Bio- ja ympäristötieteellinen tiedekunta: Recent submissions

Now showing items 1-20 of 922
  • Spoljaric, Inkeri (Helsingin yliopisto, 2019)
    Spontaneously arising network events are a characteristic feature of all developing neural networks. This activity is crucial for normal neuronal development and the establishment of appropriate synaptic connectivity. In the developing hippocampus, depolarizing GABAergic drive is essential in generation of early network events, known as giant depolarizing potentials (GDPs). Blockade of GABAergic signaling leads to hypersynchronization of the network and emergence of ictal-like events, pointing to dual, both excitatory and inhibitory roles for GABA, in regulation of these events. In Studies I-III of this thesis, we examined the role of GABAA receptor (GABAAR) -mediated neurotransmission with some parallel work on glycinergic signaling as well as neuronal Cl- regulation in modulation of GDPs in the developing rodent hippocampus. In Study I, we demonstrate that low levels of GABA and glycine suppress GDPs by activating extrasynaptic receptors. This implies that regardless of the depolarizing drive for Cl- currents at this developmental stage, a low conductance via Cl- -permeable GABAARs and glycine receptors (GlyRs) can cause efficient shunting and inhibition of the network events. In Study II, we discovered that sustained activation of a subset of hippocampal interneurons, caused by the neuropeptide arginine vasopressin (AVP), silences the network events in the perinatal hippocampus, regardless of the maturational level of the GABAergic system as compared across species. This is attributed to decreased synchronous interneuronal input that is essential for the GDP generation. In Study III, we demonstrate that transport-functional K-Cl cotransporter 2 (KCC2) is present in the CA3 pyramidal neurons already in the perinatal stages in mice and rats. Cl- extrusion by KCC2 counteracts the dominant Na-K-2Cl cotransporter 1 (NKCC1) -mediated Cl- uptake and restrains the depolarizing GABAergic drive onto the CA3 pyramidal cells. Thereby, function of KCC2 limits pyramidal neuron spiking and synchronization during GDPs and participates in the modulation of GDPs from their developmental onset. This work describes novel physiological GABAergic mechanisms that control GDPs in the perinatal rodents and establishes a role for KCC2 in regulation of pyramidal neuron excitability and synchronization during GDPs starting from their developmental onset.
  • Camarena Gómez, María Teresa (Helsingin yliopisto, 2019)
    Global warming is one of the most alarming pressures affecting marine ecosystems worldwide. One of the indirect effects of the increasing surface-water temperature is the change in phytoplankton community composition, shifting in some ecosystems from diatom predominance towards the dinoflagellate predominance or co-occurrence with diatoms during blooms. These distinct phytoplankton groups vary in the quality and/or quantity of the dissolved organic matter (DOM) they release, which may have contrasting effects on the associated bacterioplankton communities, in terms of structure and function, and also on the carbon flux passing through the microbial loop. The main objective of this thesis was to assess the effect of diatoms and dinoflagellates on shaping the bacterial community composition and dynamics in different ecosystems in which either one or both of these two groups dominate the phytoplankton bloom, such as the Baltic Sea and the Humboldt Current System (HCS) off Chile. This was achieved by conducting both experimental and field studies in these areas. Phytoplankton community composition and the stage of the bloom phase clearly affected to the bacterial community composition and dynamics in both ecosystems. Alphaproteobacteria, dominated by SAR11 and Rhodobacteraceae, was the most abundant bacterial class in all studies. The oligotrophic SAR11 dominated in pre-bloom conditions and was associated with dinoflagellates. In contrast, copiotrophic bacteria belonging to the classes Flavobacteriia, Gammaproteobacteria, Betaproteobacteria and the family Rhodobacteraceae (genera Loktanella, Planktotalea, Planktomarina and Amylibacter) were associated with diatom species such as Achnanthes taeniata, Chaetoceros spp., Skeletonema costatum and Thalassiosira levanderi in the Baltic Sea and with Thalassiosira spp. in the HCS. In addition, in the Baltic Sea, bacterial communities dominated by copiotrophs had higher bacterial production rates than in SAR11 dominated bacterial communities. Hence, the diatom-released DOM boosted the development of more productive bacterial communities during phytoplankton blooms. Further differences in the bacterial community composition were driven by the different salinities in these two ecosystems; Betaproteobacteria, Planctomycetes and Actinobacteria were more abundant in the brackish Baltic Sea than in the HCS. The results of this thesis highlight that the shift to the dinoflagellate dominance or co-occurrence with diatoms may affect the bacterial community composition and activity during bloom events. Certain diatom species promote the growth of copiotrophic bacteria, which contribute largely to high bacterial production rates and recycling of carbon. In contrast, the increase in dinoflagellate abundance associated with global warming may potentially change the pelagic remineralization of organic matter, which could reduce the carbon flux to higher trophic levels.
