High Transcriptional Activity Research Articles

BACH1 deficiency prevents neointima formation and maintains the differentiated phenotype of vascular smooth muscle cells by regulating chromatin accessibility.

The transcription factor BTB and CNC homology 1(BACH1) has been linked to coronary artery disease risk by human genome-wide association studies, but little is known about the role of BACH1 in vascular smooth muscle cell (VSMC) phenotype switching and neointima formation following vascular injury. Therefore, this study aims to explore the role of BACH1 in vascular remodeling and its underlying mechanisms. BACH1 was highly expressed in human atherosclerotic plaques and has high transcriptional factor activity in VSMCs of human atherosclerotic arteries. VSMC-specific loss of Bach1 in mice inhibited the transformation of VSMC from contractile to synthetic phenotype and VSMC proliferation and attenuated the neointimal hyperplasia induced by wire injury. Mechanistically, BACH1 suppressed chromatin accessibility at the promoters of VSMC marker genes via recruiting histone methyltransferase G9a and cofactor YAP and maintaining the H3K9me2 state, thereby repressing VSMC marker genes expression in human aortic smooth muscle cells (HASMCs). BACH1-induced repression of VSMC marker genes was abolished by the silencing of G9a or YAP. Thus, these findings demonstrate a crucial regulatory role of BACH1 in VSMC phenotypic transition and vascular homeostasis and shed light on potential future protective vascular disease intervention via manipulation of BACH1.

Changes in the Expression of Genes, Associated with Calcium Processes, in the Hippocampus of Mice under the Influence of Chronic Social Defeat Stress

Using data from a complete transcriptome analysis, changes in the expression of genes encoding proteins involved in calcium regulation processes in the hippocampus of male mice with symptoms of depression caused by chronic social defeat stress were investigated. The expression of Cacna1g, Cacnb3, Camk1g, Camk2d, Camk2n2, Caly, Caln1, S100a16, Slc24a4 genes in the hippocampus of depressed mice was increased in relation to control, while the Cacna2d1, Cacng5, Grin2a, Calm2 gene expression was reduced. The greatest number of significant correlations between the expression of the gene Calm2, which has the highest transcriptional activity, and other differentially expressed genes was revealed. It is assumed that calcium signaling in the hippocampus of mice is disrupted under the influence of chronic social defeat stress. The involvement of the Calm2 gene and the Сamk1g, Camk2d and Camk2n2 genes in this process is discussed.

Structures of NF-κB p52 homodimer-DNA complexes rationalize binding mechanisms and transcription activation.

The mammalian NF-κB p52:p52 homodimer together with its cofactor Bcl3 activates transcription of κB sites with a central G/C base pair (bp), while it is inactive toward κB sites with a central A/T bp. To understand the molecular basis for this unique property of p52, we have determined the crystal structures of recombinant human p52 protein in complex with a P-selectin(PSel)-κB DNA (5'-GGGGTGACCCC-3') (central bp is underlined) and variants changing the central bp to A/T or swapping the flanking bp. The structures reveal a nearly two-fold widened minor groove in the central region of the DNA as compared to all other currently available NF-κB-DNA complex structures, which have a central A/T bp. Microsecond molecular dynamics (MD) simulations of free DNAs and p52 bound complexes reveal that free DNAs exhibit distinct preferred conformations, and p52:p52 homodimer induces the least amount of DNA conformational changes when bound to the more transcriptionally active natural G/C-centric PSel-κB, but adopts closed conformation when bound to the mutant A/T and swap DNAs due to their narrowed minor grooves. Our binding assays further demonstrate that the fast kinetics favored by entropy is correlated with higher transcriptional activity. Overall, our studies have revealed a novel conformation for κB DNA in complex with NF-κB and pinpoint the importance of binding kinetics, dictated by DNA conformational and dynamic states, in controlling transcriptional activation for NF-κB.

Low-affinity CTCF binding drives transcriptional regulation whereas high-affinity binding encompasses architectural functions

CTCF is a DNA-binding protein which plays critical roles in chromatin structure organization and transcriptional regulation; however, little is known about the functional determinants of different CTCF-binding sites (CBS). Using a conditional mouse model, we have identified one set of CBSs that are lost upon CTCF depletion (lost CBSs) and another set that persists (retained CBSs). Retained CBSs are more similar to the consensus CTCF-binding sequence and usually span tandem CTCF peaks. Lost CBSs are enriched at enhancers and promoters and associate with active chromatin marks and higher transcriptional activity. In contrast, retained CBSs are enriched at TAD and loop boundaries. Integration of ChIP-seq and RNA-seq data has revealed that retained CBSs are located at the boundaries between distinct chromatin states, acting as chromatin barriers. Our results provide evidence that transient, lost CBSs are involved in transcriptional regulation, whereas retained CBSs are critical for establishing higher-order chromatin architecture.

