Browsing by Subject "TRANSCRIPTOME"

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  • PanScan PanC4 consortia; Walsh, Naomi; Zhang, Han; Männistö, Satu; Weiderpass, Elisabete (2019)
    Background Genome-wide association studies (GWAS) identify associations of individual single-nucleotide polymorphisms (SNPs) with cancer risk but usually only explain a fraction of the inherited variability. Pathway analysis of genetic variants is a powerful tool to identify networks of susceptibility genes. Methods We conducted a large agnostic pathway-based meta-analysis of GWAS data using the summary-based adaptive rank truncated product method to identify gene sets and pathways associated with pancreatic ductal adenocarcinoma (PDAC) in 9040 cases and 12 496 controls. We performed expression quantitative trait loci (eQTL) analysis and functional annotation of the top SNPs in genes contributing to the top associated pathways and gene sets. All statistical tests were two-sided. Results We identified 14 pathways and gene sets associated with PDAC at a false discovery rate of less than 0.05. After Bonferroni correction (P Conclusion Our agnostic pathway and gene set analysis integrated with functional annotation and eQTL analysis provides insight into genes and pathways that may be biologically relevant for risk of PDAC, including those not previously identified.
  • Nikparvar, Bahareh; Andreevskaya, Margarita; Duru, Ilhan C.; Bucur, Florentina I.; Grigore-Gurgu, Leontina; Borda, Daniela; Nicolau, Anca I.; Riedel, Christian U.; Auvinen, Petri; Bar, Nadav (2021)
    Background The pathogen Listeria (L.) monocytogenes is known to survive heat, cold, high pressure, and other extreme conditions. Although the response of this pathogen to pH, osmotic, temperature, and oxidative stress has been studied extensively, its reaction to the stress produced by high pressure processing HPP (which is a preservation method in the food industry), and the activated gene regulatory network (GRN) in response to this stress is still largely unknown. Results We used RNA sequencing transcriptome data of L. monocytogenes (ScottA) treated at 400 MPa and 8(circle)C, for 8 min and combined it with current information in the literature to create a transcriptional regulation database, depicting the relationship between transcription factors (TFs) and their target genes (TGs) in L. monocytogenes. We then applied network component analysis (NCA), a matrix decomposition method, to reconstruct the activities of the TFs over time. According to our findings, L. monocytogenes responded to the stress applied during HPP by three statistically different gene regulation modes: survival mode during the first 10 min post-treatment, repair mode during 1 h post-treatment, and re-growth mode beyond 6 h after HPP. We identified the TFs and their TGs that were responsible for each of the modes. We developed a plausible model that could explain the regulatory mechanism that L. monocytogenes activated through the well-studied CIRCE operon via the regulator HrcA during the survival mode. Conclusions Our findings suggest that the timely activation of TFs associated with an immediate stress response, followed by the expression of genes for repair purposes, and then re-growth and metabolism, could be a strategy of L. monocytogenes to survive and recover extreme HPP conditions. We believe that our results give a better understanding of L. monocytogenes behavior after exposure to high pressure that may lead to the design of a specific knock-out process to target the genes or mechanisms. The results can help the food industry select appropriate HPP conditions to prevent L. monocytogenes recovery during food storage.
  • Zhang, Sidi; Samocha, Kaitlin E.; Rivas, Manuel A.; Karczewski, Konrad J.; Daly, Emma; Schmandt, Ben; Neale, Benjamin M.; MacArthur, Daniel G.; Daly, Mark J. (2018)
    Variation in RNA splicing (i.e., alternative splicing) plays an important role in many diseases. Variants near 5' and 3' splice sites often affect splicing, but the effects of these variants on splicing and disease have not been fully characterized beyond the two "essential" splice nucleotides flanking each exon. Here we provide quantitative measurements of tolerance to mutational disruptions by position and reference allele-alternative allele combinations. We show that certain reference alleles are particularly sensitive to mutations, regardless of the alternative alleles into which they are mutated. Using public RNA-seq data, we demonstrate that individuals carrying such variants have significantly lower levels of the correctly spliced transcript, compared to individuals without them, and confirm that these specific substitutions are highly enriched for known Mendelian mutations. Our results propose a more refined definition of the "splice region" and offer a new way to prioritize and provide functional interpretation of variants identified in diagnostic sequencing and association studies.