  • Misiewicz, Zuzanna (Helsingin yliopisto, 2019)
    Anxiety disorders manifest themselves as a prolonged or exaggerated response to a threatening situation, which can be either real or perceived. Their high prevalence (14%) places them as one of the most common mental disorders within the European Union. This conveys an important message about the necessity of finding new clinically relevant drug targets leading to the development of novel personalized treatment practices. To facilitate this process, efforts should be focused on gaining a deeper understanding of the complex molecular, biochemical, and system-level mechanisms behind the neurobiology of stress, and the role of stress as one of the main etiological factors in anxiety-related psychiatric disorders. The phenotypic heterogeneity of human populations and high variability of external environmental factors, along with limited access to brain tissue samples, presents some of the main challenges to studying anxiety disorders in humans. As these aspects can be controlled for in animals, animal models are often used to administer specific stressors in a uniform manner and to obtain brain tissue at a precisely chosen time point. Thereby, within the scope of this thesis, we take advantage of the fact that anxiety is an evolutionarily conserved response and address the integration of both human and mouse data obtained from a variety of approaches, including genomic, transcriptomic, and proteomic methods. First, to identify genetic loci predisposing to a specific phobia, the fear of heights, we conducted a genome-wide parametric and non-parametric linkage scan, followed by joint linkage and association analysis in a small population isolate with reduced genetic and environmental heterogeneity. Our results implicated three regions with suggestive evidence for linkage, including region 8q24.2-q24.3 (LOD = 2.09), which encompasses 49 genes, including several candidate genes for psychiatric disorders. Second, we identified molecules and biological pathways affected by chronic social defeat stress (CSDS), a mouse model of chronic psychosocial stress, in the following three brain regions: medial prefrontal cortex (PFCM), ventral hippocampus (HIPV), and bed nucleus of the stria terminalis (BNST). We used two inbred mouse strains with different basal anxiety levels, the innately non-anxious C57BL/6NCrl (B6) and innately anxious DBA/2NCrl (D2). Following analysis of RNA sequencing results, we discovered that differentially expressed (DE) oligodendrocyte-related genes were over-represented in gene set enrichment analysis (GSEA) of all studied brain regions. As oligodendrocytes are known for their function in axon myelination, we followed the results from transcriptomic analyses with transmission electron microscopy (TEM) and established that B6 stress-susceptible mice had thicker myelin in BNST axons compared to controls. Third, using the CSDS model, we further investigated the role of the BNST through additional studies of gene regulatory networks (GRN) of mRNAs and miRNAs and protein-protein interaction networks. Subsequently, we followed with an integration analysis of the results from both transcriptomic (mRNA sequencing, as well as AGO2 miRNA, and mRNA immunoprecipitation sequencing) and proteomic (liquid chromatography–tandem mass spectrometry) experiments. Furthermore, to translate our results to human anxiety disorders, we performed transcriptome profiling in blood cells of CSDS-subjected mice and compared it with gene expression patterns from blood cells of panic disorder patients who underwent exposure-induced panic attacks. We then followed with integrative gene set enrichment analysis of mouse and human data, which showed systemic genetic background-specific enrichment of mitochondria-related gene sets. Importantly, our results showed downregulation of the oxidative phosphorylation pathway in the CSDS-subjected D2 strain and panic disorder patients after a panic attack. To conclude, our results suggest (1) brain-region and mouse strain-specific differences in myelination in susceptibility and resilience to stress and (2) dysregulation of mitochondrial pathways associated with anxiety-related behavior in both mice and humans. Taken together, our results provide further insight into the complex genetic architecture of anxiety disorders and support the suitability of cross-species approaches to studying biological mechanisms underlying anxiety disorders. Keywords: anxiety disorders, chronic social defeat stress (CSDS), panic disorder, acrophobia, linkage analysis, gene expression, protein abundance, multi-omics studies
  • Uusheimo, Sari (Helsingin yliopisto, 2019)
    ABSTRACT Massive amounts of industrially-fixed reactive nitrogen (N) results in excess N amounts in the environment. High N loading from agricultural catchments and wastewater treatment plants can be mitigated by natural N removal processes in aquatic environments. These microbe meditated processes, mainly denitrification, remove the reactive N returning it back to the atmosphere mainly in the form of harmless N2 gas. These processes occur in anoxic sediments and are dependent on the surrounding temperature. Generally, microbial N removal has been found to be efficient in wetlands, compared with other aquatic environments. In a cold climate, the process rates of microbial N removal are assumed to be low, however they are poorly understood. In this study, N removal rates were measured in a shallow agricultural wetland and in a wetland polishing treated wastewater situated in southern Finland. In the agricultural wetland, the influence of environmental factors were studied by conducting the study in field conditions. Seasonal denitrification rates were governed mainly by sediment temperature, followed by sediment oxygen conditions and water turbidity. Diurnally, denitrification was regulated by light affecting sediment oxygen conditions. Rates measured in the field merely during daytime may lead to underestimations of N removal potential of shallow wetlands. Nutrient retention and the contribution of N removal processes to the total N retention was estimated in the wetland receiving treated wastewater. The efficient N retention found was estimated to be mostly based on microbial processes, mainly on denitrification and slightly on anammox. In addition, the interference arising from organic carbon to nitrate measurements with optical sensors was studied and the information was used in improving the accuracy of sensor nitrate results in an agricultural brook. Increasing organic carbon concentrations was found to increase the nitrate concentrations by sensors and the effect was manifold in bog water. The interference was resolved by taking account the dissolved organic carbon in the correction of the sensor results. This study increases the information of actual N removing process rates and improves the knowledge of the extent constructed wetlands can remove excess N in cold climate, and provides information for the use of optical sensors in monitoring N loading.
  • Amiryousefi, Ali (Helsingin yliopisto, 2019)
    Chloroplasts are cytoplasmic organelles chiefly responsible for the photosynthesis. Their genes have been used extensively during the past decades in phylogenetic analyses of various photosynthetic eukaryotes, particularly plants. The genomic content of this organelle and its very architecture can be used for a deeper insight in evolution and towards robust phylogenetic hypotheses. Ever since this importance was recognized concurrently with the advancements of methods in both providing a basic genetic material through sequencing and advanced methods to analyze the data, we have witnessed the introduction of a couple of thousands plastid genomes up to this date. This process, by no means is in its decline or even stationary state, as this pace is projected to be accelerated in the coming years, with the inevitable advances in our technologies and our need to understand the nature as accurate as possible. The aim of this study as represented in the sequel chapters is twofold; 1) to introduce the complete chloroplast genomes of two species from the euasterid clade and provide their phylogenetic analyses; Solanum dulcamara L. as a native Old World diploid member of the nightshade family, and Ambrosia trifida L. as a recognized invasive plant originated and evolved from North America. 2) To provide two analytical tools for more advanced treatment of the genetic information of plastids in bioinformatics. By comparative analysis for bittersweet and giant ragweed, the result show that synteny and the genomic content of both belonging to the families Solanaceae and Asteraceae, respectively, have a conserved structure. We also noted that many submitted annotations in the nightshade family are far from acceptable quality, and further on, we improved them with reannotation of the existing sequences. On the other hand, a novel tool (IRscope) to detect and plot the Inverted Repeat (IR) regions of the chloroplast genome was introduced. IRscope, with the help of iterative search algorithm, allows the depiction of genes in the vicinity of the Junction Sites (JS), of up to ten different chloroplast genomes of embryophytes (land plants). Moreover, we constructed an online calculative suite (iMEC) to return the result of the seven different molecular markers against the provided input file. This tool is useful particularly in studies aimed to assess the efficiency of different marker systems linked to plastid genome variation.