A Semi-mechanistic Model to Characterize the Long-Term Dynamics of Hepatitis B Virus Markers During Treatment With Lamivudine and Pegylated Interferon.

Antiviral treatments against hepatitis B virus (HBV) suppress viral replication but do not eradicate the virus, and need therefore to be taken lifelong to avoid relapse. Mathematical models can be useful to support the development of curative anti-HBV agents; however, they mostly focus on short-term HBV DNA data and neglect the complex host-pathogen interaction. This work aimed to characterize the effect of treatment with lamivudine and/or pegylated interferon (Peg-IFN) in 1,300 patients (hepatitis B envelope antigen (HBeAg)-positive and HBeAg-negative) treated for 1 year. A mathematical model was developed incorporating two populations of infected cells, namely , with a high transcriptional activity, that progressively evolve into , at a rate , representing cells with integrated HBV DNA that have a lower transcriptional activity. Parameters of the model were estimated in patients treated with lamivudine or Peg-IFN alone (N=894), and the model was then validated in patients treated with lamivudine plus Peg-IFN (N=436) to predict the virological response after a year of combination treatment. Lamivudine had a larger effect in blocking viral production than Peg-IFN (99.4-99.9% vs. 91.8-95.1%); however, Peg-IFN had a significant immunomodulatory effect, leading to an enhancement of the loss rates of (×1.7 in HBeAg-positive patients), (> ×7 irrespective of HBeAg status), and (×4.6 and ×2.0 in HBeAg-positive and HBeAg-negative patients, respectively). Using this model, we were able to describe the synergy of the different effects occurring during treatment with combination and predicted an effect of 99.99% on blocking viral production. This framework can therefore support the optimization of combination therapy with new anti-HBV agents.

SOX2-high cancer cells exhibit an aggressive phenotype, with increases in stemness, proliferation and invasion, as well as higher metabolic activity and ATP production.

Cancer stem cells (CSCs) are responsible for cancer recurrence, treatment failure and metastatic dissemination. As such, the elimination of CSCs represents one of the most important approaches for the future of cancer treatment. Among other properties, CSCs show the activation of particular cell signalling pathways and the over-expression of certain transcription factors, such as SOX2. Herein, we describe a new model system to isolate stem-like cancer cells, based on the functional transcriptional activity of SOX2. Briefly, we employed a SOX2-enhancer-GFP-reporter system to isolate cancer cells with high SOX2 transcriptional activity by FACS sorting. The over-expression of SOX2 in this sub-population was validated by Western blot analysis and flow cytometry. SOX2-high cancer cells showed CSCs features, such as greater mammosphere forming ability, validating that this sub-population was enriched in CSCs. To further explore the model, we analysed other stemness characteristics in MCF7 and MDA-MB-231 breast cancer cell lines, corroborating that SOX2-high cells were more metabolically active, proliferative, migratory, invasive, and drug-resistant. SOX2-high MDA-MB-231 cells also showed a loss of E-cadherin expression, and increased Vimentin expression, consistent with an epithelial-mesenchymal transition (EMT). Therefore, endogenous SOX2 transcriptional activity and protein levels are mechanistically linked to aggressive phenotypic behaviours and energy production in CSCs.

Zinc Finger Protein BCL11A Contributes to the Abortive Infection of Hirame novirhabdovirus (HIRRV) in B Lymphocytes of Flounder (Paralichthys olivaceus).

