Comprehensive transcriptome analysis reveals altered mRNA splicing and post-transcriptional changes in the aged mouse brain.

A comprehensive understanding of molecular changes during brain aging is essential to mitigate cognitive decline and delay neurodegenerative diseases. The interpretation of mRNA alterations during brain aging is influenced by the health and age of the animal cohorts studied. Here, we carefully consider these factors and provide an in-depth investigation of mRNA splicing and dynamics in the aging mouse brain, combining short- and long-read sequencing technologies with extensive bioinformatic analyses. Our findings encompass a spectrum of age-related changes, including differences in isoform usage, decreased mRNA dynamics and a module showing increased expression of neuronal genes. Notably, our results indicate a reduced abundance of mRNA isoforms leading to nonsense-mediated RNA decay and suggest a regulatory role for RNA-binding proteins, indicating that their regulation may be altered leading to the reshaping of the aged brain transcriptome. Collectively, our study highlights the importance of studying mRNA splicing events during brain aging.

PAPAS promotes differentiation of mammary epithelial cells and suppresses breast carcinogenesis.

Extensive remodeling of the female mammary epithelium during development and pregnancy has been linked to cancer susceptibility. The faithful response of mammary epithelial cells (MECs) to hormone signaling is key to avoiding breast cancer development. Here, we show that lactogenic differentiation of murine MECs requires silencing of genes encoding ribosomal RNA (rRNA) by the antisense transcript PAPAS. Accordingly, knockdown of PAPAS derepresses rRNA genes, attenuates the response to lactogenic hormones, and induces malignant transformation. Restoring PAPAS levels in breast cancer cells reduces tumorigenicity and lung invasion and activates many interferon-regulated genes previously linked to metastasis suppression. Mechanistically, PAPAS transcription depends on R-loop formation at the 3' end of rRNA genes, which is repressed by RNase H1 and replication protein A (RPA) overexpression in breast cancer cells. Depletion of PAPAS and upregulation of RNase H1 and RPA in human breast cancer underpin the clinical relevance of our findings.

The Root-Colonizing Endophyte Piriformospora indica Supports Nitrogen-Starved Arabidopsis thaliana Seedlings with Nitrogen Metabolites.

The root-colonizing endophytic fungus Piriformospora indica promotes the root and shoot growth of its host plants. We show that the growth promotion of Arabidopsis thaliana leaves is abolished when the seedlings are grown on media with nitrogen (N) limitation. The fungus neither stimulated the total N content nor did it promote 15NO3- uptake from agar plates to the leaves of the host under N-sufficient or N-limiting conditions. However, when the roots were co-cultivated with 15N-labelled P. indica, more labels were detected in the leaves of N-starved host plants but not in plants supplied with sufficient N. Amino acid and primary metabolite profiles, as well as the expression analyses of N metabolite transporter genes suggest that the fungus alleviates the adaptation of its host from the N limitation condition. P. indica alters the expression of transporter genes, which participate in the relocation of NO3-, NH4+ and N metabolites from the roots to the leaves under N limitation. We propose that P. indica participates in the plant's metabolomic adaptation against N limitation by delivering reduced N metabolites to the host, thus alleviating metabolic N starvation responses and reprogramming the expression of N metabolism-related genes.

Identification of altered miRNAs and their targets in placenta accreta.