  • Aho, Velma T. E. (Helsingin yliopisto, 2019)
    Parkinson's disease is the second most common neurodegenerative disease in the world, and in spite of decades of research, the cause of the non-familial form of the disease is not known. There are currently no medications to slow down the progression of the disease nor good biomarkers for early diagnosis, even though the earliest non-motor symptoms can appear years or even decades before the onset of motor symptoms. The microbial inhabitants of the human body have recently been implicated in various medical conditions, including neurodegenerative disorders. They could offer new insight into the pathogenesis of Parkinson's disease, particularly since a microbial agent has long been suspected to play a part in the process. A typical question in studies surveying human microbiota is which specific microbial taxa differ between groups of interest, such as patients with a disease and control subjects. There are many statistical tools for performing these analyses, also known as differential abundance comparisons. The aim of my doctoral thesis was to explore the potential associations of Parkinson's disease and human microbiota, particularly of the mouth, the nose and the gut, with an additional focus on the statistical tools used for comparing differentially abundant bacterial taxa. All four publications included in my thesis were based on samples from the same subjects: 76 patients with Parkinson's disease and 76 control subjects with no signs of parkinsonism. The studies also used the same methodology, 16S rRNA gene amplicon sequencing, to compare bacteria from oral and nasal swab samples and fecal samples of these subjects. Over the course of the four publications and in a previously unpublished analysis, I compared differentially abundant taxa with six tools: Metastats, LEfSe, metagenomeSeq, DESeq2, ANCOM, and an approach based on random forests. Our results suggested that the bacterial communities of the gut and the mouth differ between Parkinson's patients and control subjects, with statistically significant differences in beta diversity and in the abundances of several bacterial taxa. Differences in gut microbiota could also be detected at a follow-up time point with samples collected two years after the initial sampling. Additionally, there were differences between the gut bacteria of Parkinson's patients with and without irritable bowel syndrome -like symptoms. For nasal bacteria, there were no differences between the patient and control groups in diversity nor the amounts of specific bacteria. Regarding the differential abundance analyses, comparing gut bacteria of patients and controls from the same samples with six different tools highlighted the wide variation in the lists of significant results, which often did not overlap except for a handful of taxa. While a few benchmarking studies have previously contrasted some of the tools, there is a definite need for further standardized testing to guide researchers in choosing between them. Despite these discrepancies, all tools tested in this thesis supported Parkinson's patients having a decreased abundance of the family Prevotellaceae in their gut. This difference in abundance could also be detected at the follow-up time point. As several other research groups have reported seeing a decrease in Prevotellaceae after our pilot publication, it is emerging as one of the key changes in microbiota associated with Parkinson's disease.
  • Kuony, Alison (Helsingin yliopisto, 2019)
    The lacrimal apparatus products the tear film protecting the eye surface. The lacrimal gland, located in the orbit, produces 80% of the tear film components. These include growth factors destined to the avascular cornea, necessary for corneal epithelium maturation and renewal. A defective tear film leads to dry eye diseases which affect an increasing percentage of the population (more than 35% in the elderly). Although dry eye diseases can lead to visual loss in extreme cases, no definitive treatment has been developed so far. My thesis work focused on unravelling lacrimal gland early development, postnatal maturation and subsequent function in mouse models. I notably used a dry eye disease mouse model to study the implication of Ectodysplasin (Eda) pathway in lacrimal gland development, maturation, function, as well as in the cornea/lacrimal gland feedback loop during corneal healing process. In addition of improving the general knowledge on lacrimal gland biology, my work gave some insights on the dry eye phenotype associated to Ectodysplasin loss-of-function mutation and on the resulting defective cornea wound healing.
  • Leppänen, Jaakko (Helsingin yliopisto, 2019)
    The mining industry has a critical role in modern societies. However, the environmental harm induced by mining activities may be drastic. One of the most important aspects of mining pollution is water contamination due to accidental or controlled waste water release, or due to harmful leachate originating from waste areas. Whereas the water quality in receiving water systems is usually monitored, the ecological impacts of mine waters are assessed more rarely. One of the challenges when ecological impacts are studied is the lack of long-term data regarding the pre-mining conditions. Palaeolimnological methods allow the reconstruction of historical conditions of water bodies. Sedimentary geochemistry and subfossil remains of biological communities (e.g. cladoceran crustacea, diatom algae) can be used to assess the natural pre-disturbance variability, the impact of the disturbance and the post-disturbance dynamics. In this thesis, mining pollution is assessed within three projects, conducted on two boreal lakes in Finland. In Lake Kirkkojärvi, the impact of historical mining activity, namely, the direct release of mineral tailings was studied from a sediment core covering also the pre-mining environmental conditions. The impacts of the mine pollutants were reflected in cladoceran communities as declining species richness and diversity. The lake Kirkkojärvi bay was studied by using a palaeolimnological top-bottom approach. The bay has been receiving acid mine drainage for nearly 50 years. However, the cladoceran community change in the bay does not reflect any changes in acidity or metal pollution. In Lake Kivijärvi the Na2SO4- and metal-contaminated mine water has had a clear impact on cladoceran and diatom communities as both groups exhibit changes due to mining pollution. Based on the results, the most important contaminants in the studied systems are saline effluents and mineral tailings. Cladocera hold great potential in palaeolimnological mining pollution studies due to their sensitivity to changes in the water quality and due to their central location in food-webs. However, more research is still needed: Most importantly, some taxonomic issues must be clarified and in-situ toxicity studies should be conducted in order to find the most sensitive indicator species for different types of pollution. The results of this work present important information regarding pollution impact assessment. In particular, this study highlights the utility of Cladocera as early warning indicators, while supporting the use of multiple sediment proxies in palaeolimnological pollution research in order to provide information about the timing, direction, and magnitude of ecosystem impacts caused by pollution events. This is of essence for environmental management as globally the exploitation of poor-grade large mine deposits will drastically increase in the near future. Only by providing such crucial information on the short- and long-term impacts of mining on aquatic ecosystems and their resilience, a technologically, socially and environmentally sustainable management of mines can be guaranteed in the future.