Hirame novirhabdovirus (HIRRV) infection is characterized by a pronounced viremia, and the high viral load is typically detected in immune-related organs and the circulatory system. In the present study, we demonstrated that HIRRV has the capacity to invade part of flounder membrane-bound IgM (mIgM+) B lymphocyte. Eight quantitative real-time PCR (qRT-PCR) standard curves involving HIRRV genomic RNA (gRNA), cRNA, and six mRNAs were established based on the strand-specific reverse transcription performed with tagged primers. It was revealed that viral RNA synthesis, especially the replication of gRNA, was inhibited in B cells, and the intracellular HIRRV even failed to produce infectious viral particles. Moreover, a range of genes with nucleic acid binding activity or related to viral infection were screened out based on the transcriptome analysis of HIRRV-infected B cells, and five molecules were further selected because of their different expression patterns in HIRRV-infected B cells and hirame natural embryo (HINAE) cells. The overexpression of these genes followed by HIRRV infection and RNA binding protein immunoprecipitation (RIP) assay revealed that the flounder B cell lymphoma/leukemia 11A (BCL11A), a highly conserved zinc finger transcription factor, is able to inhibit the proliferation of HIRRV by binding with full-length viral RNA mainly via its zinc finger domains at the C terminus. In conclusion, these data indicated that the high transcriptional activity of BCL11A in flounder mIgM+ B lymphocytes is a crucial factor for the abortive infection of HIRRV, and our findings provide new insights into the interaction between HIRRV and teleost B cells. IMPORTANCE HIRRV is a fish rhabdovirus that is considered as an important pathogen threatening the fish farming industry represented by flounder because of its high infectivity and fatality rate. To date, research toward understanding the complex pathogenic mechanism of HIRRV is still in its infancy and faces many challenges. Exploration of the relationship between HIRRV and its target cells is interesting and necessary. Here, we revealed that flounder mIgM+ B cells are capable of suppressing viral RNA synthesis and result in an unproductive infection of HIRRV. In addition, our results demonstrated that zinc finger protein BCL11A, a transcription factor in B cells, is able to suppress the replication of HIRRV. These findings increased our understanding of the underlying characteristics of HIRRV infection and revealed a novel antiviral mechanism against HIRRV based on the host restriction factor in teleost B cells, which sheds new light on the research into HIRRV control.

Analyses of widely targeted metabolic profiling reveals mechanisms of metabolomic variations during Tibetan sheep (Ovis aries) testis development

In mammals, the testis is the organ with the highest transcriptional activity. After gene transcription, translation, and post-translational protein modification, the transcriptional results are finally presented at the metabolic level. Metabolites not only essential for cell signaling and energy transfer, but also directly influenced by the physiological and pathological changes in tissues and accurately reflect the physiological changes. The fact that the testes are oxygen-deprived organs can explain why Sertoli cells and germ cells may use distinctive metabolic pathways to obtain energy in their different stages of development. Therefore, studying metabolic changes during testis development can better elucidate metabolic profile of the testis, which is essential to revealing characteristic metabolic pathways. The present study applied a widely targeted UPLC-MS/MS-based metabolomics approach with large-scale detection, identification and quantification to investigate the widespread metabolic changes during Tibetan sheep testis development. Firstly, a total of 847 metabolites were detected in the sheep testis, and their changes along with the three testis-development stages were further investigated. The results indicated that those metabolites were clustered into amino acids and their derivatives, carbohydrates and their derivatives, organic acids and their derivatives, benzene and substituted derivatives, alcohols and amines, lipids, nucleotides and their derivatives, bile acids, coenzymes and vitamins, hormones and hormone-related compounds, etc. Among them, the most abundant metabolites in the testis were amino acids and lipid metabolites. The results showed that most of the lipids, carbohydrates with their derivatives, as well as alcohol and amines metabolites were high in sexually immature sheep while organic acids, amino acids and nucleotides showed a continuously increasing trend along with testis development stages. Among them, the content of metabolites with antioxidant effects increased along with testis development, while those related with energy synthesis was downregulated with age. Further correlation analyses of each metabolite–metabolite pair emphasized the cross talk between differential metabolisms across testis development, suggesting a significant correlation between lipids and other metabolites. Finally, based on KEGG pathway analysis, we found that the metabolic pathways in Tibetan sheep testis development were mainly clustered into energy metabolism, gonadal development, and anti-oxidative stress. Reactive oxygen species (ROS) are by-products of normal cellular metabolism and are inevitable during testicular energy metabolism. Thus, the anti-oxidative stress function is a key process in maintaining the normal physiological function of testis. These results contributed to a broader view of the testis metabolome and a comprehensive analysis on metabolomic variation among different testis-development stages, providing a theoretical basis for us to understand the sheep testis metabolic mechanism.

