Browsing by Title

Epigenetics is the study of changes in gene expression without alterations in the DNA sequence. Epigenetic modifications, of which DNA methylation (DNAm) is the most known, are crucial for many biological events, especially for normal development and genomic imprinting. In imprinted genes, only one of the parental alleles is consistently monoallelically expressed. The epigenome can be altered by various environmental factors such as diet and chemicals. Moreover, evidence indicates that assisted reproductive technology (ART) is associated with distinct DNAm patterns. Still, it is unclear whether these alterations are a consequence of ART procedures themselves or the underlying subfertility. This thesis aimed to study whether ART procedures and subfertility are associated with aberrant DNAm at the imprinted DLK1-DIO3 locus in the human placenta. The genes encoded from this paternally imprinted locus are essential in mammals' fetal and placental development. Moreover, recent evidence suggests that downregulation of the DLK1 gene is associated with ART and subfertility. Therefore, two gene regions, the IG-DMR and the DLK1 promoter, were chosen as study objects. The IG-DMR acts as the main regulator of gene expression at the whole DLK1-DIO3 locus, and promoter regions are considered important regulators of the expression of their corresponding genes. Eight ART, four subfertility, and six control samples of human placental DNA were studied. The subfertility group consisted of couples who were committed to initiating fertility treatments but eventually got pregnant spontaneously. The study was performed using traditional bisulfite sequencing, after which the differences in DNAm levels between study groups were statistically analyzed. As expected, significantly decreased DNAm level was observed in the placentas of subfertile couples compared to controls. Surprisingly, no significant differences were addressed between ART and control groups in either region. The partly unexpected results are explained by the fact that aberrant methylation at distinct imprinted DMRs, including the IG-DMR, is caused by the in vitro culture media. Moreover, the results indicate that the regulation of DLK1 expression is more complicated than that solely by the IG-DMR and the DLK1 promoter. Further research should be dedicated to differentiating the impacts of ART and subfertility on the epigenome and phenotype to better understand the health implications of ART.

The brainstem monoaminergic neuronal systems are involved in regulation of mood, reward system, memory processing etc. Any defects or damage in these cells lead to many neurological disorders. The brainstem inhibitory GABAergic and excitatory glutamatergic cells in turn control these neuromodulatory neurons. The glutamatergic neurons are found in the Laterodorsal tegmental nucleus (LDTg), Interpeduncular nucleus (IPN) as well as in the Ventral tegmental are (VTA). The LDTg in particular sends these glutamatergic projections to the VTA to regulate their Dopaminergic (DA) neurons. During embryonic development, the brainstem GABAergic and glutamatergic neurons, that regulate the monoaminergic systems, are produced in the ventral rhombomere 1. Their subtypes are known to express various transcription factors (TFs), such as Nkx6-1, Vsx2 and Skor1 marking the glutamatergic neuron precursors in the ventral rhombomere 1. In this thesis project, I studied the expression of another TF, Skor2 in the embryonic brainstem precursors. The basis of the experiment came from an embryonic brainstem single cell mRNA sequencing study performed earlier, where Skor2 expression was observed in the cluster of neurons containing Nkx6-1, Vsx2 and Skor1 expressing cells. Based on this, I hypothesized that Skor2 expression could be seen in glutamatergic precursors in the ventral rhombomere (rV2) domain as well as later in the LDTg nucleus derived from these precursors. To test this, I performed immunohistochemistry (IMHC) studies on a transgenic mouse line expressing Green Fluorescent Protein (GFP) from the Skor2 locus. In the second part of the thesis, I hypothesized that the Skor2 positive cells need this TF for their differentiation. To study this aspect, I performed similar IMHC studies on homozygous Skor2GFP/GFP mice, where Skor2 had been inactivated. My study showed that Skor2 positive cells expressed markers Nkx6-1 and Vsx2 and represented a specific subgroup of early embryonic post-mitotic precursors in the rV2 domain. Later in the brainstem, in contrast to my initial hypothesis, I did not observe Skor2 expression in the LDTg glutamatergic region. Instead, I observed Skor2 positive cells in a region more lateral to the Ventral and Dorsal tegmental nuclei of Gudden. In the homozygous Skor2 mutants, I observed no changes in cell fate during embryonic development. Based on my results, the TF Skor2 is expresses in the glutamatergic precursors and neurons in the rhombomere 1, but form a part of a new cluster of cells away from the LDTg. These neurons have not been studied in detail. However, the Ventral and Dorsal tegmental nuclei of Gudden have been shown to regulate memory and navigation. It is possible that the Skor2 expressing neurons also participate in these functions. Identification of specific molecular markers, such as Skor2, for these neurons now allows their focused functional studies. Skor2 and Skor1 are related TFs belonging to Ski family of transcriptional repressors and are seen to be expressed together. Further investigations into the roles and functional redundancy of these two TFs can be performed using mice carrying mutations in both of these genes.