Placenta accreta spectrum (PAS) is one of the major causes of maternal morbidity and mortality worldwide with increasing incidence. PAS refers to a group of pathological conditions ranging from the abnormal attachment of the placenta to the uterus wall to its perforation and, in extreme cases, invasion into surrounding organs. Among them, placenta accreta is characterized by a direct adhesion of the villi to the myometrium without invasion and remains the most common diagnosis of PAS. Here, we identify the potential regulatory miRNA and target networks contributing to placenta accreta development. Using small RNA-Seq followed by RT-PCR confirmation, altered miRNA expression, including that of members of placenta-specific miRNA clusters (e.g., C19MC and C14MC), was identified in placenta accreta samples compared to normal placental tissues. In situ hybridization (ISH) revealed expression of altered miRNAs mostly in trophoblast but also in endothelial cells and this profile was similar among all evaluated degrees of PAS. Kyoto encyclopedia of genes and genomes (KEGG) analyses showed enriched pathways dysregulated in PAS associated with cell cycle regulation, inflammation, and invasion. mRNAs of genes associated with cell cycle and inflammation were downregulated in PAS. At the protein level, NF-κB was upregulated while PTEN was downregulated in placenta accreta tissue. The identified miRNAs and their targets are associated with signaling pathways relevant to controlling trophoblast function. Therefore, this study provides miRNA:mRNA associations that could be useful for understanding PAS onset and progression.

Draft genome assembly and sequencing dataset of the marine diatom Skeletonema cf. costatum RCC75.

Diatoms (Bacillariophyceae) are a major constituent of the phytoplankton and have a universally recognized ecological importance. Between 1,000 and 1,300 diatom genera have been described in the literature, but only 10 nuclear genomes have been published and made available to the public up to date. Skeletonema costatum is a cosmopolitan marine diatom, principally occurring in coastal regions, and is one of the most abundant members of the Skeletonema genus. Here we present a draft assembly of the Skeletonema cf. costatum RCC75 genome, obtained from PacBio and Illumina NovaSeq data. This dataset will expand the knowledge of the Bacillariophyceae genetics and contribute to the global understanding of phytoplankton's physiological, ecological, and environmental functioning.

Age-dependent expression changes of circadian system-related genes reveal a potentially conserved link to aging.

The circadian clock system influences the biology of life by establishing circadian rhythms in organisms, tissues, and cells, thus regulating essential biological processes based on the day/night cycle. Circadian rhythms change over a lifetime due to maturation and aging, and disturbances in the control of the circadian system are associated with several age-related pathologies. However, the impact of chronobiology and the circadian system on healthy organ and tissue aging remains largely unknown. Whether aging-related changes of the circadian system's regulation follow a conserved pattern across different species and tissues, hence representing a common driving force of aging, is unclear. Based on a cross-sectional transcriptome analysis covering 329 RNA-Seq libraries, we provide indications that the circadian system is subjected to aging-related gene alterations shared between evolutionarily distinct species, such as Homo sapiens, Mus musculus, Danio rerio, and Nothobranchius furzeri. We discovered differentially expressed genes by comparing tissue-specific transcriptional profiles of mature, aged, and old-age individuals and report on six genes (per2, dec2, cirp, klf10, nfil3, and dbp) of the circadian system, which show conserved aging-related expression patterns in four organs of the species examined. Our results illustrate how the circadian system and aging might influence each other in various tissues over a long lifespan and conceptually complement previous studies tracking short-term diurnal and nocturnal gene expression oscillations.

VIDHOP, viral host prediction with deep learning.

MOTIVATION: Zoonosis, the natural transmission of infections from animals to humans, is a far-reaching global problem. The recent outbreaks of Zikavirus, Ebolavirus and Coronavirus are examples of viral zoonosis, which occur more frequently due to globalization. In case of a virus outbreak, it is helpful to know which host organism was the original carrier of the virus to prevent further spreading of viral infection. Recent approaches aim to predict a viral host based on the viral genome, often in combination with the potential host genome and arbitrarily selected features. These methods are limited in the number of different hosts they can predict or the accuracy of the prediction. RESULTS: Here, we present a fast and accurate deep learning approach for viral host prediction, which is based on the viral genome sequence only. We tested our deep neural network (DNN) on three different virus species (influenza A virus, rabies lyssavirus and rotavirus A). We achieved for each virus species an AUC between 0.93 and 0.98, allowing highly accurate predictions while using only fractions (100-400 bp) of the viral genome sequences. We show that deep neural networks are suitable to predict the host of a virus, even with a limited amount of sequences and highly unbalanced available data. The trained DNNs are the core of our virus-host prediction tool VIrus Deep learning HOst Prediction (VIDHOP). VIDHOP also allows the user to train and use models for other viruses. AVAILABILITY AND IMPLEMENTATION: VIDHOP is freely available under https://github.com/flomock/vidhop. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

EpiDope: a deep neural network for linear B-cell epitope prediction.