Genetic adaptations to potato starch digestion in thePeruvian Andes

AbstractObjectivesPotatoes are an important staple crop across the world and particularly in the Andes, where they were cultivated as early as 10,000 years ago. Ancient Andean populations that relied upon this high‐starch food to survive could possess genetic adaptation(s) to digest potato starch more efficiently. Here, we analyzed genomic data to identify whether this putative adaptation is still present in their modern‐day descendants, namely Peruvians of Indigenous American ancestry.Materials and methodsWe applied several tests to detect signatures of natural selection in genes associated with starch‐digestion,AMY1,AMY2,SI, andMGAMin Peruvians. These were compared to two populations who only recently incorporated potatoes into their diets, Han Chinese and West Africans.ResultsOverlapping statistical results identified a regional haplotype inMGAMthat is unique to Peruvians. The age of this haplotype was estimated to be around 9547 years old.DiscussionTheMGAMhaplotype in Peruvians lies within a region of high transcriptional activity associated with the REST protein. The timing of this haplotype suggests that it arose in response to increased potato cultivation and attendant consumption. For Peruvian populations that relied upon the high‐starch potato as a major source of nutrition, natural selection likely favored theseMGAMvariant(s) that led to more efficient digestion and increased glucose production. This research provides further support that detecting subtle shifts in human diet can be a major driver of human evolutionary change, as these results indicate that there is global variation in human ability to better digest high‐starch foods.

Cellular Plasticity As Mechanisms to Escape from NOTCH1-Inhibition in T-ALL

Cellular plasticity has recently emerged as an underlying mechanism for treatment refraction in cancer. Hence, characterizing cell states and the transcriptional changes required for state transitions is critical for the development of efficient targeted therapies. Acute lymphoblastic leukemia (T-ALL) is an aggressive hematopoietic malignancy in children and adolescents that is associated with high rates of treatment failure and early relapse. T-ALL patients frequently harbor NOTCH1 activating mutations as the driving oncogene in this disease. Using T-ALL patient-derived xenograft (PDX) models carrying activating NOTCH1 mutations, we aim to u First, to determine how cell state diversity drives treatment refraction in T-ALL, we performed full-length single-cell transcriptome analysis on 3188 immune cells collected from 2 sensitive and 3 refractory PDX models. PDX models were treated with either with NOTCH inhibitor DBZ (Dipenzazepine; 10 μM/kg every other day intravenously) or vehicle. Profiled cells were collected at both a short (5 days) and a late time point (x-150 days) post-treatment to assess the effects of DBZ. Analysis of early hematopoietic and thymic precursor signatures demonstrate expression of immature hematopoietic signatures and acquisition of alternative lineage identities coexisting within the same cell, predominantly in refractory models. Analyses of pre-treatment samples indicated that disruption of the normal developmental hierarchy was associated with blockade at an early stage of T-cell commitment and can predict response to NOTCH inhibition in vivo. Upon treatment, cells underwent dramatic transcriptional reprogramming resulting in expansion of 2 distinct populations that greatly differ in both differentiation stage and developmental trajectory. One population expresses lymphoid differentiation programs, whereas the other population is enriched in genes associated with RNA processing and more immature hematopoietic signatures. Cell fate trajectories assessment using Monocle 2, revealed that consistent with their immature hematopoietic signature, refractory cells map at the start point of the differentiation trajectory, while sensitive cells clustered at the endpoint. We next sought to characterize the differentiation hierarchies that define the immature states. Developmental potential as assessed by CytoTRACE was higher in refractory cells when compared to their sensitive counterpart. To identify determinants of cell fate in a functionally relevant manner, we inferred gene regulatory network configurations using SCENIC and defined state specific regulon activity. Among the most prominently expressed regulons present in mature states, we identified transcriptional regulators involved in T-cell differentiation such as IKAROS, GATA3, and SOX5, while in immature states we observed high transcriptional regulon activity in transcription factors associated with renewal of hematopoietic progenitors such as ATF4, BCLAF1, and MYC in addition to the chromatin remodelers SMARCA4 and EZH2. To assess whether transcriptional rewiring in refractory cells results from greater chromatin accessibility, we performed single-cell ATAC sequencing (10x Genomics) on splenic cells from 1 sensitive and 1 refractory T-ALL PDX model treated with DBZ or vehicle. After integration with corresponding single-cell RNA-sequencing data, annotated ATAC profiles demonstrated greater chromatin accessibility, predominantly in active promotor and enhancer regions, in the refractory model compared to the sensitive model. Peak to gene linkage analysis revealed widespread alterations in cis-regulatory regions near genes associated with immature hematopoietic precursor signatures such as CD33. Furthermore, greater chromatin accessibility in refractory cells was associated with an increased number of enhancer loops per expressed gene as determined by H3K27ac HiChIP analysis. In conclusion, we demonstrated that presence of highly plastic cellular states defined by aberrant differentiation trajectories, distinct transcriptional circuitries, and remodeled chromatin architecture results in treatment escape in T-ALL.