  • Madissoon, Elo; Jouhilahti, Eeva-Mari; Vesterlund, Liselotte; Tohonen, Virpi; Krjutskov, Kaarel; Petropoulos, Sophie; Einarsdottir, Elisabet; Linnarssons, Sten; Lanner, Fredrik; Mansson, Robert; Hovatta, Outi; Burglin, Thomas R.; Katayama, Shintaro; Kere, Juha (2016)
    PAIRED (PRD)-like homeobox genes belong to a class of predicted transcription factor genes. Several of these PRD-like homeobox genes have been predicted in silico from genomic sequence but until recently had no evidence of transcript expression. We found recently that nine PRD-like homeobox genes, ARGFX, CPHX1, CPHX2, DPRX, DUXA, DUXB, NOBOX, TPRX1 and TPRX2, were expressed in human preimplantation embryos. In the current study we characterized these PRD-like homeobox genes in depth and studied their functions as transcription factors. We cloned multiple transcript variants from human embryos and showed that the expression of these genes is specific to embryos and pluripotent stem cells. Overexpression of the genes in human embryonic stem cells confirmed their roles as transcription factors as either activators (CPHX1, CPHX2, ARGFX) or repressors (DPRX, DUXA, TPRX2) with distinct targets that could be explained by the amino acid sequence in homeodomain. Some PRD-like homeodomain transcription factors had high concordance of target genes and showed enrichment for both developmentally important gene sets and a 36 bp DNA recognition motif implicated in Embryo Genome Activation (EGA). Our data implicate a role for these previously uncharacterized PRD-like homeodomain proteins in the regulation of human embryo genome activation and preimplantation embryo development.
  • Byers, Kelsey J. R. P.; Darragh, Kathy; Fernanda Garza, Sylvia; Abondano Almeida, Diana; Warren, Ian A.; Rastas, Pasi M. A.; Merrill, Richard M.; Schulz, Stefan; McMillan, W. Owen; Jiggins, Chris D. (2021)
    The degree to which loci promoting reproductive isolation cluster in the genome-that is, the genetic architecture of reproductive isolation-can influence the tempo and mode of speciation. Tight linkage between these loci can facilitate speciation in the face of gene flow. Pheromones play a role in reproductive isolation in many Lepidoptera species, and the role of endogenously produced compounds as secondary metabolites decreases the likelihood of pleiotropy associated with many barrier loci. Heliconius butterflies use male sex pheromones to both court females (aphrodisiac wing pheromones) and ward off male courtship (male-transferred antiaphrodisiac genital pheromones), and it is likely that these compounds play a role in reproductive isolation between Heliconius species. Using a set of backcross hybrids between H. melpomene and H. cydno, we investigated the genetic architecture of putative male pheromone compound production. We found a set of 40 significant quantitative trait loci (QTL) representing 33 potential pheromone compounds. QTL clustered significantly on two chromosomes, chromosome 8 for genital compounds and chromosome 20 for wing compounds, and chromosome 20 was enriched for potential pheromone biosynthesis genes. There was minimal overlap between pheromone QTL and known QTL for mate choice and color pattern. Nonetheless, we did detect linkage between a QTL for wing androconial area and optix, a color pattern locus known to play a role in reproductive isolation in these species. This tight clustering of putative pheromone loci might contribute to coincident reproductive isolating barriers, facilitating speciation despite ongoing gene flow.
