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  • Juuri, Juuso (Helsingin yliopisto, 2015)
    Kainate-type of ionotropic glutamate (KA) receptors are associated with the modulation of neuronal excitability, synaptic transmission, and activity of neuronal networks. They are believed to have an important role in the development of neuronal connections. In this thesis, the role of KA receptors in the early brain development was assessed by conducting in vitro electrophysiological recordings from individual neurons at CA3 region in acute slices of neonatal rodent hippocampi. It was found that activation of separate KA receptor populations promoted action potential firing in both glutamatergic pyramidal neurons and GABAergic interneurons. The receptors in pyramidal neurons displayed a high affinity for agonist kainate, appeared to lack subunit GluK1, and promoted spontaneous firing of pyramidal neurons without depolarizing them. The receptors in interneurons contained subunit GluK1 and their activation suppressed afterhyperpolarizing current of medium duration (ImAHP). Receptors in both neuron types appeared to be activated tonically by ambient glutamate, suggesting that their physiological role may be to act as a modulatory mechanism sensitive to changes in extracellular glutamate concentration. Changes in activity of neurons at CA3 by activation of KA receptors were reflected on the network level. Promotion of pyramidal cell firing by pharmacological activation of high-affinity KA receptors lead to enhanced glutamatergic drive and generation of network bursts in the CA3 region. The ImAHP in interneurons was also suppressed by apamin, a blocker of SK potassium channels that mediate majority of this current, and apamin enhanced generation of network bursts. This suggests that also KA receptor mediated regulation of ImAHP may modulate network activity. It was also found that there was an interaction between KA receptors and ethanol in the modulation of hippocampal network: ethanol decreased the occurrence of the network bursts at postnatal days 1 (P1) and P10, whereas it increased bursting at P5. The network effects of ethanol were partially or completely counteracted by specific pharmacological block of GluK1 subunit-containing KA receptors. The findings disclose that via regulation of activity of individual neurons, KA receptors are capable of robust modulation of network activity in immature hippocampus. Additionally, exogenous agents affecting KA receptors may perturb activity dependent developmental processes that are central for the synaptic development. The results shed light on the mechanisms underlying development of hippocampal connectivity, and may help to understand early pathologies of the brain that have developmental origins.
  • Puskarjov, Martin (Helsingin yliopisto, 2013)
    Active extrusion of Cl- from the neuronal cytoplasm by the neuron-specific K-Cl co-transporter isoform KCC2 is necessary for the hyperpolarizing inhibitory Cl- currents mediated by the GABA receptors (GABAARs). Early in development and following cellular trauma or seizures, GABAAR-mediated signaling is often depolarizing and may even, in contrast to its classical inhibitory action, promote action potential firing. Developmental up-regulation of KCC2 is largely responsible for the shift from depolarizing to hyperpolarizing GABAAR-mediated signaling, and conditions associated with brain pathology often lead to loss of KCC2 and re-emergence of depolarizing GABAAR responses. The molecular mechanisms responsible for the up-regulation of KCC2 during development and those mediating its down-regulation, however, remain elusive. The present Thesis demonstrates that the low level of KCC2 protein in immature neurons is not a limiting factor for its functional activation. A single seizure episode induced with kainate triggers a fast transient enhancement of neuronal Cl- extrusion capacity paralleled by a large increase in surface-expressed but not total KCC2 protein in the hippocampus of neonatal rodents. This post-translational activation of KCC2 appears to be mediated by BDNF-TrkB signaling, as evidenced by its sensitivity to Trk inhibition and its absence in BDNF knockout mice. In contrast to these fast changes in functional expression of KCC2, no requirement for endogenous BDNF was observed for the developmental up-regulation of KCC2 protein. Another key finding of this work is that down-regulation and inactivation of KCC2 following intense NMDA receptor (NMDAR) activation is mediated via cleavage and truncation of KCC2 by the calcium-activated protease calpain. Importantly, the data obtained using inhibitors of protein degradation and protein synthesis indicate that the basal turn-over of KCC2 protein is slow and, consequently, down-regulation under pathological conditions is likely to result from enhanced degradation rather than from reduced de novo KCC2 synthesis. Together, the present findings highlight post-translational regulation as an important mediator of changes in the functional expression of KCC2 in response to conditions of enhanced neuronal activity, such as epileptic seizures. KCC2 has been traditionally regarded to have the most clearly defined physio-logical role of all the K-Cl cotransporters, as it is uniquely expressed in central neurons, and determines the neuronal response to activation of GABAA and glycine receptors. However, such a view has changed drastically following the unexpected observation that KCC2 has also a structural role in the morphological maintenance of dendritic spines, one that is independent of its ability to transport ions. The intimate temporal coincidence between the developmental onset of KCC2 expression and the most intense phase of synaptogenesis during the brain growth spurt points to a possible role for this protein in synapse formation. Importantly, whether KCC2 plays a role in spinogenesis i.e. in induction of spines during the brain growth spurt has not been investigated so far. The results of the present work demonstrate that expression of KCC2 is not only a necessary but also a sufficient condition for the induction of functional glutamatergic spines during the brain growth spurt. The results of this work support the idea of KCC2 as an important synchronizing factor in the functional development of glutamatergic and GABAergic signaling.