Genome-wide map of R-loops reveals its interplay with transcription and genome integrity during germ cell meiosis

IntroductionThe R-loop is a naturally formed three-strand nucleic acid structure that recently has been reported to participate in multiple biological processes and helped answer some previously unexplained scientific questions. Meiosis process involves multiple chromatin-related events such as DNA double-stranded breaks (DSB) formation, repairing and transcriptional dynamics. ObjectivesExplore the regulatory roles and physiological functions of R-loops in the mammalian meiosis process. MethodsIn our study, using genome-wide S9.6 CUT & Tag seq, we first mapped the genomic distribution and dynamic changes of R-loop during the meiotic process in mice, from spermatogonia to secondary spermatocytes. And we further explore the role of R-loop in physiological conditions by constructing conditional knockout mice of Rnaseh1, which deleted the R-loop endonuclease before meiosis entry. ResultsR-loop predominantly distributes at promoter-related regions and varies across different meiotic stages. By joint analysis with the corresponding transcriptome, we found that the R-loop was closely related to transcription during the meiotic process. The high frequency of promoter-related R-loop in meiotic cells is usually accompanied by high transcription activity, and we further verified this in the leptotene/zygotene to the pachytene transition process. Moreover, the lack of RNase H1 caused sterility in male mice with R-loop accumulation and abnormal DSB repair in spermatocytes. Further analysis showed that abnormal R-loop accumulation in the leptotene/zygotene stages influenced transcriptional regulation in the pachytene stage. ConclusionThe mutual regulation of the R-loop and transcription plays an essential role in spermatogenesis. And R-loop is also important for the normal repair process of DSB during meiosis.

Frequent Occurrence and Metabolic Versatility of Marinifilaceae Bacteria as Key Players in Organic Matter Mineralization in Global Deep Seas.

Transfer of animal and plant detritus of both terrestrial and marine origins to the deep sea occurs on a global scale. Microorganisms play an important role in mineralizing them therein, but these are yet to be identified in situ. To observe key bacteria involved, we conducted long-term in situ incubation and found that members of the family Marinifilaceae (MF) occurred as some of the most predominant bacteria thriving on the new inputs of plant and animal biomasses in the deep sea in both marginal and oceanic areas. This taxon is diverse and ubiquitous in marine environments. A total of 11 MAGs belonging to MF were retrieved from metagenomic data and diverged into four subgroups in the phylogenomic tree. Based on metagenomic and metatranscriptomic analyses, we described the metabolic features and in situ metabolizing activities of different subgroups. The MF-2 subgroup, which dominates plant detritus-enriched cultures, specializes in polysaccharide degradation and lignin oxidation and has high transcriptional activities of related genes in situ. Intriguingly, members of this subgroup encode a nitrogen fixation pathway to compensate for the shortage of nitrogen sources inside the plant detritus. In contrast, other subgroups dominating the animal tissue-supported microbiomes are distinguished from MF-2 with regard to carbon and nitrogen metabolism and exhibit high transcriptional activity for proteolysis in situ. Despite these metabolic divergences of MF lineages, they show high in situ transcriptional activities for organic fermentation and anaerobic respiration (reductions of metal and/or dimethyl sulfoxide). These results highlight the role of previously unrecognized Marinifilaceae bacteria in organic matter mineralization in marine environments by coupling carbon and nitrogen cycling with metal and sulfur. IMPORTANCE Microbial mineralization of organic matter has a significant impact on the global biogeochemical cycle. This report confirms the role of Marinifilaceae in organic degradation in the oceans, with a contribution to ocean carbon cycling that has previously been underestimated. It was the dominant taxon thriving on plant and animal biomasses in our in situ incubator, as well as in whale falls and wood falls. At least 9 subgroups were revealed, and they were widely distributed in oceans globally but predominant in organic-matter-rich environments, with an average relative abundance of 8.3%. Different subgroups display a preference for the degradation of different macromolecules (polysaccharides, lignin, and protein) and adapt to their environments via special metabolic mechanisms.

Two Novel Functional Mutations in Promoter Region of SCN3B Gene Associated with Atrial Fibrillation.