  • Alonso Serra, Juan Antonio; Shi, Xueping; Peaucelle, Alexis; Rastas, Pasi; Bourdon, Matthieu; Immanen, Juha; Takahashi, Junko; Koivula, Hanna; Eswaran, Gugan; Muranen, Sampo Johannes; Help-Rinta-Rahko, Hanna; Smolander, Olli-Pekka; Su, Chang; Safronov, Omid; Gerber, Lorenz; Salojärvi, Jarkko; Hagqvist, Risto; Mähönen, Ari Pekka; Helariutta, Yrjö; Nieminen, Kaisa (2020)
    Tree architecture has evolved to support a top-heavy above-ground biomass, but this integral feature poses a weight-induced challenge to trunk stability. Maintaining an upright stem is expected to require vertical proprioception through feedback between sensing stem weight and responding with radial growth. Despite its apparent importance, the principle by which plant stems respond to vertical loading forces remains largely unknown. Here, by manipulating the stem weight of downy birch (Betula pubescens) trees, we show that cambial development is modulated systemically along the stem. We carried out a genetic study on the underlying regulation by combining an accelerated birch flowering program with a recessive mutation at the ELIMAKI locus (EKI), which causes a mechanically defective response to weight stimulus resulting in stem collapse after just 3 months. We observed delayed wood morphogenesis in eki compared with WT, along with a more mechanically elastic cambial zone and radial compression of xylem cell size, indicating that rapid tissue differentiation is critical for cambial growth under mechanical stress. Furthermore, the touch-induced mechanosensory pathway was transcriptionally misregulated in eki, indicating that the ELIMAKI locus is required to integrate the weight-growth feedback regulation. By studying this birch mutant, we were able to dissect vertical proprioception from the gravitropic response associated with reaction wood formation. Our study provides evidence for both local and systemic responses to mechanical stimuli during secondary plant development.
  • Sablok, Gaurav; Powell, Jonathan J.; Kazan, Kemal (2017)
    Plants use a wide range of mechanisms to adapt to different environmental stresses. One of the earliest responses displayed under stress is rapid alterations in stress responsive gene expression that has been extensively analyzed through expression profiling such as microarrays and RNA-sequencing. Recently, expression profiling has been complemented with proteome analyses to establish a link between transcriptional and the corresponding translational changes. However, proteome profiling approaches have their own technical limitations. More recently, ribosome-associated mRNA profiling has emerged as an alternative and a robust way of identifying translating mRNAs, which are a set of mRNAs associated with ribosomes and more likely to contribute to proteome abundance. In this article, we briefly review recent studies that examined the processes affecting the abundance of translating mRNAs, their regulation during plant development and tolerance to stress conditions and plant factors affecting the selection of translating mRNA pools. This review also highlights recent findings revealing differential roles of alternatively spliced mRNAs and their translational control during stress adaptation. Overall, better understanding of processes involved in the regulation of translating mRNAs has obvious implications for improvement of stress tolerance in plants.
  • Kuosmanen, Anna; Norri, Tuukka; Mäkinen, Veli (2018)
    Transcript prediction can be modeled as a graph problem where exons are modeled as nodes and reads spanning two or more exons are modeled as exon chains. Pacific Biosciences third-generation sequencing technology produces significantly longer reads than earlier second-generation sequencing technologies, which gives valuable information about longer exon chains in a graph. However, with the high error rates of third-generation sequencing, aligning long reads correctly around the splice sites is a challenging task. Incorrect alignments lead to spurious nodes and arcs in the graph, which in turn lead to incorrect transcript predictions. We survey several approaches to find the exon chains corresponding to long reads in a splicing graph, and experimentally study the performance of these methods using simulated data to allow for sensitivity/precision analysis. Our experiments show that short reads from second-generation sequencing can be used to significantly improve exon chain correctness either by error-correcting the long reads before splicing graph creation, or by using them to create a splicing graph on which the long-read alignments are then projected. We also study the memory and time consumption of various modules, and show that accurate exon chains lead to significantly increased transcript prediction accuracy. Availability: The simulated data and in-house scripts used for this article are available at http://www.cs.helsinki.fi/group/gsa/exon-chains/exon-chains-bib.tar.bz2.