  • Viita, Tiina (Helsingin yliopisto, 2019)
    Actin is best-known from its functions in the cytoplasm, where it is a key component of the cytoskeleton. Nevertheless, functions of actin are not restricted to the cytoplasm, since actin is also present in the nucleus, where it has been linked to multiple functions from gene activation to chromatin remodeling. However, molecular mechanisms by which actin operates in the nucleus are still poorly understood. This is mainly because of the lack of well-characterized binding partners for nuclear actin. Therefore, the aim of this thesis was to identify and characterize novel nuclear actin-binding partners and elucidate the molecular mechanisms behind actin regulated transcription factors. To identify nuclear actin-binding partners, we used two complementary mass spectrometry (MS) techniques, affinity purification combined with MS (AP-MS) and proximity dependent biotin identification with MS (BioID). Analysis of our interactome data revealed that actin can form stable complexes with proteins related to chromatin remodeling but seems to function in a dynamic fashion in other nuclear processes, such as transcription and DNA replication. In our experimental setup actin seemed to be monomeric when it associated with nuclear complexes. We also discovered a novel actin-containing complex, human Ada-Two-A-containing complex (hATAC). HATAC is a histone modifying complex and actin directly binds one of it subunits, lysine acetyltransferase 14 (KAT14). We showed that actin-binding modulates histone acetyl transferase (HAT) activity of KAT14 in vitro and in cells. Our interactome data also indicated that actin could be involved in RNA splicing and mRNA processing. Further experiments showed, for the first time, that actin has a functional role in mRNA splicing, as alterations in nuclear actin levels disturbed survival motor neuron protein 2 (SMN2) alternative exon skipping. In addition, the nuclear actin interactome analysis provided new insights into nuclear processes already earlier linked to actin, such as chromatin remodeling, transcription and DNA replication, and hence this work provides a protein interaction platform for further mechanistic studies of nuclear actin-dependent functions. One well-known protein, which is known to bind nuclear actin and to regulate transcription is myocardin-related transcription factor A (MRTF-A), a co-activator of serum response factor (SRF). MRTF-A has a RPEL domain, which can regulate the localization and activity of MRTF-A in response to actin dynamics. Interestingly, also phosphatase and actin regulator (Phactr) protein family contains RPEL domain and binds protein phosphatase 1 (PP1). The molecular properties of MRTF-A intrigued us to study if the Phactr-proteins could be putative binding partners of nuclear actin. We revealed that the RPEL domain of Phactr4 do not regulate the subcellular localization of Phactr4. Instead, Phactr4 controls the activity of PP1 in response to actin monomer levels in the cytoplasm, thereby creating a feedback loop that maintains actin homeostasis in cells. In addition, we were able to show that MRTF-A subcellular localization is not solely dependent on actin, since DDX19, an ATP-dependent RNA helicase, can regulate nuclear import of full-length MRTF-A independently of actin dynamics. This thesis work has thereby broadened the knowledge of nuclear actin-binding partners as well as revealed novel regulatory properties of actin-regulated RPEL domain containing proteins.
  • Yadav, Leena (Hansaprint Oy, 2019)
    Nearly all eukaryotic processes are regulated by cyclic phosphorylation-dephosphorylation of cellular signaling proteins, catalyzed by protein kinases and phosphatases, respectively. Until now, the kinases dominated the scientific research, however, in recent years only, the protein phosphatases have emerged as crucial regulators for setting up the levels and amplitude of phosphorylation-driven signaling events. Nevertheless, our molecular and biological awareness of these enzymes is still poor. This thesis aims to gather information on these vital aspects of phosphatase biology. Through our high-throughput systematic affinity-purification coupled to mass spectrometry (AP-MS) approach, we provide the first glimpse on the cellular protein protein interactions and functional networks of human protein phosphatases from three broad families: phosphoprotein phosphatase, metal-dependent phosphatase, and protein tyrosine phosphatase. Next, we obtained the comprehensive view on their enzymatic structure-activity relationship by utilizing inhibitor Okadaic acid, revealing the effect of catalytic disruption on the phosphatase interaction profile and on the phosphorylation status of their various sub-complex components. From here, we expanded the thesis by investigating the functional relevance of unique phosphates-kinase interaction in the context of specific signaling. From dephosphorylation and luciferase assays we established the role of PTPRA phosphatase in negatively regulating RET kinase activities and its down-stream Ras-MAPK pathway, showing RET as novel PTPRA substrate. By including RET cancer mutants- MEN2A and MEN2B, we, further, shed light on the tumor suppressor functions of PTPRA in human cancers. Taken together, this thesis not only provides a global landscape of human protein phosphatase complexes and their functions but also furnish a reliable resource for futuristic investigations.
  • Zimm, Roland (Helsingin yliopisto, 2019)
    The turtle carapace has been considered one of the classic examples of an evolutionary novelty. It is diagnostic of the turtle clade and probably largely responsible for its success in evolution. In more detail, it consists of ossified dermal plates overlaying laterally extended ribs, as well as a regular array of epidermal scutes. Interestingly, the unique and complex mosaic pattern formed by the carapacial scutes is highly conserved amongst almost all extant turtle taxa. Since the morphogenesis of this remarkable pattern has not been studied much before, I present, in this dissertation, a novel computational approach to elucidating carapacial scute development. In a mathematical model, a combination of two mechanistically different reaction-diffusion systems together with growth reproduces scute formation in the carapace. In order to disentangle the underlying gene regulatory network, we disturb carapace development experimentally in embryos of Trachemys scripta and use the model to understand the resulting phenotypes. We show how the carapacial ridge acts as a signaling center which is required for the onset of scute development. Furthermore, the model suggests how phenotypic differences between species might arise and explains, in line with statistical data from different turtle species, how environmental stress, rather than genetic mutations, leads to the very specific patterns of variation found within turtle populations. Thus, our results suggest that environmental stress might be an underrated source of phenotypic variation within species. Developmental insight helps us understand why this particular variation occurs and at what frequencies. In order to further understand how environmental factors might interfere with development and to be able to account for more realistic tissue growth and biomechanics, I present a more general multiscale model of animal development that integrates biomechanics, GRN dynamics and cell behaviours in different, interacting cell types and tissues. Thus, this model does not only allow an advancement of our understanding of mechanisms by which phenotypic variation in the developing turtle scute might be generated, it is also a useful tool to understand more general features of development and variation. Finally, I discuss my work in the context of previous hypotheses about the origins of turtle scutes as well as their variation. By a comparison with the development of other ectodermal organs in different vertebrates, I try to integrate turtle scutes into a wider evolutionary background.