  • Bargum, Katja (Helsingin yliopisto, 2007)
    Social groups are common across animal species. The reasons for grouping are straightforward when all individuals gain directly from cooperating. However, the situation becomes more complex when helping entails costs to the personal reproduction of individuals. Kin selection theory has offered a fruitful framework to explain such cooperation by stating that individuals may spread their genes not only through their own reproduction, but also by helping related individuals reproduce. However, kin selection theory also implicitly predicts conflicts when groups consist of non-clonal individuals, i.e. relatedness is less than one. Then, individual interests are not perfectly aligned, and each individual is predicted to favour the propagation of their own genome over others. Social insects provide a solid study system to study the interplay between cooperation and conflict. Breeding systems in social insects range from solitary breeding to eusocial colonies displaying complete division of reproduction between the fertile queen and the sterile worker caste. Within colonies, additional variation is provided by the presence of several reproductive individuals. In many species, the queen mates multiply, which causes the colony to consist of half-sib instead of full-sib offspring. Furthermore, in many species colonies contain multiple breeding queens, which further dilutes relatedness between colony members. Evolutionary biology is thus faced with the challenge to answer why such variation in social structure exists, and what the consequences are on the individual and population level. The main part of this thesis takes on this challenge by investing the dynamics of socially polymorphic ant colonies. The first four chapters investigate the causes and consequences of different social structures, using a combination of field studies, genetic analyses and laboratory experiments. The thesis ends with a theoretical chapter focusing on different social interactions (altruism and spite), and the evolution of harming traits. The main results of the thesis show that social polymorphism has the potential to affect the behaviour and traits of both individuals and colonies. For example, we found that genetic polymorphism may increase the phenotypic variation between individuals in colonies, and that socially polymorphic colonies may show different life history patterns. We also show that colony cohesion may be enhanced even in multiple-queen colonies through patterns of unequal reproduction between queens. However, the thesis also demonstrates that spatial and temporal variation between both populations and environments may affect individual and colony traits, to the degree that results obtained in one place or at one time may not be applicable in other situations. This opens up potential further areas of research to explain these differences.
  • Helanterä, Heikki (Helsingin yliopisto, 2004)
  • Räty, Riitta (Helsingin yliopisto, 2013)
    Respiratory infections are the most common illnesses worldwide, and a large fraction of those are of viral origin. These infections are associated with significant morbidity and mortality and have substantial socioeconomic impact. Rapid identification of a causing agent provides guidance for treatment decisions and may help prevent the further spread of the virus. Noteworthy that virus infections can be complicated by subsequent bacterial infections. Over a period of 16 years, the occurrence of seven common respiratory viruses has been studied in certain segments of the Finnish population. A laboratory test (TR-FIA) for the rapid detection of viral proteins in respiratory secretions has been used in these studies. From many of the patients also acute- and convalescent-phase serum specimens have been collected for the detection of antibodies to these seven viruses and also to Mycoplasma pneumoniae, a bacterial pathogen. During the 16-year period of observation, influenza viruses have caused epidemics every year. Some of these influenza findings have been further investigated in the National Influenza Center and relevant results have been reported to the WHO and to other international organizations. Also the respiratory syncytial virus (RSV) has been found to circulate every year. This virus is an important respiratory pathogen of small children, but also of the elderly. Mycoplasma pneumoniae caused epidemics at intervals of several years. Typically, Mycoplasma pneumoniae affects families with school-age children, and often these infections do not require treatment. Members of the Finnish Defense Forces have been overrepresented in this surveillance, and a significant number of mycoplasma findings have been from young service men hospitalized with pneumonia. Although serological methods rarely help establish a diagnosis during the early phase of an infection, this study has shown that serology performed on paired sera can identify a substantial number of infections that would have gone undetected by rapid diagnostic methods. Besides finding the causative agent of a patient s illness, surveillance of respiratory infections provides information about pathogens that cause epidemics in the population. Through such surveillance it might be possible to identify previously unknown viruses, or common viruses with novel characteristics. As an example, one could mention zoonotic influenza viruses or influenza viruses that have acquired resistance to antiviral drugs.