  • Eising, Else; Huisman, Sjoerd M. H.; Mahfouz, Ahmed; Vijfhuizen, Lisanne S.; Anttila, Verneri; Winsvold, Bendik S.; Kurth, Tobias; Ikram, M. Arfan; Freilinger, Tobias; Kaprio, Jaakko; Boomsma, Dorret I.; van Duijn, Cornelia M.; Jarvelin, Marjo-Riitta R.; Zwart, John-Anker; Quaye, Lydia; Strachan, David P.; Kubisch, Christian; Dichgans, Martin; Smith, George Davey; Stefansson, Kari; Palotie, Aarno; Chasman, Daniel I.; Ferrari, Michel D.; Terwindt, Gisela M.; de Vries, Boukje; Nyholt, Dale R.; Lelieveldt, Boudewijn P. F.; van den Maagdenberg, Arn M. J. M.; Reinders, Marcel J. T. (2016)
    Migraine is a common disabling neurovascular brain disorder typically characterised by attacks of severe headache and associated with autonomic and neurological symptoms. Migraine is caused by an interplay of genetic and environmental factors. Genome-wide association studies (GWAS) have identified over a dozen genetic loci associated with migraine. Here, we integrated migraine GWAS data with high-resolution spatial gene expression data of normal adult brains from the Allen Human Brain Atlas to identify specific brain regions and molecular pathways that are possibly involved in migraine pathophysiology. To this end, we used two complementary methods. In GWAS data from 23,285 migraine cases and 95,425 controls, we first studied modules of co-expressed genes that were calculated based on human brain expression data for enrichment of genes that showed association with migraine. Enrichment of a migraine GWAS signal was found for five modules that suggest involvement in migraine pathophysiology of: (i) neurotransmission, protein catabolism and mitochondria in the cortex; (ii) transcription regulation in the cortex and cerebellum; and (iii) oligodendrocytes and mitochondria in subcortical areas. Second, we used the high-confidence genes from the migraine GWAS as a basis to construct local migraine-related co-expression gene networks. Signatures of all brain regions and pathways that were prominent in the first method also surfaced in the second method, thus providing support that these brain regions and pathways are indeed involved in migraine pathophysiology.
  • Katayama, Shintaro; Skoog, Tiina; Jouhilahti, Eeva-Mari; Siitonen, H. Annika; Nuutila, Kristo; Tervaniemi, Mari H.; Vuola, Jyrki; Johnsson, Anna; Lonnerberg, Peter; Linnarsson, Sten; Elomaa, Outi; Kankuri, Esko; Kere, Juha (2015)
    Background: Keratinocytes (KCs) are the most frequent cells in the epidermis, and they are often isolated and cultured in vitro to study the molecular biology of the skin. Cultured primary cells and various immortalized cells have been frequently used as skin models but their comparability to intact skin has been questioned. Moreover, when analyzing KC transcriptomes, fluctuation of polyA+ RNA content during the KCs' lifecycle has been omitted. Results: We performed STRT RNA sequencing on 10 ng samples of total RNA from three different sample types: i) epidermal tissue (split-thickness skin grafts), ii) cultured primary KCs, and iii) HaCaT cell line. We observed significant variation in cellular polyA+ RNA content between tissue and cell culture samples of KCs. The use of synthetic RNAs and SAMstrt in normalization enabled comparison of gene expression levels in the highly heterogenous samples and facilitated discovery of differences between the tissue samples and cultured cells. The transcriptome analysis sensitively revealed genes involved in KC differentiation in skin grafts and cell cycle regulation related genes in cultured KCs and emphasized the fluctuation of transcription factors and non-coding RNAs associated to sample types. Conclusions: The epidermal keratinocytes derived from tissue and cell culture samples showed highly different polyA+ RNA contents. The use of SAMstrt and synthetic RNA based normalization allowed the comparison between tissue and cell culture samples and thus proved to be valuable tools for RNA-seq analysis with translational approach. Transciptomics revealed clear difference both between tissue and cell culture samples and between primary KCs and immortalized HaCaT cells.