MOTIVATION: By binding to specific structures on antigenic proteins, the so-called epitopes, B-cell antibodies can neutralize pathogens. The identification of B-cell epitopes is of great value for the development of specific serodiagnostic assays and the optimization of medical therapy. However, identifying diagnostically or therapeutically relevant epitopes is a challenging task that usually involves extensive laboratory work. In this study, we show that the time, cost and labor-intensive process of epitope detection in the lab can be significantly reduced using in silico prediction. RESULTS: Here, we present EpiDope, a python tool which uses a deep neural network to detect linear B-cell epitope regions on individual protein sequences. With an area under the curve between 0.67 ± 0.07 in the receiver operating characteristic curve, EpiDope exceeds all other currently used linear B-cell epitope prediction tools. Our software is shown to reliably predict linear B-cell epitopes of a given protein sequence, thus contributing to a significant reduction of laboratory experiments and costs required for the conventional approach. AVAILABILITYAND IMPLEMENTATION: EpiDope is available on GitHub (http://github.com/mcollatz/EpiDope). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Tissue-specific Gene Expression Changes Are Associated with Aging in Mice.

Aging is a complex process that can be characterized by functional and cognitive decline in an individual. Aging can be assessed based on the functional capacity of vital organs and their intricate interactions with one another. Thus, the nature of aging can be described by focusing on a specific organ and an individual itself. However, to fully understand the complexity of aging, one must investigate not only a single tissue or biological process but also its complex interplay and interdependencies with other biological processes. Here, using RNA-seq, we monitored changes in the transcriptome during aging in four tissues (including brain, blood, skin and liver) in mice at 9 months, 15 months, and 24 months, with a final evaluation at the very old age of 30 months. We identified several genes and processes that were differentially regulated during aging in both tissue-dependent and tissue-independent manners. Most importantly, we found that the electron transport chain (ETC) of mitochondria was similarly affected at the transcriptome level in the four tissues during the aging process. We also identified the liver as the tissue showing the largest variety of differentially expressed genes (DEGs) over time. Lcn2 (Lipocalin-2) was found to be similarly regulated among all tissues, and its effect on longevity and survival was validated using its orthologue in Caenorhabditis elegans. Our study demonstrated that the molecular processes of aging are relatively subtle in their progress, and the aging process of every tissue depends on the tissue's specialized function and environment. Hence, individual gene or process alone cannot be described as the key of aging in the whole organism.

Robustness during Aging-Molecular Biological and Physiological Aspects.

Understanding the process of aging is still an important challenge to enable healthy aging and to prevent age-related diseases. Most studies in age research investigate the decline in organ functionality and gene activity with age. The focus on decline can even be considered a paradigm in that field. However, there are certain aspects that remain surprisingly stable and keep the organism robust. Here, we present and discuss various properties of robust behavior during human and animal aging, including physiological and molecular biological features, such as the hematocrit, body temperature, immunity against infectious diseases and others. We examine, in the context of robustness, the different theories of how aging occurs. We regard the role of aging in the light of evolution.

Correction to: A miRNA catalogue and ncRNA annotation of the short-living fish Nothobranchius furzeri.

Following the publication of this article [1], the authors reported that the images of Figs. 1, 2 and 3 were published in the incorrect order, whereby they mismatch with their captions.

Conserved aging-related signatures of senescence and inflammation in different tissues and species.

