioxidant whose role is partially implemented by protein S-sulfhydration. However, protein S-sulfhydration has not been reported in germ cells. Therefore, we investigated whether asthenozoospermia could be associated with sperm protein S-sulfhydration. S-sulfhydrated proteins in human sperm were enriched via biotin-switch assay and analyzed using LC-MS/MS spectrometry. Two hundred forty-four S-sulfhydrated proteins were identified. Importantly, we validated that sperm histones H3.1 and H3.3 were the S-sulfhydrated proteins. Their S-sulfhydrated amino acid residue was Cysteine111. Abundances of S-sulfhydrated H3 (sH3) and S-sulfhydrated H3.3 (sH3.3) were significantly down-regulated in asthenozoospermic sperm, compared with the fertile controls, and were significantly correlated with progressive motility. Retinoic acid (RA) up-regulated level of sH3.3 in primary round spermatids and the C18-4 cells (a mouse spermatogonial stem cell line). Overexpression of the mutant H3.3 (Cysteine111 was replaced with serine) affected expression of 759 genes and raised growth rate of C18-4 cells. For the first time, S-sulfhydration H3 and H3.3 were demonstrated in the present study. Our results highlight that aberrant S-sulfhydration of H3 is a new pathophysiological basis in male infertility.

Signatures of Divergence, Invasiveness, and Terrestrialization Revealed by Four Apple Snail Genomes.

The family Ampullariidae includes both aquatic and amphibious apple snails. They are an emerging model for evolutionary studies due to the high diversity, ancient history, and wide geographical distribution. Insight into drivers of ampullariid evolution is hampered, however, by the lack of genomic resources. Here, we report the genomes of four ampullariids spanning the Old World (Lanistes nyassanus) and New World (Pomacea canaliculata, P. maculata, and Marisa cornuarietis) clades. The ampullariid genomes have conserved ancient bilaterial karyotype features and a novel Hox gene cluster rearrangement, making them valuable in comparative genomic studies. They have expanded gene families related to environmental sensing and cellulose digestion, which may have facilitated some ampullarids to become notorious invasive pests. In the amphibious Pomacea, novel acquisition of an egg neurotoxin and a protein for making the calcareous eggshell may have been key adaptations enabling their transition from underwater to terrestrial egg deposition.

Association between prenatal bisphenol a exposure and promoter hypermethylation of CAPS2, TNFRSF25, and HKR1 genes in cord blood.

In utero exposure to bisphenol A (BPA) in early stages of development has been reported to exert adverse health effects on offspring later in life. Epigenetic alterations, particularly DNA methylation, may be one plausible biological mechanism involved. We examined the association between maternal BPA exposure and DNA methylation in cord blood. We randomly selected 96 paired samples of maternal urine and infant cord blood collected from the Shanghai-Minhang Birth Cohort. BPA levels in maternal urine were measured using high-performance liquid chromatography (HPLC). Three cord blood samples with maternal BPA levels >2.0 µg/g Cr and three samples with undetected BPA were randomly selected for genome-wide methylation analysis using methylated DNA binding domain sequencing (MBD-Seq). The genes with hypermethylated promoter regions were chosen for validation using quantitative methylation-specific polymerase chain reaction (Q-MSP). Based on MBD-seq results, we observed that maternal BPA exposure was primarily associated with hypermethylation of genes involved in signal transduction in the nervous system. Using Q-MSP, we further validated the association between maternal BPA exposure and promoter hypermethylation of three genes in multiple linear regression models: a log unit increase in BPA was associated with 12.63% (95%CI: 7.99, 17.26), 11.17%, (95%CI: 3.31, 19.02), and 16.57% (95% CI: 10.59, 22.56) increase in promoter of CAPS2, TNFRSF25, and HKR1 methylation, respectively. Our findings provide evidence that in utero exposure to BPA could alter the offspring's epigenome by altering DNA methylation pattern.

Genomic basis of recombination suppression in the hybrid between Caenorhabditis briggsae and C. nigoni.

DNA recombination is required for effective segregation and diversification of genomes and for the successful completion of meiosis. Recent studies in various species hybrids have demonstrated a genetic link between DNA recombination and speciation. Consistent with this, we observed a striking suppression of recombination in the hybrids between two nematodes, the hermaphroditic Caenorhabditis briggsae and the gonochoristic C. nigoni. To unravel the molecular basis underlying the recombination suppression in their hybrids, we generated a C. nigoni genome with chromosome-level contiguity and produced an improved C. briggsae genome with resolved gaps up to 2.8 Mb. The genome alignment reveals not only high sequence divergences but also pervasive intra- and inter-chromosomal sequence re-arrangements between the two species, which are plausible culprits for the observed suppression. Comparison of recombination boundary sequences suggests that recombination in the hybrid requires extensive sequence homology, which is rarely seen between the two genomes. The new genomes and genomic libraries form invaluable resources for studying genome evolution, hybrid incompatibilities and sex evolution for this pair of model species.