  • Heiskanen, Jouni (Helsingin yliopisto, 2015)
    Greenhouse gas (GHG) emissions from lakes result from processes between the watershed, lake characteristics and the atmosphere. The organic matter loading from the watershed both provides carbon for the lake biota and in major part defines water clarity, which, in addition to wind and heat flux, is essential in thermocline formation. Thermal stratification suppresses the wind-induced momentum input to the surface water preventing effective mixing of gases throughout the water column. The lake biota process the organic matter, producing carbon dioxide (CO2) in oxic surface water and also methane (CH4) if the near-bottom water becomes anoxic, and thus influence the chemical properties of the water column. Finally, air-water GHG exchange occurs over a thin layer at the water surface. Lakes are typically supersaturated with CO2 and CH4 in relation to average atmospheric mixing ratios causing fluxes of these gases to the atmosphere. Even though lakes cover only 2 % of the world s land surface, it has been estimated that lakes release about 10 % of the carbon fixed annually by the terrestrial ecosystems back to the atmosphere. A critical parameter in the gas exchange estimates is the gas transfer velocity (k), which is governed by turbulence. The implementation of direct flux measurement using the eddy covariance (EC) technique allows the detailed measurements needed to estimate k. However, on lakes, the EC method is a novel subject and as of yet, there has been no published estimates of the error related to these measurements nor fully established set of accepted procedures. The aim of this thesis was to assess the current global CO2 evasion estimates from lakes to the atmosphere by comparing parameterizations for k and the significance of wind and heat flux to the gas transfer in small lakes. To improve future predictions of gas evasion from lakes, we focused on the changes in water clarity and how they affect water column physics and processes in the air-water interface. We used the EC method for the high precision data needed, and therefore also aimed to improve the EC methodology on lakes. The air-water gas transfer was related to both wind and heat loss during times of seasonal stratification, but only to wind during autumn overturn, and the mean value for k of CO2 was 6.0 cm h-1 in Lake Kuivajärvi. When wind-induced thermocline tilting and resulting spatial variability in surface water CO2 concentrations was accounted for, k derived from the measurements dropped to 5.2 cm h-1. This was still over twice the estimate (2.2 cm h-1) calculated with a widely used model for k in lakes suggesting that the global estimates of gas evasion from lakes might be underestimations. Unsolved question is that how important factor the thermocline tilting is in other lakes in defining spatial variability? Our results showed that k for CH4 was higher than for CO2, a result which has been reported in some other studies, but as of yet, no solid explanation has been found. Water clarity was a significant parameter defining the thermal stratification of the lake: a change from clear to dark water would lead to shorter stratification period and lower water column temperatures in small lakes and therefore have significant impact on the lake-atmosphere exchange processes. Important question is how changes is climate will affect lake water clarity e.g. via precipitation and runoff related DOC loading. We concluded that the EC method produces reliable results even in a small lake after rigorous data processing. After these procedures, about half of the CO2 and turbulent heat flux data were of good quality with relative random errors of 10 % and 26 % for heat and CO2 fluxes, respectively. When measuring GHG fluxes accurately from lakes, methods that integrate over time and space are a necessity. These will provide more detailed knowledge on the complex processes that contribute to the gas transfer, from large scale physical phenomena such as thermocline tilting to small scale such as near-surface turbulence. Since most of the world s lakes are small and in northern latitudes, our studies have wide implications even to the global level. Better understanding of the lake biogeochemistry will allow us to make more accurate estimates of GHG evasion from lakes in different regions as well as predictions of how the climate change will affect the lake-atmosphere GHG fluxes.