The significance of adhesion to the basement membrane (BM) for the growth and survival of healthy mammary epithelial cells is well-established. In ductal carcinoma in situ (DCIS), a non-invasive form of breast cancer, luminal cells detach from the BM and proliferate within the ductal lumen of the mammary gland. This suggests that malignant cells are capable of evading detachment induced cell death, called anoikis during DCIS. The alterations that contribute to anoikis resistance may be essential for anchorage-independent survival, a phenomenon that remains largely unexplored in the context of BM. Hence, in this study, I aimed to investigate the association between anchorage-independent survival of breast cancer cells and the expression of BM components. I utilized a low-adhesion 3D suspension culture devoid of any matrix to evaluate the efficiency of mammosphere formation in luminal A and triple-negative breast cancer cell lines. To understand the ability of cancer cells to survive in an anchorage-independent environment, I investigated the expression profiles of BM components (including laminins and type IV collagen) across various breast cancer cell lines. Gene expression analysis of BM components was conducted in conventional 2D monolayer cultures, while immunofluorescence analysis was performed on mammospheres or single cell clusters derived from low-adhesion 3D cultures. Furthermore, to explore their functional significance, I examined the effect of laminin α5 (LAMA5) downregulation on the spheroid formation ability of luminal A cell line ZR75-1. The primary results of the study highlighted the ability of breast cancer cells to survive in low anchorage conditions by forming mammospheres or as single-cell clusters. Significant variations in the expression of BM components were observed across breast cancer cell lines and within the subtypes. Moreover, I observed the atypical expression and localization of BM components produced by the various breast cancer cell lines in the 3D culture. The potential functional relevance of BM was demonstrated by the decreased mammosphere-forming ability of the ZR75-1 with LAMA5 downregulation. Although further investigations are needed, these observations suggest a crucial role of BM produced by tumour cells in facilitating anoikis resistance and anchorage-independent survival of cancer cells.

Master's thesis project includes the backbone assignment of the human activity-regulated cytoskeleton-associated protein C-lobe (hArc, Uniprot ID: Q7LC44), 7-fluoroindole-based tryptophan-labeling method, and comparing that with the 100% double-labeled and 20%(13C) fractionally labeled samples. The project focuses on the effects of 7-fluoroindole-based fluorotryptophan-labeling. hArc C-lobe has only one tryptophan, which makes the analysis easier. Typically fluorotryptophan-labeling is a costly method – fluorotryptophan itself is very expensive and attaching the fluorine to the tryptophan while expressing is expensive and complicated. Fluoroindolebased labeling circles around the problem, as indole and serine are used in procaryotic systems for tryptophan biosynthesis – meaning that fluoroindole, which is cheap, could be used as an alternative for previous methods. Fluoro-labeled tryptophan is used in protein NMR; for example, in binding studies – fluorine-probes are sensitive, and binding of ligand or protein would move these peaks, indicating binding. This project aims to get an insight into the application of this labeling method. The goal is to see if one could utilize one sample with both (1H, 15N, 13C) labeling and 7-fluorotryptophan labeling for binding and structural studies. However, fluorine is very electronegative, affecting surrounding structures and possibly sequentially nearby amino acids. This possible effect will be observed and determined by comparing the 1H15N-chemical shifts between well-established labeling methods and fluoroindolebased labeling. To determine what amino acids in the protein are affected, if they are affected, will be determined by using the backbone assignment results and the results from the sample comparisons.

Ecosystem accounting is a new framework for integrating the value of nature into decision-making. Previous measures of natural capital accounting have not been able to achieve policy relevancy and have only limited use cases. It is important to look into the usability of ecosystem accounting, to ensure that it is implemented in a way that supports decision-making. The goal of my research is to provide a categorization of barriers and enablers of use, as well as provide some insight into how to make ecosystem accounting usable for decision-making. There is only limited literature on ecosystem accounting, so I conducted a scoping literature review on barriers and enablers of use of natural capital accounting to look into the issues and opportunities in making ecosystem accounting usable. I complemented this literature review with interviews of ecosystem accounting experts from Norway and Germany. Both countries have started the implementation journey and thus provide a view into the active development phase. My main results are the categorization of barriers and enablers of use of ecosystem accounting, as well as main messages for creating usable accounts. I categorized the barriers and enablers into 12 categories under three themes of political, structural and relevancy related barriers and enablers. The categories can be used when planning implementation to identify potential issues and to plan accounts to be as usable as possible. I have three main messages for compiling usable ecosystem accounts. First, sufficient resources are essential in gathering comprehensive accounts. Second, compiling the accounts rely on cooperation of multiple institutions, thus there is a need to agree on forms of coordination. Third, the categories of barriers and enablers are interconnected, and thus there are positive feedback loops that can support making ecosystem accounts usable for policymaking.