  • Psychiat Genomics Consortium; 23andMe Res Team; Psychosis Endopheno-types Int Cons; Wellcome Trust Case Control Consor; Lee, Phil H.; Anttila, Verneri; Won, Hyejung; Kaprio, Jaakko; Keski-Rahkonen, Anna; Churchhouse, Claire; Rehnström, Karola; Raevuori, Anu; Palotie, Aarno; Daly, Mark J.; Neale, Benjamin M. (2019)
    Genetic influences on psychiatric disorders transcend diagnostic boundaries, suggesting substantial pleiotropy of contributing loci. However, the nature and mechanisms of these pleiotropic effects remain unclear. We performed analyses of 232,964 cases and 494,162 controls from genome-wide studies of anorexia nervosa, attention-deficit/hyper-activity disorder, autism spectrum disorder, bipolar disorder, major depression, obsessive-compulsive disorder, schizophrenia, and Tourette syndrome. Genetic correlation analyses revealed a meaningful structure within the eight disorders, identifying three groups of inter-related disorders. Meta-analysis across these eight disorders detected 109 loci associated with at least two psychiatric disorders, including 23 loci with pleiotropic effects on four or more disorders and 11 loci with antagonistic effects on multiple disorders. The pleiotropic loci are located within genes that show heightened expression in the brain throughout the lifespan, beginning prenatally in the second trimester, and play prominent roles in neurodevelopmental processes. These findings have important implications for psychiatric nosology, drug development, and risk prediction.
  • Sood, Sanjana; Szkop, Krzysztof J.; Nakhuda, Asif; Gallagher, Iain J.; Murie, Carl; Brogan, Robert J.; Kaprio, Jaakko; Kainulainen, Heikki; Atherton, Philip J.; Kujala, Urho M.; Gustafsson, Thomas; Larsson, Ola; Timmons, James A. (2016)
    DNA microarrays and RNAseq are complementary methods for studying RNA molecules. Current computational methods to determine alternative exon usage (AEU) using such data require impractical visual inspection and still yield high false-positive rates. Integrated Gene and Exon Model of Splicing (iGEMS) adapts a gene-level residuals model with a gene size adjusted false discovery rate and exon-level analysis to circumvent these limitations. iGEMS was applied to two new DNA microarray datasets, including the high coverage Human Transcriptome Arrays 2.0 and performance was validated using RT-qPCR. First, AEU was studied in adipocytes treated with (n = 9) or without (n = 8) the anti-diabetes drug, rosiglitazone. iGEMS identified 555 genes with AEU, and robust verification by RT-qPCR (similar to 90%). Second, in a three-way human tissue comparison (muscle, adipose and blood, n = 41) iGEMS identified 4421 genes with at least one AEU event, with excellent RT-qPCR verification (95%, n = 22). Importantly, iGEMS identified a variety of AEU events, including 3'UTR extension, as well as exon inclusion/exclusion impacting on protein kinase and extracellular matrix domains. In conclusion, iGEMS is a robust method for identification of AEU while the variety of exon usage between human tissues is 5-10 times more prevalent than reported by the Genotype-Tissue Expression consortium using RNA sequencing.
  • Eising, Else; de Leeuw, Christiaan; Min, Josine L.; Anttila, Verneri; Verheijen, Mark H. G.; Terwindt, Gisela M.; Dichgans, Martin; Freilinger, Tobias; Kubisch, Christian; Ferrari, Michel D.; Smit, August B.; de Vries, Boukje; Palotie, Aarno; van den Maagdenberg, Arn M. J. M.; Posthuma, Danielle; Int Headache Genetics Consortium (2016)
    Background Migraine is a common episodic brain disorder characterized by recurrent attacks of severe unilateral headache and additional neurological symptoms. Two main migraine types can be distinguished based on the presence of aura symptoms that can accompany the headache: migraine with aura and migraine without aura. Multiple genetic and environmental factors confer disease susceptibility. Recent genome-wide association studies (GWAS) indicate that migraine susceptibility genes are involved in various pathways, including neurotransmission, which have already been implicated in genetic studies of monogenic familial hemiplegic migraine, a subtype of migraine with aura. Methods To further explore the genetic background of migraine, we performed a gene set analysis of migraine GWAS data of 4954 clinic-based patients with migraine, as well as 13,390 controls. Curated sets of synaptic genes and sets of genes predominantly expressed in three glial cell types (astrocytes, microglia and oligodendrocytes) were investigated. Discussion Our results show that gene sets containing astrocyte- and oligodendrocyte-related genes are associated with migraine, which is especially true for gene sets involved in protein modification and signal transduction. Observed differences between migraine with aura and migraine without aura indicate that both migraine types, at least in part, seem to have a different genetic background.