  • Parajuli, Anirudra (Helsingin yliopisto, 2019)
    Microbes play important roles in the development of the human immune system. Human commensal microbiota of the gut and skin have been shown to modulate the immune system, and high diversity of this microbiota is likely to confer protection against immune-mediated diseases. However, sufficient contacts with the diverse microbiota in our environment have also been thought to be equally important. It is suggested that the rising incidences of immune-mediated diseases such as allergies and asthma in the western urban society having less proportion of natural environment could be due to insufficient contacts with environmental microbes. This thesis aimed to find out the effect of living environment, including the coverage of built area, vegetation type and pollution, on environmental microbiota, their human exposure as well as the composition of human commensal microbiota. This thesis also aimed to study whether applying soil and plant based materials on skin is effective in establishing better contact with microbes. The effect of pollution on the composition of the environmental microbiota was studied by contaminating four different types of soils with polyaromatic hydrocarbons (PAHs). It was found that PAH-pollution altered the community composition of bacteria, increased the abundance of bacterial phyla shown to be unfavorable and reduced the abundance of those recognized to be favorable for the human immune system. Since exposure to environmental microbes affects human microbiota and the immune function, pollutants such as PAHs likely affect human exposure to environmental microbes and also lead to disturbances in the composition of the human commensal microbiota. The effect of land cover type on indoor exposure to environmental microbes was investigated by studying the composition of bacteria in the debris deposited in experimental doormats installed inside the main door of urban and rural houses in southern Finland. The quantity of environmental litter and bacterial diversity was significantly less in the doormats of houses in heavily built urban area compared to those in sparsely built rural areas. Stool samples from elderly volunteers living in those houses were analyzed to investigate the effect of land cover type and garden vegetation diversity on the composition of gut microbiota. The number of shrub and non-woody flowering plant species in the yard were associated with an increased abundance of several bacterial taxa known to promote the human immune system. The effect of built area was similar to the shifts in the gut microbiota observed in urban inhabitants from highly industrialized areas relative to rural hunter-gatherer population in previous studies suggesting that high incidences of immune disorders in western population could be associated with a greater coverage of built area. Urban volunteers were asked to apply soil and plant-based materials on their hands to see if the use of natural ingredients is effective in changing the composition of bacteria on the skin. This led to a significant increase in the diversity and biomass as well as the relative abundance of several bacterial taxa suggesting that microbial exposure via skin could potentially be employed as a route for preventing or even treating immune-mediated disorders by increasing the diversity of skin microbiota, which needs to be investigated. This thesis concludes that urban living environment characterized by increasing proportion of built areas and high pollution levels could lead to adverse changes in the environmental microbes, reduced indoor exposure to those microbes as well as unfavorable shifts in the composition of the gut microbiota. Shrubs and flowering plants could contribute positively to gut microbial homeostasis. Likewise, a direct contact with soil and plant based materials can be effective in increasing the diversity of the skin microbiota.
  • Trotta, Luca (Helsingin yliopisto, 2019)
    Primary Immunodeficiency (PIDs) categorize a broad and heterogeneous group of inborn immunity errors. Despite being generally quite rare, PIDs collectively account for consistent morbidity and mortality. Currently, more than 350 monogenic PIDs have been recognised to embody clinical phenotypes ranging from life-threatening infections to autoimmune/inflammatory diseases, allergies and/or malignancy. Many PIDs display genetic and allelic heterogeneity with an overlap of symptoms among different syndromes, often making diagnoses challenging. In the past few years, advancements in genomic technologies have revolutionised the world of genetic testing, and currently, next-generation sequencing (NGS)-based approaches are widely applied to routine genetic diagnostics of human disorders. Among the different methods, whole-exome sequencing (WES) proved highly efficient in revealing the genetic variants behind rare disorders. To further depict the genetics of PIDs, a WES-based approach was carried out, targeting the possible disease-causing variants in Finnish subjects lacking a clinical diagnosis. The cohort included patients with a clinical suspicion of immune or/and haematological disorders (n= 212). In the first study, a Finnish founder mutation in the AICDA gene was identified in patients affected by hyper-IgM syndrome type 2 (HIGM2). The disease is a primary antibody deficiency characterised by early-onset recurrent infections, autoimmunity and an absence/low levels of IgG, IgA and IgE but elevated/normal levels of IgM. The retrieved ancestral founder allele is significantly enriched in Finns compared to other European populations (38.56-fold) and has accounted for all HIGM2 cases diagnosed in Finns thus far. In the second study, biallelic ADA2 mutations that cause a deficiency of adenosine deaminase 2 (DADA2) were identified in seven PID patients, all sharing one of the causal variants, which were significantly enriched in Finns (3.31-fold). DADA2 was originally associated with systemic autoinflammation, polyarteritis nodosa-type vasculitis and mild immunodeficiency. Only a fraction of the identified DADA2 patients presented with vasculopathies. In addition, recurrent haematological manifestations are noted, and for the first time, the occurrence of lymphoproliferation is described for some of the patients, expanding the phenotypic spectrum of DADA2. Finally, novel causal variants in telomere biology disorders (TBDs)-associated genes were identified in three families with heterogeneous phenotypes that lacked the classic clinical pathognomonic signs of telomeropathies. The phenotypes ranged from mild signs of Dyskeratosis congenita (DKC) to SCID. The genetic diagnosis was confirmed by an assessment of shortened telomere lengths in patients. In addition, the spectrum of TBD-associated phenotypes was enlarged, showing variable degrees of cytopenia in some patients. This work attests to the validity of clinical WES testing to identify rare disease-causing variants despite the heterogeneous and/or atypical clinical presentations of PIDs. The achievement of a genetic diagnosis allowed for updating the spectrum of reported phenotypes as well as including atypical clinical presentations that might have otherwise remained undiagnosed. In addition, the enrichment of some rare PID-causing mutations in Finland has been depicted, highlighting the correlation of the population history with the distribution of rare deleterious variants of clinical relevance.
  • Tikhonov, Gleb (Helsingin yliopisto, 2018)
    In the last decades, the aims of research in community ecology have been shifting from the mere description of observed patterns towards a mechanistic perspective that seeks to understand the processes shaping observed species communities. Simultaneously, the technical advances in data collection techniques dramatically raised the amount and quality of ecological data annually obtained and provided opportunities to address more comprehensive research questions. The combination of these novel aims and data increased the interest in the statistical ecology, seeking analytical methods capable to harness the full potential of the emerging data. A special interest has been focused on the development of approaches capable to combine multiple types of existing data and jointly model the dynamics and distributions of entire species communities or ecosystems. This doctoral thesis contributes to the ongoing methodological development of analytical tools for the joint species modeling. In the presented research I combine both perspectives of the statistical ecology: the ecologist’s practical point of view and the statistician’s methodological/theoretical vision. The thesis consists of four Chapters that are arranged to form a coherent narrative. I start with a synthesis of the recent advances in joint species modeling and propose a unifying statistical framework that enables scientists to easily address many common questions in community ecology simultaneously. This framework, called Hierarchical Model of Species Communities (HMSC), is capable to incorporate information on species occurrences, environmental covariates, species traits and phylogenic relationships, as well as the structure of study design. Next, I devise and present two important extensions to this framework. The first extension enables HMSC to neatly assess the variation in species associations and relate it to environmental factors. My second extension aims to achieve better numerical properties for the HMSC-based analysis of numerous spatial observations. I carry out a set of simulated data experiments to assess the performances of the proposed extensions in comparison to existing methods. To demonstrate how the proposed methods can be used in practice, I accompany these methodological developments with real-data examples and additionally present one detailed applied ecological study. My results demonstrate that the unifying HMSC framework can be robustly used to address a wide set of fundamental and applied ecological questions for various natural systems and contexts. Conducted simulation experiments verify that the proposed extensions considerably expand the framework’s potential. The developed software implementation of the HMSC and detailed user manual provide a practical guidance for ecologists on how to apply this framework for analysis of their own data on species communities. Although this thesis is a completed research item, it should be seen as a solid foundation for further developments in the field of joint species modeling. Some of these potential developments are related to how more comprehensive ecological questions could be answered with statistical models, while other correspond to the numerical challenges posed by emerging types and amounts of ecological data. I believe that advances and results of my study will enable future research to tackle these challenges and that the joint species modeling framework will become generally applicable and insightful for a wide array of real-world problems.