  • Sillanpää, Maarit (Helsingin yliopisto, 2003)
  • Nurmi, Susanna (Helsingin yliopisto, 2008)
    Integrins are heterodimeric transmembrane adhesion receptors composed of alpha- and beta-subunits and they are vital for the function of multicellular organisms. Integrin-mediated adhesion is a complex process involving both affinity regulation and coupling to the actin cytoskeleton. Integrins also function as bidirectional signaling devices, regulating cell adhesion and migration after inside-out signaling, but also signal into the cell to regulate growth, differentiation and apoptosis after ligand binding. The LFA-1 integrin is exclusively expressed in leukocytes and is of fundamental importance for the function of the immune system. The LFA-1 integrins have short intracellular tails, which are devoid of catalytic activity. These cytoplasmic domains are important for integrin regulation and both the alpha and beta chain become phosphorylated. The alpha chain is constitutively phosphorylated, but the beta chain becomes phosphorylated on serine and functionally important threonine residues only after cell activation. The cytoplasmic tails of LFA-1 bind to many cytoskeletal and signaling proteins regulating numerous cell functions. However, the molecular mechanisms behind these interactions have been poorly understood. Phosphorylation of the cytoplasmic tails of the LFA-1 integrin could provide a mechanism to regulate integrin-mediated cytoskeletal interactions and take part in T cell signaling. In this study, the effects of phosphorylation of LFA-1 integrin cytoplasmic tails on different cellular functions were examined. Site-specific phosphorylation of both the alpha- and beta-chains of the LFA-1 was shown to have a role in the regulation of the LFA-1 integrin.Alpha-chain Ser1140 is needed for integrin conformational changes after chemokine- or integrin ligand-induced activation or after activation induced by active Rap1, whereas beta-chain binds to 14-3-3 proteins through the phosphorylated Thr758 and mediates cytoskeletal reorganization. Thr758 phosphorylation also acts as a molecular switch to inhibit filamin binding and allows 14-3-3 protein binding to integrin cytoplasmic domain, and it was also shown to lead to T cell adhesion, Rac-1/Cdc42 activation and expression of the T cell activation marker CD69, indicating a signaling function for Thr758 phosphorylation in T cells. Thus, phosphorylation of the cytoplasmic tails of LFA-1 plays an important role in different functions of the LFA-1 integrin in T cells. It is of vital importance to study the mechanisms and components of integrin regulation since leukocyte adhesion is involved in many functions of the immune system and defects in the regulation of LFA-1 contributes to auto-immune diseases and fundamental defects in the immune system.
  • Happonen, Lotta (Helsingin yliopisto, 2012)
    Viruses are the most abundant replicating entities on Earth, and they infect cells from all three domains of life - where there are cells, there are viruses. Extremophilic organisms and viruses thrive in hostile environments including hot, acidic springs, oceanic hydrothermal vents, and salt lakes. Due to their adaptation to extreme environments, these organisms and their viruses have been exploited for enzymes useful for industrial and biotechnological applications. Such enzymes include starch processing, cellulose degrading, proteolytic and DNA-processing enzymes. The latter ones are used in molecular biology applications such as polymerase chain reactions and DNA-sequencing. The aim of this study was to characterize novel, extremophilic viruses living in hot springs. I solved the three dimensional structure of two such viruses using electron cryo-microscopy and three dimensional image reconstruction, and explored the presence of extremophilic enzymes based on their genome sequence. One of the viruses characterized in this study is P23-77 that infects the thermophilic bacterium Thermus thermophilus living in alkaline hot springs. P23-77 has been proposed to belong to the Tectiviridae family of viruses characterized by an internal lipid bilayer surrounded by an icosahedral protein capsid. The structure of the icosahedral P23-77 was initially solved to 1.4 nm resolution, and subsequently to 1.0 nm resolution. The reconstruction, together with thin-layer chromatography, confirmed the presence of an internal lipid bilayer composed of neutral lipids. Analysis of the P23-77 protein profile revealed it to have 10 structural proteins, two of which were major ones based on their abundance in SDS-PAGE gels. These proteins were suggested to form the capsomers with hexameric bases of the P23-77 T = 28d capsid lattice. Surprisingly, P23-77 closely resembles the haloarchaeal virus SH1, both of which are suggested to have single β-barrel major capsid proteins, and together forming a novel viral lineage. The other virus characterized in this study is the Sulfolobus turreted icosahedral virus 2 (STIV2) infecting the crenarchaeon Sulfolobus islandicus that lives in acidic hot springs. The genome of STIV2 was sequenced, and some of its structural proteins were determined by mass-peptide fingerprinting. The structure of STIV2 was solved to 2.0 nm resolution. The genome sequence and the structure of STIV2 revealed it to resemble most closely STIV, infecting S. solfataricus. Like