  • Hakonen, Elina; Chandra, Vikash; Fogarty, Christopher L.; Yu, Nancy Yiu-Lin; Ustinov, Jarkko; Katayama, Shintaro; Galli, Emilia; Danilova, Tatiana; Lindholm, Paivi; Vartiainen, Aki; Einarsdottir, Elisabet; Krjutskov, Kaarel; Kere, Juha; Saarma, Mart; Lindahl, Maria; Otonkoski, Timo (2018)
    There is a great need to identify factors that could protect pancreatic beta cells against apoptosis or stimulate their replication and thus prevent or reverse the development of diabetes. One potential candidate is mesencephalic astrocyte-derived neurotrophic factor (MANF), an endoplasmic reticulum (ER) stress inducible protein. Manf knockout mice used as a model of diabetes develop the condition because of increased apoptosis and reduced proliferation of beta cells, apparently related to ER stress. Given this novel association between MANF and beta cell death, we studied the potential of MANF to protect human beta cells against experimentally induced ER stress. Primary human islets were challenged with proinflammatory cytokines, with or without MANF. Cell viability was analysed and global transcriptomic analysis performed. Results were further validated using the human beta cell line EndoC-beta H1. There was increased expression and secretion of MANF in human beta cells in response to cytokines. Addition of recombinant human MANF reduced cytokine-induced cell death by 38% in human islets (p <0.05). MANF knockdown in EndoC-beta H1 cells led to increased ER stress after cytokine challenge. Mechanistic studies showed that the protective effect of MANF was associated with repression of the NF-kappa B signalling pathway and amelioration of ER stress. MANF also increased the proliferation of primary human beta cells twofold when TGF-beta signalling was inhibited (p <0.01). Our studies show that exogenous MANF protein can provide protection to human beta cells against death induced by inflammatory stress. The antiapoptotic and mitogenic properties of MANF make it a potential therapeutic agent for beta cell protection.
  • van der Kolk, Birgitta W.; Saari, Sina; Lovric, Alen; Arif, Muhammad; Alvarez, Marcus; Ko, Arthur; Miao, Zong; Sahebekhtiari, Navid; Muniandy, Maheswary; Heinonen, Sini; Oghabian, Ali; Jokinen, Riikka; Jukarainen, Sakari; Hakkarainen, Antti; Lundbom, Jesper; Kuula, Juho; Groop, Per-Henrik; Tukiainen, Taru; Lundbom, Nina; Rissanen, Aila; Kaprio, Jaakko; Williams, Evan G.; Zamboni, Nicola; Mardinoglu, Adil; Pajukanta, Paivi; Pietiläinen, Kirsi H. (2021)
    Tissue-specific mechanisms prompting obesity-related development complications in humans remain unclear. We apply multiomics analyses of subcutaneous adipose tissue and skeletal muscle to examine the effects of acquired obesity among 49 BMI-discordant monozygotic twin pairs. Overall, adipose tissue appears to be more affected by excess body weight than skeletal muscle. In heavier co-twins, we observe a transcriptional pattern of downregulated mitochondrial pathways in both tissues and upregulated inflammatory pathways in adipose tissue. In adipose tissue, heavier co-twins exhibit lower creatine levels; in skeletal muscle, glycolysis- and redox stress-related protein and metabolite levels remain higher. Furthermore, metabolomics analyses in both tissues reveal that several proinflammatory lipids are higher and six of the same lipid derivatives are lower in acquired obesity. Finally, in adipose tissue, but not in skeletal muscle, mitochondrial downregulation and upregulated inflammation are associated with a fatty liver, insulin resistance, and dyslipidemia, suggesting that adipose tissue dominates in acquired obesity.