Increasing evidence indicates that chronic inflammation and senescence are the cause of many severe age-related diseases, with both biological processes highly upregulated during aging. However, until now, it has remained unknown whether specific inflammation- or senescence-related genes exist that are common between different species or tissues. These potential markers of aging could help to identify possible targets for therapeutic interventions of aging-associated afflictions and might also deepen our understanding of the principal mechanisms of aging. With the objective of identifying such signatures of aging and tissue-specific aging markers, we analyzed a multitude of cross-sectional RNA-Seq data from four evolutionarily distinct species (human, mouse and two fish) and four different tissues (blood, brain, liver and skin). In at least three different species and three different tissues, we identified several genes that displayed similar expression patterns that might serve as potential aging markers. Additionally, we show that genes involved in aging-related processes tend to be tighter controlled in long-lived than in average-lived individuals. These observations hint at a general genetic level that affect an individual's life span. Altogether, this descriptive study contributes to a better understanding of common aging signatures as well as tissue-specific aging patterns and supplies the basis for further investigative age-related studies.

Identification of miRNAs and associated pathways regulated by Leukemia Inhibitory Factor in trophoblastic cell lines.

INTRODUCTION: Leukemia Inhibitory Factor (LIF) regulates behavior of trophoblast cells and their interaction with immune and endothelial cells. In vitro, trophoblast cell response to LIF may vary depending on the cell model. Reported differences in the miRNA profile of trophoblastic cells may be responsible for these observations. Therefore, miRNA expression was investigated in four trophoblastic cell lines under LIF stimulation followed by in silico analysis of altered miRNAs and their associated pathways. METHODS: Low density TaqMan miRNA assays were used to quantify levels of 762 mature miRNAs under LIF stimulation in three choriocarcinoma-derived (JEG-3, ACH-3P and AC1-M59) and a trophoblast immortalized (HTR-8/SVneo) cell lines. Expression of selected miRNAs was confirmed in primary trophoblast cells and cell lines by qPCR. Targets and associated pathways of the differentially expressed miRNAs were inferred from the miRTarBase followed by a KEGG Pathway Enrichment Analysis. HTR-8/SVneo and JEG-3 cells were transfected with miR-21-mimics and expression of miR-21 targets was assessed by qPCR. RESULTS: A similar number of miRNAs changed in each tested cell line upon LIF stimulation, however, low coincidence of individual miRNA species was observed and occurred more often among choriocarcinoma-derived cells (complete data set at http://www.ncbi.nlm.nih.gov/geo/ under GEO accession number GSE130489). Altered miRNAs were categorized into pathways involved in human diseases, cellular processes and signal transduction. Six cascades were identified as significantly enriched, including JAK/STAT and TGFB-SMAD. Upregulation of miR-21-3p was validated in all cell lines and primary cells and STAT3 was confirmed as its target. DISCUSSION: Dissimilar miRNA responses may be involved in differences of LIF effects on trophoblastic cell lines.

Effects of allelic variations in the human myxovirus resistance protein A on its antiviral activity.

Only a minority of patients infected with seasonal influenza A viruses exhibit a severe or fatal outcome of infection, but the reasons for this inter-individual variability in influenza susceptibility are unclear. To gain further insights into the molecular mechanisms underlying this variability, we investigated naturally occurring allelic variations of the myxovirus resistance 1 (MX1) gene coding for the influenza restriction factor MxA. The interferon-induced dynamin-like GTPase consists of an N-terminal GTPase domain, a bundle signaling element, and a C-terminal stalk responsible for oligomerization and viral target recognition. We used online databases to search for variations in the MX1 gene. Deploying in vitro approaches, we found that non-synonymous variations in the GTPase domain cause the loss of antiviral and enzymatic activities. Furthermore, we showed that these amino acid substitutions disrupt the interface for GTPase domain dimerization required for the stimulation of GTP hydrolysis. Variations in the stalk were neutral or slightly enhanced or abolished MxA antiviral function. Remarkably, two other stalk variants altered MxA's antiviral specificity. Variations causing the loss of antiviral activity were found only in heterozygous carriers. Interestingly, the inactive stalk variants blocked the antiviral activity of WT MxA in a dominant-negative way, suggesting that heterozygotes are phenotypically MxA-negative. In contrast, the GTPase-deficient variants showed no dominant-negative effect, indicating that heterozygous carriers should remain unaffected. Our results demonstrate that naturally occurring mutations in the human MX1 gene can influence MxA function, which may explain individual variations in influenza virus susceptibility in the human population.