  • Ilves, Marit (Helsingin yliopisto, 2018)
    Nanotechnology is exerting a huge impact on various sectors of everyday life as it has a tremendous potential for revolutionizing a long list of consumer products and industrial applications. The key to success in the nanotechnology field lies in the fact that materials at the nanoscale possess novel and enhanced properties such as greater strength and improved conductivity when compared with their bulk-sized equivalents. The most probable occupational and consumer routes of exposure to engineered nanomaterials (ENM) are via the respiratory tract and skin. Due to their small size, ENM are able to bypass physical and chemical barriers in the human body and come into contact with the immune system which is capable of recognizing foreign structures including ENM. However, the downscaling of the materials also increases their chemical reactivity, which in combination with the small size and other physicochemical properties, means that ENM could influence our immune system exerting possibly beneficial but also adverse effects on our health. The aim of this thesis was to investigate modulatory effects and physiological outcomes of ENM on a healthy and a compromised immune system in the lungs and skin. The main findings of the thesis were that rigid, rod-like but not long and tangled carbon nanotubes (CNT) were able to induce a condition similar to allergic airway inflammation via activation of innate immunity. Although nanofibrillated celluloses triggered acute pulmonary inflammation, their effects subsided within one month and regardless of the material’s biopersistence, their health outcomes differed significantly from the long-term pathologies of rigid, rod-like CNT. Uncoated and functionalized CuO nanomaterials demonstrated an ability to worsen allergic asthma by eliciting pulmonary neutrophilia, however it was found that surface PEGylation significantly suppressed the inflammatory potential of the pristine CuO ENM; this effect was especially evident at the transcriptional level. Topical exposure to nano-sized ZnO in a murine model of atopic dermatitis revealed that the particles were able to pass through mechanically injured allergic skin. This penetration of the material resulted in a local inhibition of pro-inflammatory and allergic reactions and a systemic exacerbation of IgE antibody production. This work provides knowledge of pulmonary and dermal effects of ENM. The results of this thesis demonstrate that ENM with different physicochemical characteristics possess an ability to modulate our immune system. These observations emphasize the diversity and complexity of the materials as well as highlighting their impacts on the immune system and the resulting consequences on health. These data contribute to the safety assessment of ENM as well as information that can be useful in nanomedicine.
  • Saulamo, Kari (Helsingin yliopisto, 2018)
    Fisheries management needs to address an array issues to achieve long-term sustainability. Economically viable fisheries require healthy fish stocks. Our knowledge of fish populations is still limited, as fish cannot be observed directly and they are typically highly mobile. Fish populations also fluctuate temporally and spatially depending on the biophysical characteristics of the environment and interactions in the ecosystem. By increasing our knowledge of the ecology of exploited species and how they respond to the impacts of fishing, we increase the probability of success with the management of the fish populations. The main contribution of this thesis is improved scientific understanding of the biology of the pikeperch and how this relates to the management of fisheries. In the thesis, the movements of the pikeperch are studied during the spawning season to gather information on biological characteristics, such as changes in size distribution, sex ratio and maturity between fish ascending to the spawning grounds at different times. The interaction between biology and the management of the pikeperch is studied by analysing the situation in where pikeperch are exploited by several management units. The effects of high fishing pressure and selective fishing on growth and the production of pikeperch stock are also studied. It is demonstrated that there are differences in the biological characteristics between pikeperch arriving at the spawning ground at different times, and signs that there can be early and late spawners in the populations. It is also shown that in cases where the pikeperch migrates over several management areas, overfishing is inevitable if there is no agreed cooperation between the areas. It is proven that excessive fishing pressure with highly selective fishing gears creates strong phenotypic selection for slower growth in the studied pikeperch population. Fisheries managers are advised to consider the spatial and temporal dynamics of the pikeperch stocks in order to protect the different stock components. Pikeperch managers should focus on distributing harvesting with caution regarding space, time and also between age groups to avoid adverse ecological effects from fishing, and to avoid the risk of decreasing the long-term productivity of the stock.