  • Nair, Preethy Sasidharan; Kuusi, Tuire; Ahvenainen, Minna; Philips, Anju K.; Järvelä, Irma (2019)
    Musical training and performance require precise integration of multisensory and motor centres of the human brain and can be regarded as an epigenetic modifier of brain functions. Numerous studies have identified structural and functional differences between the brains of musicians and non-musicians and superior cognitive functions in musicians. Recently, music-listening and performance has also been shown to affect the regulation of several genes, many of which were identified in songbird singing. MicroRNAs affect gene regulation and studying their expression may give new insights into the epigenetic effect of music. Here, we studied the effect of 2 hours of classical music-performance on the peripheral blood microRNA expressions in professional musicians with respect to a control activity without music for the same duration. As detecting transcriptomic changes in the functional human brain remains a challenge for geneticists, we used peripheral blood to study music-performance induced microRNA changes and interpreted the results in terms of potential effects on brain function, based on the current knowledge about the microRNA function in blood and brain. We identified significant (FDR
  • Kachroo, Priyanka; Eraso, Jesus M.; Olsen, Randall J.; Zhu, Luchang; Kubiak, Samantha L.; Pruitt, Layne; Yerramilli, Prasanti; Cantu, Concepcion C.; Saavedra, Matthew Ojeda; Pensar, Johan; Corander, Jukka; Jenkins, Leslie; Kao, Lillian; Granillo, Alejandro; Porter, Adeline R.; DeLeo, Frank R.; Musser, James M. (2020)
    A fundamental goal of contemporary biomedical research is to understand the molecular basis of disease pathogenesis and exploit this information to develop targeted and more-effective therapies. Necrotizing myositis caused by the bacterial pathogen Streptococcus pyogenes is a devastating human infection with a high mortality rate and few successful therapeutic options. We used dual transcrip-tome sequencing (RNA-seq) to analyze the transcriptomes of S. pyogenes and host skeletal muscle recovered contemporaneously from infected nonhuman primates. The in vivo bacterial transcriptome was strikingly remodeled compared to organisms grown in vitro, with significant upregulation of genes contributing to virulence and altered regulation of metabolic genes. The transcriptome of muscle tissue from infected nonhuman primates (NHPs) differed significantly from that of mock-infected animals, due in part to substantial changes in genes contributing to inflammation and host defense processes. We discovered significant positive correlations between group A streptococcus (GAS) virulence factor transcripts and genes involved in the host immune response and inflammation. We also discovered significant correlations between the magnitude of bacterial virulence gene expression in vivo and pathogen fitness, as assessed by previously conducted genome-wide transposon-directed insertion site sequencing (TraDIS). By integrating the bacterial RNA-seq data with the fitness data generated by TraDIS, we discovered five new pathogen genes, namely, S. pyogenes 0281 (Spy0281 [dahA]), ihk-irr, slr, isp, and ciaH, that contribute to necrotizing myositis and confirmed these findings using isogenic deletion-mutant strains. Taken together, our study results provide rich new information about the molecular events occurring in severe invasive infection of primate skeletal muscle that has extensive translational research implications. IMPORTANCE Necrotizing myositis caused by Streptococcus pyogenes has high morbidity and mortality rates and relatively few successful therapeutic options. In addition, there is no licensed human S. pyogenes vaccine. To gain enhanced understanding of the molecular basis of this infection, we employed a multidimensional analysis strategy that included dual RNA-seq and other data derived from experimental infection of nonhuman primates. The data were used to target five streptococcal genes for pathogenesis research, resulting in the unambiguous demonstration that these genes contribute to pathogen-host molecular interactions in necrotizing infections. We exploited fitness data derived from a recently conducted genome-wide transposon mutagenesis study to discover significant correlation between the magnitude of bacterial virulence gene expression in vivo and pathogen fitness. Collectively, our findings have significant implications for translational research, potentially including vaccine efforts.
  • Tervaniemi, Mari H.; Katayama, Shintaro; Skoog, Tiina; Siitonen, H. Annika; Vuola, Jyrki; Nuutila, Kristo; Sormunen, Raija; Johnsson, Anna; Linnarsson, Sten; Suomela, Sari; Kankuri, Esko; Kere, Juha; Elomaa, Outi (2016)
    Psoriatic skin differs distinctly from normal skin by its thickened epidermis. Most gene expression comparisons utilize full-thickness biopsies, with substantial amount of dermis. We assayed the transcriptomes of normal, lesional, and non-lesional psoriatic epidermis, sampled as split-thickness skin grafts, with 5'-end RNA sequencing. We found that psoriatic epidermis contains more mRNA per total RNA than controls, and took this into account in the bioinformatic analysis. The approach highlighted innate immunity-related pathways in psoriasis, including NOD-like receptor (NLR) signaling and inflammasome activation. We demonstrated that the NLR signaling genes NOD2, PYCARD, CARD6, and IFI16 are upregulated in psoriatic epidermis, and strengthened these findings by protein expression. Interestingly, PYCARD, the key component of the inflammasome, showed an altered expression pattern in the lesional epidermis. The profiling of non-lesional skin highlighted PSORS4 and mitochondrially encoded transcripts, suggesting that their gene expression is altered already before the development of lesions. Our data suggest that all components needed for the active inflammasome are present in the keratinocytes of psoriatic skin. The characterization of inflammasome pathways provides further opportunities for therapy. Complementing previous transcriptome studies, our approach gives deeper insight into the gene regulation in psoriatic epidermis.