The sodium voltage-gated channel beta subunit 3 (SCN3B) plays a crucial role in electrically excitable cells and conduction tissue in the heart. Some previous studies have established that genetic modification in sodium voltage-channel genes encoding for the cardiac β-subunits, such as SCN1B, SCN2B, SCN3B and SCN4B, can result in atrial fibrillation (AF). In the current study, we identified two rare variants in 5'UTR (NM_018400.4: c.-324C>A, rs976125894 and NM_018400.4: c.-303C>T, rs1284768362) of SCN3B in two unrelated lone AF patients. Our further functional studies discovered that one of them, the A allele of c.-324C>A (rs976125894), can improve transcriptional activity and may raise SCN3B expression levels. The A allele of c.-324C>A (rs976125894) has higher transcriptional activity when it interacts with GATA4, as we confirmed transcription factor GATA4 is a regulator of SCN3B. To the best of our knowledge, the current study is the first to demonstrate that the gain-of-function mutation of SCN3B can produce AF and the first to link a mutation occurring in the non-coding 5'UTR region of SCN3B to lone AF. The work also offers empirical proof that GATA4 is a critical regulator of SCN3B gene regulation. Our findings may serve as an encyclopedia for AF susceptibility variants and can also provide insight into the investigation of the functional mechanisms behind AF variants discovered by genetic methods.

A Single Nucleotide Polymorphism in the WIF1 Promoter Region Regulates the Wool Length in Rabbits

Wool length is an important trait in the rabbit wool industry. Wnt inhibitory factor 1 (WIF1), a key gene, regulates hair follicle growth and development. This study explores the WIF1 gene relationship with wool production traits in three different rabbit breeds, Rex rabbit, New Zealand rabbit, and Angora rabbit. The rabbit WIF1 gene sequence was obtained from GenBank, and PCR and Sanger sequencing were applied to screen SNP (single nucleotide polymorphism) from the gene exons and promoter regions. Sequencing results found no SNP in the PCR products of 10 exons from the rabbit WIF1 gene, indicating that the WIF1 gene was not polymorphic. Thus, the gene was conserved in all three rabbit breeds. Only one SNP was found in the WIF1 promoter region, associated with wool-producing traits in rabbits. Luciferase activity assays showed that the AA genotype had higher transcriptional activities than the AG or GG genotype (p < 0.01). In addition, WIF1 was significantly upregulated in the Rex rabbit compared to the New Zealand rabbit and Angora rabbit (p < 0.01). In summary, a key SNP in the WIF1 promoter region was found to be associated with rabbit fiber length trait.

A grazing-driven positive nutrient feedback loop and active sexual reproduction underpin widespread Noctiluca green tides

The mixoplankton green Noctiluca scintillans (gNoctiluca) is known to form extensive green tides in tropical coastal ecosystems prone to eutrophication. In the Arabian Sea, their recent appearance and annual recurrence have upended an ecosystem that was once exclusively dominated by diatoms. Despite evidence of strong links to eutrophication, hypoxia and warming, the mechanisms underlying outbreaks of this mixoplanktonic dinoflagellate remain uncertain. Here we have used eco-physiological measurements and transcriptomic profiling to ascribe gNoctiluca’s explosive growth during bloom formation to the form of sexual reproduction that produces numerous gametes. Rapid growth of gNoctiluca coincided with active ammonium and phosphate release from gNoctiluca cells, which exhibited high transcriptional activity of phagocytosis and metabolism generating ammonium. This grazing-driven nutrient flow ostensibly promotes the growth of phytoplankton as prey and offers positive support successively for bloom formation and maintenance. We also provide the first evidence that the host gNoctiluca cell could be manipulating growth of its endosymbiont population in order to exploit their photosynthetic products and meet critical energy needs. These findings illuminate gNoctiluca’s little known nutritional and reproductive strategies that facilitate its ability to form intense and expansive gNoctiluca blooms to the detriment of regional water, food and the socio-economic security in several tropical countries.

Human papillomavirus-related neoplasia of the ocular adnexa.