P23-77, both STIV and STIV2 have an outer protein capsid surrounding the internal lipid bilayer and the double-stranded (ds) DNA genome. The most striking difference between STIV and STIV2 resides in the host-cell recognition and attachments structures, which in STIV2 lacks the petal-like appendages present in STIV. Based on difference imaging, homology modeling and comparison to STIV, a model for the organization of the STIV2 virion was proposed. Furthermore, based on sequence data and homology modeling I identified the postulated genome packaging NTPase B204 of STIV2. I expressed and purified B204, and studied the nucleotide hydrolysis catalyzed by it. I furthermore solved four structures of B204 more precisely, in complex with a sulphate ion, adenosine monosphosphate, the product adenosine diphosphate, and the substrate analogue adenylylmethylenediphosphonate. B204 is the first genome packaging NTPase of a membrane-containing virus for which the structure has been solved. Based on the structure of B204, comparison to other known DNA-translocating enzymes, and other genome packaging NTPases of dsDNA and dsRNA viruses, I propose a model for the genome packaging of STIV2.
  • Koutaniemi, Sanna (Helsingin yliopisto, 2007)
    Lignin is a hydrophobic polymer that is synthesised in the secondary cell walls of all vascular plants. It enables water conduction through the stem, supports the upright growth habit and protects against invading pathogens. In addition, lignin hinders the utilisation of the cellulosic cell walls of plants in pulp and paper industry and as forage. Lignin precursors are synthesised in the cytoplasm through the phenylpropanoid pathway, transported into the cell wall and oxidised by peroxidases or laccases to phenoxy radicals that couple to form the lignin polymer. This study was conducted to characterise the lignin biosynthetic pathway in Norway spruce (Picea abies (L.) Karst.). We focused on the less well-known polymerisation stage, to identify the enzymes and the regulatory mechanisms that are involved. Available data for lignin biosynthesis in gymnosperms is scarce and, for example, the latest improvements in precursor biosynthesis have only been verified in herbaceous plants. Therefore, we also wanted to study in detail the roles of individual gene family members during developmental and stress-induced lignification, using EST sequencing and real-time RT-PCR. We used, as a model, a Norway spruce tissue culture line that produces extracellular lignin into the culture medium, and showed that lignin polymerisation in the tissue culture depends on peroxidase activity. We identified in the culture medium a significant NADH oxidase activity that could generate H2O2 for peroxidases. Two basic culture medium peroxidases were shown to have high affinity to coniferyl alcohol. Conservation of the putative substrate-binding amino acids was observed when the spruce peroxidase sequences were compared with other peroxidases with high affinity to coniferyl alcohol. We also used different peroxidase fractions to produce synthetic in vitro lignins from coniferyl alcohol; however, the linkage pattern of the suspension culture lignin could not be reproduced in vitro with the purified peroxidases, nor with the full complement of culture medium proteins. This emphasised the importance of the precursor radical concentration in the reaction zone, which is controlled by the cells through the secretion of both the lignin precursors and the oxidative enzymes to the apoplast. In addition, we identified basic peroxidases that were reversibly bound to the lignin precipitate. They could be involved, for example, in the oxidation of polymeric lignin, which is required for polymer growth. The dibenzodioxocin substructure was used as a marker for polymer oxidation in the in vitro polymerisation studies, as it is a typical substructure in wood lignin and in the suspension culture lignin. Using immunolocalisation, we found the structure mainly in the S2+S3 layers of the secondary cell walls of Norway spruce tracheids. The structure was primarily formed during the late phases of lignification. Contrary to the earlier assumptions, it appears to be a terminal structure in the lignin macromolecule. Most lignin biosynthetic enzymes are encoded for by several genes, all of which may not participate in lignin biosynthesis. In order to identify the gene family members that are responsible for developmental lignification, ESTs were sequenced from the lignin-forming tissue culture and developing xylem of spruce. Expression of the identified lignin biosynthetic genes was studied using real-time RT-PCR. Candidate genes for developmental lignification were identified by a coordinated, high expression of certain genes within the gene families in all lignin-forming tissues. However, such coordinated expression was not found for peroxidase genes. We also studied stress-induced lignification either during compression wood formation by bending the stems or after Heterobasidion annosum infection. Based on gene expression profiles, stress-induced monolignol biosynthesis appeared similar to the developmental process, and only single PAL and C3H genes were specifically up-regulated by stress. On the contrary, the up-regulated peroxidase genes differed between developmental and stress-induced lignification, indicating specific responses.