Oil spills in aquatic environments are devastating disasters with both biological and economic impacts. Fish populations are among the many subjects of these impacts. In literature, there are numerous assessments of oil spill impacts on fish populations. From all applied research methods, the focus of this thesis is on Bayesian methods. In prior research, several Bayesian models have been developed for assessing oil spill impacts on fish populations. These models, however, have focused on the assessment of impacts from past spills. They have not been used for predicting impacts of possible future oil spills. Furthermore, the models have not utilized data from laboratory studies. Some examples can be found of models assessing economic impacts of oil spills on fish populations however, none of them assess the economic impacts that follow from decreases in biomass. The aim of this thesis is to develop a Bayesian bioeconomic prediction model, which would be able to predict oil spill impacts on Baltic Sea main basin herring population, and the consequential economic impacts on fishermen. The idea is to predict the impacts of several hypothetical oil spill scenarios. As a result of this thesis, a bioeconomic prediction model was developed, which can predict both biological and economic impacts of oil spills on Baltic Sea main basin herring through additional oil induced mortality of herring eggs. The model can be applied to other fish populations in other regions as well. The model utilizes laboratory studies for assessing population level impacts. The model can be used for both assessing risks of the impacts of possible future oil spills, and for decision analysis after a spill has already occurred. Furthermore, the model can be used for assessing unknown aspects of past oil spills. The economic predictions can be used, for example, to estimate the compensations that could possibly be paid to fishermen. In the future, the prediction model should be developed further, especially regarding its stock-recruitment relationship assumptions. In addition, the model’s assumptions regarding the calculation of oil induced additional mortality and the economic impacts, should be expanded.

Forest fires are a growing issue of concern because of climate change. Forest fires impact forest ecosystems partially or completely and can cause negative and positive consequences. Negative consequences include, for instance, carbon release, and positive consequences include, for instance, increase in biodiversity. Forest fires are prevented by various measures, among which there are those targeted at human behavior. The aim of this thesis is to research current forest fire prevention strategies of Finnish national parks, find gaps and problems, and suggest possible solutions. The thesis focuses on the five largest Finnish national parks ac- cording to the number of visits, because the parks can demonstrate examples of education and modification of people’s behavior. The focus is on human behavior because the majority of forest fires are human-caused. The research questions of the thesis are: 1) “What are the behavior-oriented forest fire prevention measures in Finnish national parks?” and 2) “What are the problems of the current prevention measures?” The data of this study was collected via semi-structured interviews with national parks' workers who have knowledge about the topic. This method was chosen because interviewees can provide explanations and case examples. Five interviews were done via Zoom or Teams, while two responses were received via email. The content analysis of the data was performed in qualitative data analysis software ATLAS.ti. The theoretical frame- work of the thesis is risk governance and, more precisely, risk management and communication. In addition, the framework includes prevention practices identified in previous research. Behavior-oriented forest fire prevention measures identified in this thesis classify as risk management and risk communication. Risk management includes construction and maintenance of safe facilities, forbidding unsafe campfire ignition methods, and regulations mentioned online and on-site. Risk communication, in turn, includes safety instructions, wildfire warnings, organized activities, promotion of visits without campfires, nudge, and verbal advice. The problems related to the measures are making campfires when and where it is forbidden, irresponsible behavior in the context of ashes and rubbish, and leaving fires unattended. In addition, problems are related to how and where forest fire safety information is provided. The knowledge and findings of this study can be used to improve current forest fire prevention measures. Possible solutions include delivering explanations of measures’ reasons to visitors, highlighting the urgency of a problem, and organizing more educational group activities.