A miRNA catalogue and ncRNA annotation of the short-living fish Nothobranchius furzeri.

BACKGROUND: The short-lived fish Nothobranchius furzeri is the shortest-lived vertebrate that can be cultured in captivity and was recently established as a model organism for aging research. Small non-coding RNAs, especially miRNAs, are implicated in age dependent control of gene expression. RESULTS: Here, we present a comprehensive catalogue of miRNAs and several other non-coding RNA classes (ncRNAs) for Nothobranchius furzeri. Analyzing multiple small RNA-Seq libraries, we show most of these identified miRNAs are expressed in at least one of seven Nothobranchius species. Additionally, duplication and clustering of N. furzeri miRNAs was analyzed and compared to the four fish species Danio rerio, Oryzias latipes, Gasterosteus aculeatus and Takifugu rubripes. A peculiar characteristic of N. furzeri, as compared to other teleosts, was a duplication of the miR-29 cluster. CONCLUSION: The completeness of the catalogue we provide is comparable to that of the zebrafish. This catalogue represents a basis to investigate the role of miRNAs in aging and development in this species.

Massive Effect on LncRNAs in Human Monocytes During Fungal and Bacterial Infections and in Response to Vitamins A and D.

Mycoses induced by C.albicans or A.fumigatus can cause important host damage either by deficient or exaggerated immune response. Regulation of chemokine and cytokine signaling plays a crucial role for an adequate inflammation, which can be modulated by vitamins A and D. Non-coding RNAs (ncRNAs) as transcription factors or cis-acting antisense RNAs are known to be involved in gene regulation. However, the processes during fungal infections and treatment with vitamins in terms of therapeutic impact are unknown. We show that in monocytes both vitamins regulate ncRNAs involved in amino acid metabolism and immune system processes using comprehensive RNA-Seq analyses. Compared to protein-coding genes, fungi and bacteria induced an expression change in relatively few ncRNAs, but with massive fold changes of up to 4000. We defined the landscape of long-ncRNAs (lncRNAs) in response to pathogens and observed variation in the isoforms composition for several lncRNA following infection and vitamin treatment. Most of the involved antisense RNAs are regulated and positively correlated with their sense protein-coding genes. We investigated lncRNAs with stimulus specific immunomodulatory activity as potential marker genes: LINC00595, SBF2-AS1 (A.fumigatus) and RP11-588G21.2, RP11-394l13.1 (C.albicans) might be detectable in the early phase of infection and serve as therapeutic targets in the future.

Differential transcriptional responses to Ebola and Marburg virus infection in bat and human cells.

The unprecedented outbreak of Ebola in West Africa resulted in over 28,000 cases and 11,000 deaths, underlining the need for a better understanding of the biology of this highly pathogenic virus to develop specific counter strategies. Two filoviruses, the Ebola and Marburg viruses, result in a severe and often fatal infection in humans. However, bats are natural hosts and survive filovirus infections without obvious symptoms. The molecular basis of this striking difference in the response to filovirus infections is not well understood. We report a systematic overview of differentially expressed genes, activity motifs and pathways in human and bat cells infected with the Ebola and Marburg viruses, and we demonstrate that the replication of filoviruses is more rapid in human cells than in bat cells. We also found that the most strongly regulated genes upon filovirus infection are chemokine ligands and transcription factors. We observed a strong induction of the JAK/STAT pathway, of several genes encoding inhibitors of MAP kinases (DUSP genes) and of PPP1R15A, which is involved in ER stress-induced cell death. We used comparative transcriptomics to provide a data resource that can be used to identify cellular responses that might allow bats to survive filovirus infections.