Human papillomaviruses (HPV) are involved in approximately 5% of solid cancers worldwide. The mucosotropic genotypes infect the stratified epithelium of various locations, where persistent infection may lead to invasive carcinomas. While the causative role of HPV in certain anogenital and head and neck carcinomas is well established, the role of HPV in carcinomas arising in the mucosal membranes of the ocular adnexal tissue (the lacrimal drainage system and the conjunctiva) has been a topic of great uncertainty. Therefore, we conducted a series of studies to assess the correlation between HPV and carcinomas arising in the mucosa of the ocular adnexal tissue and characterize the clinical, histopathological, and genomic features of the tumors in the context of HPV status in a Danish nationwide cohort. We collected clinical and histopathological data and tumor specimens from patients with carcinomas of the conjunctiva and the lacrimal drainage system, and their potential precursors, identified in Danish nationwide registries. The HPV status of the tumors was determined by the combined use of HPV DNA polymerase chain reaction (PCR), HPV E6/E7 mRNA in-situ hybridization, and p16 immunohistochemistry. The genomic profile was investigated by high-throughput DNA sequencing targeting 523 cancer-relevant genes. The literature to date on carcinomas of the lacrimal drainage system and the conjunctiva was summarized. In the Danish cohort, 67% of all carcinomas of the lacrimal drainage system and 21% of all conjunctival carcinomas were HPV-positive. HPV16 was the most frequently implicated genotype. A full-thickness expression of the viral oncogenes E6 and E7 was evident in almost all HPV DNA-positive cases. The HPV-positive carcinomas of the conjunctiva and the lacrimal drainage system shared histopathological and genomic features distinct from their HPV-negative counterparts. The HPV-positive carcinomas were characterized by a non-keratinizing morphology, p16 overexpression, high transcriptional activity of HPV E6/E7, and frequent pathogenic variants in the PI3K-AKT signaling cascade. In contrast, the HPV-negative carcinomas were characterized by a keratinizing morphology, lack of p16 and E6/E7 expression, and frequent somatic pathogenic variants in TP53, CDKN2A, and RB1. Among the patients with conjunctival tumors, HPV positivity was associated with a younger age at diagnosis and a higher risk of recurrence. In conclusion, the results support an etiological role of HPV in a subset of conjunctival and LDS carcinomas and their precursor lesions. Our investigations have shown that the HPV-positive carcinomas of the ocular adnexa share genomic and phenotypic characteristics with HPV-positive carcinomas of other anatomical locations. Therefore, these patients may be eligible for inclusion in future basket trials and future treatment regimens tailored to the more frequently occurring HPV-positive carcinomas of other locations. Future research will further elucidate the diagnostic, prognostic, and predictive role of HPV in these carcinomas.

Niche differentiation and symbiotic association among ammonia/nitrite oxidizers in a full-scale rotating biological contactor

Although the distribution of ammonia/nitrite oxidizers had been profiled in different habitats, current understanding is still limited regarding their niche differentiation in the integrated biofilm reactors, the symbiotic associations of ammonia/nitrite oxidizers, as well as the parasitic interaction between viruses and those functional organisms involved in the nitrogen cycle. Here, the integrated metagenomics and metatranscriptomics are applied to profile the ammonia/nitrite oxidizers communities and transcriptional activities changes along the flowpath of a concatenated full-scale rotating biological contactor (RBC) (frontend Stage-A and backend Stage-B). 19 metagenome-assembled genomes (MAGs) of ammonia/nitrite oxidizers were recovered by using a hybrid assembly approach, including four ammonia-oxidizing bacteria (AOB), two ammonia-oxidizing archaea (AOA), two complete ammonia oxidation bacteria (comammox), eight nitrite-oxidizing bacteria (NOB), and three anaerobic ammonium oxidation bacteria (anammox). Diverse AOB and anammox dominated Stage-A and collectively contributed to nitrogen conversion. With the decline of ammonia concentration along the flowpath, comammox and AOA appeared and increased in relative abundance in Stage-B, accounting for 8.8% of the entire community at the end of this reactor, and their dominating role in nitrogen turnover was indicated by the high transcription activity of their corresponding function genes. Moreover, the variation in the abundance of viruses infecting ammonia and nitrite oxidizers suggests that viruses likely act as a biotic factor mediating ammonia/nitrite oxidizer populations. This study demonstrates that complex factors shaped niche differentiation and symbiotic associations of ammonia/nitrite oxidizers in the RBC and highlights the importance of RBCs as model systems for the investigation of biotic and abiotic factors affecting the composition of microbiomes.

In Situ Genomics and Transcriptomics of SAR202 Subclusters Revealed Subtle Distinct Activities in Deep-Sea Water.

Deep-sea water columns are enriched with SAR202 that may conduct detrital matter degradation. There are several subclusters in SAR202, but their subtle differences in geochemical cycles are largely unknown, particularly for their in situ activities in the marine deep zone. Deep-sea DNA/RNA samples obtained from 12 continuous time periods over two days by in situ nucleic acid collection apparatus were used to re-evaluate the ecological functions of each SAR202 subcluster at a depth of ~1000 m in the South China Sea (SCS). Phylogenomics of 32 new SAR202 genomes from the SCS and western Pacific revealed their distribution in five subclusters. Metatranscriptomics analysis showed that the subclusters II and III were the dominant SAR202 groups with higher transcriptional activities in the SCS deep-sea zone than other subclusters. The analyses of functional gene expression further indicated that SAR202 subclusters II and III might be involved in different metabolic pathways in the deep-sea environment. The SAR202 subcluster III might take part in the degradation of deep-sea aromatic compounds. Time-course metagenomics and metatranscriptomics data did not show metabolic correlation of subclusters II and III over two days, suggesting diversified ecological functions of SAR202 subclusters under different organic inputs from the overlying water column. Collectively, our results indicate that the SAR202 subclusters play different roles in organic degradation and have probably undergone subtle and gradual adaptive evolution in the dynamic environment of the deep ocean.