The shallow bays and inlets of the coastal Baltic Sea consist of a patchy seafloor with different sediment types that are affected by anthropogenic pressure, such as organic matter loading. Benthic macrofauna affects the carbon and nutrient stock capacity of such coastal areas by storing these elements in their body tissues and thus working as a temporary carbon and nutrient pool. These nutrient stocks are especially stable when the animals are large and long-lived. Fluctuations in organic input have been suggested to modify the diversity, abundance, and biomass of the benthic communities and therefore affect the nutrient reserves bound within the fauna. However, there is little information on how stress, in form of increasing sediment organic matter content, change the stocks and stoichiometric ratios of benthic macrofaunal communities. This thesis investigates if the changes in sediment organic matter affect benthic community composition and organism size and its ability to function as a carbon (C), nitrogen (N) and phosphorous (P) pool by measuring: 1) how the benthic faunal diversity, abundance, and biomass composition and 2) the C:N:P content and ratios of the benthic fauna change between three different sediment categories. A significant difference was found between sandy and muddy sediments in terms of benthic biomass and abundance, as well as the carbon and phosphorus stocks. Large long-living bivalves were found in sandy sediments, but overall organism size was not significantly different between the sediment categories. In general, the results show that higher sediment organic matter content resulted in fewer individuals, biomass, and thus a smaller storage capacity of carbon and nutrients. No statistical differences were found between the C:N and N:P and C:P ratios along the three sediment types. However, somewhat more P was bound within the fauna at sites with more organic matter, compared to the sandy and mixed sediments. The results highlight the importance of macrofauna in the functional performance of the ecosystem function and their potential for shifts with increasing disturbance. In conclusion, the increase in sediment organic matter can significantly change the capacity of macrofaunal communities to bind carbon and nutrients and this might have implications for the coastal nutrient filter. More research regarding the fauna's ability to adapt to the changing environment is needed.

The goal of this study has been to compile and to further develop a collection of biodiversity indicators for Finland. Biodiversity indicators aim at bringing forth information related to biodiversity change and the factors causing it in a condensed and easy to understand manner. The factual information which indicators provide is supposed to appeal to the reader's intellect and thus to have an influence on her opinions and behavior. Biodiversity indicators also aim at influencing biodiversity related politics. Being successful at this, biodiversity indicators can make a strong contribution towards reaching the most important goal of nature conservation - the perpetuation of natural variety. The study is divided into two parts. The first part presents a preliminary collection of 75 biodiversity indicators. These indicators are based on the biodiversity monitoring and research data as well as other statistical data which were readily available at the time of their composition. In addition to the statistics and literature reviewed the study is based on interviews of numerous experts. The collection of 75 biodiversity indicators enables a reasonably comprehensive assessment of the major factors effecting biodiversity as well as of the state and development of some components of biodiversity (e.g. endangered species). It, however, contains too few indicators on the state and development of common species and on many important characteristics of ecosystems. The data for such indicators were not available. The second part of the study consists of a review of some European national biodiversity monitoring schemes, international biodiversity indicator development projects as well as Finnish national biodiversity monitoring field programs. Based on the review, recommendations for the further development of the indicator based biodiversity information system are made. The second part of the study concludes with a proposal for an expanded biodiversity indicator collection which contains 31 new indicators. The expanded biodiversity indicator collection could provide a sound factual foundation on which comprehensive assessments of state and development of biodiversity could be based. Such assessments are needed when the effects of biodiversity related politics are evaluated and decisions concerning where to focus conservation efforts are made. At the end of this study possible applications of the expanded biodiversity indicator collection are examined in the light of four different conservation objectives

The need to develop and expand urban areas is increasing in most countries, but urbanization also increases the threat for global biodiversity. Some cities have acknowledged this challenge and formed strategies and action plans for biodiversity preservation. How can we ensure that such strategies are realized in city planning? Negotiations are a crucial part of urban planning, and therefore can be a leverage point of intervention to effectively implement strategies to pro-tect biodiversity. However, little is known about the dynamics of the actual negotiation process in city planning. I applied a game theoretic approach to study how information availability influences the suc-cess and efficiency of negotiations. A role-playing game was used to simulate a negotiation on specific measures to preserve biodiversity in a residential building project. Eleven urban devel-opment specialists played the game with different sets of information. In addition to the direct outcomes of the negotiation, I analysed the post hoc discussion and arguments used to gain in-sights into perceptions of biodiversity-related negotiations in urban planning. Results indicate that information availability can increase the efficiency of negotiations. Partici-pants favour principled and integrative negotiation, but incomplete information seems to push them to take a more positional stance. The post hoc discussion also reveals some issues rele-vant to the design of urban planning process for biodiversity. The overall results suggest that a simple game-theoretic framework, implemented in (a) game-like simulation with quasi-experimental control and (b) qualitative analysis of discussions, holds potential for both under-standing (i) how decision makers frame and resolve the negotiation with conflicting interests and (ii) how to design efficient administrative processes taking into account not only the partic-ipants’ preferences but also wider public interests, such as biodiversity preservation.