  • Morales, Luis O.; Shapiguzov, Alexey; Safronov, Omid; Leppälä, Johanna; Vaahtera, Lauri; Yarmolinsky, Dmitry; Kollist, Hannes; Brosche, Mikael (2021)
    Tropospheric ozone (O-3) is a major air pollutant that decreases yield of important crops worldwide. Despite long-lasting research of its negative effects on plants, there are many gaps in our knowledge on how plants respond to O-3. In this study, we used natural variation in the model plant Arabidopsis (Arabidopsis thaliana) to characterize molecular and physiological mechanisms underlying O-3 sensitivity. A key parameter in models for O-3 damage is stomatal uptake. Here we show that the extent of O-3 damage in the sensitive Arabidopsis accession Shahdara (Sha) does not correspond with O-3 uptake, pointing toward stomata-independent mechanisms for the development of O-3 damage. We compared tolerant (Col-0) versus sensitive accessions (Sha, Cvi-0) in assays related to photosynthesis, cell death, antioxidants, and transcriptional regulation. Acute O-3 exposure increased cell death, development of lesions in the leaves, and decreased photosynthesis in sensitive accessions. In both Sha and Cvi-0, O-3-induced lesions were associated with decreased maximal chlorophyll fluorescence and low quantum yield of electron transfer from Photosystem II to plastoquinone. However, O-3-induced repression of photosynthesis in these two O-3-sensitive accessions developed in different ways. We demonstrate that O-3 sensitivity in Arabidopsis is influenced by genetic diversity given that Sha and Cvi-0 developed accession-specific transcriptional responses to O-3. Our findings advance the understanding of plant responses to O-3 and set a framework for future studies to characterize molecular and physiological mechanisms allowing plants to respond to high O-3 levels in the atmosphere as a result of high air pollution and climate change.
  • Ferreira, Pedro G.; Oti, Martin; Barann, Matthias; Wieland, Thomas; Ezquina, Suzana; Friedlander, Marc R.; Rivas, Manuel A.; Esteve-Codina, Anna; Rosenstiel, Philip; Strom, Tim M.; Lappalainen, Tuuli; Guigo, Roderic; Sammeth, Michael; GEUVADIS Consortium; Palotie, A. (2016)
    Recent advances in the cost-efficiency of sequencing technologies enabled the combined DNA-and RNA-sequencing of human individuals at the population-scale, making genome-wide investigations of the inter-individual genetic impact on gene expression viable. Employing mRNA-sequencing data from the Geuvadis Project and genome sequencing data from the 1000 Genomes Project we show that the computational analysis of DNA sequences around splice sites and poly-A signals is able to explain several observations in the phenotype data. In contrast to widespread assessments of statistically significant associations between DNA polymorphisms and quantitative traits, we developed a computational tool to pinpoint the molecular mechanisms by which genetic markers drive variation in RNA-processing, cataloguing and classifying alleles that change the affinity of core RNA elements to their recognizing factors. The in silico models we employ further suggest RNA editing can moonlight as a splicing-modulator, albeit less frequently than genomic sequence diversity. Beyond existing annotations, we demonstrate that the ultra-high resolution of RNA-Seq combined from 462 individuals also provides evidence for thousands of bona fide novel elements of RNA processing-alternative splice sites, introns, and cleavage sites-which are often rare and lowly expressed but in other characteristics similar to their annotated counterparts.