  • Jahan, Farhana (Helsingin yliopisto, 2018)
    Inflammation is a defense response of the body to an event of injury or tissue damage. Leukocytes circulating in the bloodstream are stimulated by chemokines released by endothelial cells that enable them to adhere and transmigrate to the site of infection. Leukocyte extravasation from the blood vessels towards the site of inflammation is a sequential and overlapping process involving leukocyte rolling, adhesion and transendothelial migration. Adhesion is critical for T-cell trafficking and antigen recognition and is mediated in part by integrins, a large family of αβ heterodimeric cell surface proteins. The β2-integrin lymphocyte function-associated antigen-1 (LFA-1 /αLβ2) and the β1-integrin very late antigen-4 (VLA-4/α4β1) promote T cell interactions with their ligands intercellular and vascular cell adhesion molecules (ICAMs and VCAMs) respectively which are expressed on endothelial cells. VLA-4 and LFA-1 directly participate in cell arrest under flow, whereas firm adhesion is mediated by LFA-1. Integrins have unique characteristics of signaling in both directions across the membranes. Inside-out signaling occurs by ligand binding to non-integrin receptors, which can activate integrins through intracellular signal transduction whereas outside-in signaling takes place by binding of ligands to the integrin itself. The complex signaling pathways of LFA-1 are regulated by rapid phosphorylation and dephosphorylation events of LFA-1, which affect its binding affinities and subsequently protein-protein interactions. My interest lies how the activation of LFA-1 by inside-out and outside-in signaling affects the phosphorylation of the αL- and β2-chains, the adhesive property of LFA-1, intracellular binding partners and cross-talk with the VLA-4 integrin. In my study, I have found involvement of Tiam1, a Rac GEF (Guanine nucleotide exchange factor) protein as a new key player in the intracellular signaling of LFA-1. Upon LFA-1 activation by inside-out signaling, Tiam1 forms a complex with β2 phosphorylated on Thr-758 and the scaffolding protein 14-3-3, which activates Rac1, an actin-regulating protein. Downregulation of Tiam1 inhibits Rac1 activation and furthermore, it impairs cell adhesion to the LFA-1 ligand ICAM-1. The next finding was that the activation of LFA-1 via inside-out or outside-in signaling downregulates VLA-4 binding to its ligand VCAM-1. When LFA-1 is activated, β2 is phosphorylated on Thr-758, which recruits 14-3-3 and Tiam1 and results in crosstalk to VLA-4. It leads to dephosphorylation of Thr-788/789 on the β1-chain, which results in the recruitment of more filamin but less 14-3-3 to the β1-chain. I have also demonstrated that the activation of LFA-1 by anti-LFA-1 antibodies binding to the extracellular part is mediated through phospholipase C (PLC) and protein kinase C (PKC) activation, which causes the phosphorylation of the Thr-758 on β2-chain. Furthermore, I have shown that the phosphorylation of the LFA-1 αL-chain at Ser-1140 is essential for the functionally significant phosphorylation of the β-chain Thr-758 and protein interaction with the β2-chain. Mutation of Ser-1140 to alanine in the αL-chain significantly decreased Thr-758 phosphorylation of β2-chain after SDF-1α or anti-CD3 activation. α-Actinin is an actin-binding protein, which links integrins to the actin cytoskeleton and is crucial for cell migration. We showed that mutation of Ser-1140 to alanine in the αL-chain affects the α-actinin interaction with the β2-chain and chemotactic directional migration. Phosphorylation and de-phosphorylation of αL- and β2- chains may be a way to regulate adhesion and deadhesion turnover of cells during migration. De-regulation of integrins’ function may lead to auto-immune and chronic inflammatory diseases. Here in this thesis, I have now revealed some novel, important mechanisms of the regulation of the leukocyte integrins. The knowledge of how integrins are regulated could be beneficial in clinical applications as well as for developing drugs with high specificity in future.
  • Perttilä, Niko (Helsingin yliopisto, 2018)
    Background: The term frailty is used to describe older persons who are at increased risk of adverse health outcomes. Various measures have been investigated but Fried’s criteria are used most often. However, there is still a lack of consensus on the definition of frailty. Simpler measures based on postal questionnaires have also been investigated in the assessment of frailty. Different measures may only partly identify the same persons as frail, and in addition the predicted outcomes may vary depending on the measure used. Older persons in general and older frail persons have benefited from exercise interventions. Some studies have also revealed that persons with dementia may benefit from exercise intervention. However, there is a lack of evidence on whether or not the stage of frailty affects the outcomes of exercise intervention among persons with dementia. Among older persons in general, known risk factors of falls are, for example, a history of falls, muscle weakness, gait deficit, arthritis, depression, polypharmacy, psychotropic medication and older age. Fewer studies have concerned risk factors of falls among persons with dementia. No study has been carried out to investigate possible interactions between exercise, fall-related drugs and falls among people with dementia. Aims: This study aimed to investigate how various frailty measures overlap, whether or not they identify the same persons as frail, and to explore if the outcomes are similar irrespective of the used measure. This study also investigated whether or not the frailty status of community-dwelling persons with Alzheimer Disease (AD) affects the outcomes of exercise intervention. In addition, this study explored risk and protective factors associated with falls among persons with AD and whether there are interactions between exercise intervention, fall-related drugs, and falls in this population. Subjects and methods: The subjects in Study I were the participants of the Helsinki Businessmen Study, which is a long-term observational study of men born in 1919–1934. These men were investigated by analyzing their responses in postal questionnaires in 2000 and 2005. A total of 480 men were included in the study, and their mean age was 73 years at the start of follow-up. Two phenotype-based measures, the Helsinki Businessmen Study (HBS) measure and the modified Women’s Health Initiative Observational Study (WHI-OS) measure, and the Frailty Index (FI), consisting of 20 items, were used to identify frailty through postal questionnaires. The measures were investigated in regard to how they overlapped in identifying robust, prefrail and frail individuals and how they predicted falls, health-related quality of life (HRQoL, 15D instrument), weight change and mortality during a five year follow-up period. The subjects in Studies II, III and IV were the participants of the Finnish Alzheimer disease exercise trial (FINALEX), which was an exercise intervention study among home-dwelling persons with AD. The mean age of the participants (N=194) was 78 years and 39% were women. The participants in intervention groups underwent group-based or home-based exercise for one hour twice weekly for 12 months (N=129), and the control group had normal care (N=65). In the FINALEX study, falls were recorded in fall diaries by AD persons’ spousal caregivers during the one-year follow-up period. In Study II, both the combined intervention group and the control group were subdivided into prefrail (0–1 criteria) and advanced frailty (2–5 criteria) according to modified Fried phenotypic criteria. The number of falls per person-years and changes in the Functional Independence Measure (FIM) served as outcome measures. In Study III, all participants were investigated together to reveal fall risk factors in connection with physical functioning, diseases and drugs. Study IV investigated possible interactions between exercise and fall-related drugs by comparing the incidence rate ratios (IRRs) of falls among non-users and users of various drugs in intervention and control groups. Results: Both the HBS measure and the WHI-OS measure identified 7.3% of the participants of the Helsinki Businessmen Study as frail, but only partly were they the same individuals. The FI measure identified 17.9% of the participants as frail. Altogether, 21.3% of the participants were classified as frail by at least one measure. All measures (HBS, WHI-OS, FI) predicted a significantly higher number of fallers, lower HRQoL, and higher mortality among frail participants than among prefrail and not frail participants. There were no differences as regards weight changes between the frailty stages or frailty groups. In the FINALEX study, in the prefrail groups the average rate of falls was significantly lower in the intervention group than in the control group (1.14 falls/person per year [95% confidence interval (CI): 0.90 to 1.43] and 1.82 falls/person per year [95% CI: 1.40 to 2.32], respectively; IRR 0.63 [95% CI: 0.45 to 0.89]; P=0.008 adjusted for sex, age and comorbidities). In the advanced frailty groups also, the participants in the intervention group had a significantly lower rate of falls than those in the control group (2.15 falls/person per year [95% CI: 1.76 to 2.59] and 5.32 falls/person per year [95% CI: 4.36 to 6.44], respectively; IRR 0.43 [95% CI: 0.33 to 0.57]; P<0.001 adjusted for sex, age and comorbidities). Both intervention groups also deteriorated significantly more slowly than their respective control groups according to changes in FIM scores. Better physical functioning and robustness protects against falls among persons with AD. Diseases such as chronic obstructive pulmonary disease, diabetes and arthritis, as well as drugs such as opioids, psychotropics, drugs with anticholinergic properties, and polypharmacy were associated with an increased risk of falls. Use of antihypertensives was associated with a reduced risk of falls among those undertaking exercise, but there was an increased risk of falls in the control group. Exercise also reduced fall risk among users of psychotropics. Conclusions: The HBS measure, the WHI-OS measure and the FI measure identified frail participants, but there was only a partial overlap. The FI identified more persons as frail. All three measures predicted increased falls and mortality and reduced HRQoL among the frail participants. People with AD may benefit from exercise intervention irrespective of their frailty status in respect of falls and physical functioning. Better physical functioning and robustness protect against falls. Several diseases and drugs were risk factors of falls. Exercise favorably interacted with antihypertensives and psychotropics as regards the risk of falls.