Chronic exposure to diesel exhaust particulate matter impairs meiotic progression during spermatogenesis in a mouse model.

Exposure to ambient PM2.5 may correlate with the decline of semen quality, and the underlying biological mechanism has not been fully understood. In the present study, mice were intratracheally instilled with diesel exhaust PM2.5 (DEP), and its effects on the spermatogenic process as well as the alterations of testicular gene expression profile were assessed. Our results showed that chronic exposure to DEP impaired the fertility of male mice without influencing their libido. Compared with Vehicle-exposed group, the sperm count and motility from DEP-exposed mice were significantly decreased. In addition, immunohistological staining of γH2AX and DMC1, biomarkers for meiotic double strand breaks (DSBs), demonstrated that chronic exposure to DEP comprised the repair of meiotic DSBs, thus disrupting the spermatogenesis. Deep RNA sequencing test showed altered expressions of testicular genes including the GnRH signaling pathway. In summary, our research demonstrated that chronic exposure to DEP may disrupt spermatogenesis through targeting the meiotic recombination, providing a new perspective for the research on the male reproductive system damage caused by air pollution.

MLN4924 protects against interleukin-17A-induced pulmonary inflammation by disrupting ACT1-mediated signaling.

An excessive inflammatory response in terminal airways, alveoli, and the lung interstitium eventually leads to pulmonary hypertension and chronic obstructive pulmonary disease. Proinflammatory cytokine interleukin-17A (IL-17A) has been implicated in the pathogenesis of pulmonary inflammatory diseases. MLN4924, an inhibitor of NEDD8-activating enzyme (NAE), is associated with the treatment of various types of cancers, but its role in the IL-17A-mediated inflammatory response has not been identified. Here, we report that MLN4924 can markedly reduce the expression of proinflammatory cytokines and chemokines such as IL-1ß, IL-6, and CXCL-1 and neutrophilia in a mouse model of IL-17A adenovirus-induced pulmonary inflammation. MLN4924 significantly inhibited IL-17A-induced stabilization of mRNA of proinflammatory cytokines and chemokines in vitro. Mechanistically, MLN4924 significantly blocked the activation of MAPK and NF-κB pathways and interfered with the interaction between ACT1 and tumor necrosis factor receptor-associated factor proteins (TRAFs), thereby inhibiting TRAF6 ubiquitination. Taken together, our data uncover a previously uncharacterized inhibitory effect of MLN4924 on the IL-17A-mediated inflammatory response; this phenomenon may facilitate the development of MLN4924 into an effective small-molecule drug for the treatment of pulmonary inflammatory diseases.

Specific down-regulation of spermatogenesis genes targeted by 22G RNAs in hybrid sterile males associated with an X-Chromosome introgression.

Hybrid incompatibility (HI) prevents gene flow between species, thus lying at the heart of speciation genetics. One of the most common HIs is male sterility. Two superficially contradictory observations exist for hybrid male sterility. First, an introgression on the X Chromosome is more likely to produce male sterility than on autosome (so-called large-X theory); second, spermatogenesis genes are enriched on the autosomes but depleted on the X Chromosome (demasculinization of X Chromosome). Analysis of gene expression in Drosophila hybrids suggests a genetic interaction between the X Chromosome and autosomes that is essential for male fertility. However, the prevalence of such an interaction and its underlying mechanism remain largely unknown. Here we examine the interaction in nematode species by contrasting the expression of both coding genes and transposable elements (TEs) between hybrid sterile males and its parental nematode males. We use two lines of hybrid sterile males, each carrying an independent introgression fragment from Caenorhabditis briggsae X Chromosome in an otherwise Caenorhabditis nigoni background, which demonstrate similar defects in spermatogenesis. We observe a similar pattern of down-regulated genes that are specific for spermatogenesis between the two hybrids. Importantly, the down-regulated genes caused by the X Chromosome introgressions show a significant enrichment on the autosomes, supporting an epistatic interaction between the X Chromosome and autosomes. We investigate the underlying mechanism of the interaction by measuring small RNAs and find that a subset of 22G RNAs specifically targeting the down-regulated spermatogenesis genes is significantly up-regulated in hybrids, suggesting that perturbation of small RNA-mediated regulation may contribute to the X-autosome interaction.