Tumor treating fields affect mesothelioma cell proliferation by exerting histotype-dependent cell cycle checkpoint activations and transcriptional modulations

Although clinical antitumor activity of Tumor Treating Fields (TTFields) has been reported in malignant pleural mesothelioma (MPM) patients, the mechanisms behind the different selectivity displayed by the various MPM histotypes to this physical therapy has not been elucidated yet. Taking advantage of the development of well characterized human MPM cell lines derived from pleural effusion and/or lavages of patients’ thoracic cavity, we investigated the biological effects of TTFields against these cells, representative of epithelioid, biphasic, and sarcomatoid histotypes. Growth inhibition and cell cycle perturbations caused by TTFields were investigated side by side with RNA-Seq analyses at different exposure times to identify pathways involved in cell response to treatment. We observed significant differences of response to TTFields among the cell lines. Cell cycle analysis revealed that the most sensitive cells (epithelioid CD473) were blocked in G2M phase followed by formation of polyploid cells. The least sensitive cells (sarcomatoid CD60) were only slightly affected by TTFields with a general delay in all cell cycle phases. Apoptosis was present in all samples, but while epithelioid cell death was already observed during the first 24 h of treatment, sarcomatoid cells needed longer times before they engaged apoptotic pathways. RNA-Seq experiments demonstrated that TTFields induced a transcriptional response already detectable at early time points (8 h). The number of differentially expressed genes was higher in CD473 than in CD60 cells, involving several pathways, such as those pertinent to cell cycle checkpoints, DNA repair, and histone modifications. Our data provide further support to the notion that the antitumor effects of TTFields are not simply related to a non-specific reaction to a physical stimulus, but are dependent on the biological background of the cells and the particular sensitivity to TTFields observed in epithelioid MPM cells is associated with a higher transcriptional activity than that observed in sarcomatoid models.

Genome-wide analysis of long terminal repeat retrotransposons from the cranberry Vaccinium macrocarpon

BACKGROUND: Long terminal repeat (LTR) retrotransposons are widespread in plant genomes and play a large role in the generation of genomic variation. Despite this, their identification and characterization remains challenging, especially for non-model genomes. Hence, LTR retrotransposons remain undercharacterized in Vaccinium genomes, although they may be beneficial for current berry breeding efforts. OBJECTIVE: Exemplarily focusing on the genome of American cranberry (Vaccinium macrocarpon Aiton), we aim to generate an overview of the LTR retrotransposon landscape, highlighting the abundance, transcriptional activity, sequence, and structure of the major retrotransposon lineages. METHODS: Graph-based clustering of whole genome shotgun Illumina reads was performed to identify the most abundant LTR retrotransposons and to reconstruct representative in silico full-length elements. To generate insights into the LTR retrotransposon diversity in V. macrocarpon, we also queried the genome assembly for presence of reverse transcriptases (RTs), the key domain of LTR retrotransposons. Using transcriptomic data, transcriptional activity of retrotransposons corresponding to the consensuses was analyzed. RESULTS: We provide an in-depth characterization of the LTR retrotransposon landscape in the V. macrocarpon genome. Based on 475 RTs harvested from the genome assembly, we detect a high retrotransposon variety, with all major lineages present. To better understand their structural hallmarks, we reconstructed 26 Ty1-copia and 28 Ty3-gypsy in silico consensuses that capture the detected diversity. Accordingly, we frequently identify association with tandemly repeated motifs, extra open reading frames, and specialized, lineage-typical domains. Based on the overall high genomic abundance and transcriptional activity, we suggest that retrotransposons of the Ale and Athila lineages are most promising to monitor retrotransposon-derived polymorphisms across accessions. CONCLUSIONS: We conclude that LTR retrotransposons are major components of the V. macrocarpon genome. The representative consensuses provide an entry point for further Vaccinium genome analyses and may be applied to derive molecular markers for enhancing cranberry selection and breeding.