  • Paukkunen, Juho (Helsingin yliopisto, 2018)
    Cuckoo wasps (Chrysididae) include more than 2500 described species, all of which are parasitoids or kleptoparasites of other insects, predominantly solitary wasps and bees. They favor warm, dry and sunny habitats, and their bodies are typically highly sculptured with bright metallic colours. About 490 species are currently known in Europe, but the actual number of species is estimated at 550-600. The objectives of this study were to: 1) to determine which species of cuckoo wasp occur in Northern Europe, i.e. Fennoscandia, Denmark and the Baltic countries, 2) to assess their valid names according to the International Code of Zoological Nomenclature, 3) to present an identification key for all species, based on morphological characters and 4) to study their distribution, abundance and long-term population trends. The structure, distribution, and abundance of cuckoo wasps were studied on the basis of museum and private collections, and also all published data of Northern European taxa were compiled. The validity and phylogeny of the morphologically similar species of the Chrysis ignita group were investigated by analyzing their mitochondrial DNA. The population trends of Finnish cuckoo wasps were assessed by studying changes in the occupancy of 10 x 10 km grid squares between two periods, 1840–1967 and 1968–2015. Comparative analysis was performed to investigate the correlation between the population trends and shared species traits. A total of 74 cuckoo wasp species were found in the study area, four of which were new to Northern Europe, and one, Chrysis borealis Paukkunen, Ødegaard & Soon, 2015, also new to science. Cleptes striatipleuris Rosa, Forshage, Paukkunen & Soon, 2015 was described from Central Europe. Chrysis terminata Dahlbom, 1854 represented the valid name for C. ignita Form A, and Cleptes semiauratus (Linnaeus, 1761) was re-established as a senior synonym of Cleptes pallipes Lepeletier, 1806. Two new synonyms were also found. Mitochondrial DNA sequences supported the validity of the most previously recognized species of the Chrysis ignita group and additionally suggested the existence of several cryptic species. Decreasing populations were found in 11 Finnish cuckoo wasp species, while significant increases were not observed for any species. Scarce and small-sized species that are dependent on sun-exposed dead wood had declined more than abundant and large-sized species that live in open sandy habitats. This study provides a solid basis for the taxonomy of Northern European cuckoo wasps and facilitates species identification with an illustrated determination key. The declining populations of many species indicate that cuckoo wasps are vulnerable to human-induced environmental changes and conservation measures are needed for their preservation.
  • Oghabian, Ali (Helsingin yliopisto, 2018)
    In the Eukaryotes, DNA sequences in genes are often interrupted by non-coding sequences called introns. These sequences are removed from the transcripts via a process known as splicing either while the genes are being transcribed (co-transcriptionally) or after transcription (posttranscriptionally). In higher eukaryotes two separate pre-mRNA splicing machineries have been described: the U12-dependent spliceosome which is responsible for splicing of approximately 700-800 unique introns (known as the U12-type introns), and the U2-dependent spliceosome responsible for splicing all other introns (known as the U2-type introns). The two intron types show divergent sequence elements in their 5' splice site and branch point sequences. In addition, earlier reports have indicated that U12-type introns are spliced with a slower rate comparing to the U2-type introns, suggesting that the splicing of U12-type introns is rate-limiting to the expression of the U12-type intron containing genes. This slower splicing is manifested as unspliced or retained U12-type introns in the otherwise fully processed mRNA products. In this work I developed a novel computational tool called the intron-exon retention estimator (IntEREst) which allows accurate detection, quantification and differential analysis of the intron retention levels from RNAseq data. Additional features of IntEREst include a tool for identification of U12-type introns, and a number of tools to compare the retention levels of userdefined subclasses of introns across several samples. An already published RNAseq dataset (available under accession GSE63816 in NCBI Gene expression Omnibus database) from patients and control subjects of myelodysplastic syndrome (MDS) was used to assess the functionality by benchmarking IntEREst. This dataset included RNAseq data from MDS patients featuring mutations in the ZRSRS2 gene that functions in the recognition of U12-type introns, and from control subjects that were either healthy or MDS patients without ZRSR2 mutations. Additionally, I used a Maize dataset consisting of samples with mutated and wild-type RGH3 gene, which is an ortholog of human ZRSR2. My results indicate that IntEREst is a reliable tool for analyzing intron retention events from RNAseq data producing comparable or better results than the other similar methods. I used IntEREst to globally compare the retention of the U12-type introns to that of U2-type introns. I found that U12-type introns show on average a 2-fold higher retention levels compared to that of U2-type introns both in human and plant cells. This result recapitulates the findings from earlier studies using a small set of selected genes and generalizes the increased intron retention of U12-type introns to a genome-wide scale. Furthermore, the results of this work provide evidence that transcripts containing unspliced U12-type introns are degraded in the nucleus by the nuclear exosome. Together, these results support the hypothesis that U12-type introns are globally spliced less efficiently than the U2-type introns and can thus regulate the rate of mature mRNA formation with the genes containing U12-type introns. Additionally, intron retention analysis of human/plant cells containing mutations in the U12-dependent spliceosome showed that such defects lead to a further increase in the levels of unspliced U12-type introns. In conclusion, this thesis extends current knowledge concerning the significance of the correct splicing of U12-type introns and the consequences of their abnormal splicing. Furthermore, it describes a combination of available tools together with a novel software tool (i.e. IntEREst) that can be used to measure and compare the efficiency and accuracy of RNA splicing across multiple samples. We show that these tools can reveal valuable information about the molecular mechanisms involved in various conditions, e.g. diseases caused by defective spliceosome.