  • Burgas, Daniel (Helsingin yliopisto, 2014)
    Under the current biodiversity crisis, there is the need to improve the conservation action. More areas need to be protected to curb biodiversity loss. Also, the methods for selection of both protected areas and management practices have to be well-informed in order to maximise the benefits from the limited resources allocated for conservation. However, because of limited information, decision making procedures are forced to use environmental variables and different species as surrogates of general biodiversity. Moreover, there is a bias towards charismatic and better known species like top predators. It is therefore important to forecast the consequences that favouring certain species might have on other organisms and to evaluate the effectiveness of preserving a subset of species. In this thesis I use empirical data to investigate how avian predators (the Northern Goshawk Accipiter gentilis and the Ural Owl Strix uralensis) associate to biodiversity. The focus is two-fold; First, I investigate the role of species interactions in dictating biodiversity patterns. I show that interactions within the predator assembly can have stronger effect on prey distribution (the Siberian Flying Squirrel Pteromys volans) than landscape attributes. This finding points out that individuals may be able to respond to changes in the structure of the predator assemblage. Additionally, I examine the impact of the Goshawk altering the structure of the forest bird community. I found that not only the raptor conditioned species distribution across space, but that this effect persisted over the years after the Goshawk abandoned the breeding site. Second, I evaluate the potential use of raptors as surrogates to indicate areas of relevant conservation value. On the one hand, I address how the two focal raptor species associate to different biodiversity metrics for birds and polypores (i.e. wood decaying fungi) over a landscape gradient. I found that, while both predator species indicated high biodiversity levels as compared to reference sites, the dominant Goshawk was superior to the Ural Owl. Interestingly, the surrogacy properties remained the same even if the environmental setting changed. On the other hand, I assess the value of preserving raptor nest sites in the context of the existing network of protected areas in the study region. I show that using goshawk nest sites is the most cost-efficient approach if considering only single species. However, combining both predator species further enhanced the conservation output. Concluding, this thesis highlights the idea that avian top predators have a key role on species distributions and shaping community heterogeneity in space and time. Given this disproportionate influence in ecosystems, apex predators might also merit extra conservation commitment. Furthermore, information on the spatial distribution of raptors can indicate, over large areas, locations of disproportional biodiversity value. I show that setting aside raptor nest sites is more cost-effective than other conservation approaches. Additionally, one has to consider that national-wide monitoring schemes already provide raptor nest locations at no extra cost. In the light of these results it is advisable to consider the use of raptors to complement existing and future reserve selection methods.
  • Laine, Anna-Liisa (Helsingin yliopisto, 2005)
  • Majaneva, Markus (Helsingin yliopisto, 2013)
    Protists are unicellular eukaryotes. Some protistan species may be impossible to distinguish under the light or even electron microscope, and a complete balanced study of protistan taxonomy requires molecular analysis and light and electron microscopy. One of the main applications of taxonomic work is the assessment of diversity of organisms in an ecosystem. However, uncertainty in taxonomic precision undermines the diversity measures. DNA sequence data provide assistance since they are easily transformed to numbers that can be compared systematically and in a similar way throughout the eukaryotic domain, using sequence similarity to define operational taxonomic units (OTUs). DNA-based assessment of diversity is called environmental sequencing. The most commonly used gene in the environmental gene sequencing for eukaryotes and also in the protistan taxonomic studies has been the small subunit (18S) ribosomal RNA gene of the ribosomal operon. Also, internal transcribed spacers (ITS) are used. The studies in this thesis were conducted with Baltic Sea protists. The Baltic Sea is a subarctic brackish-water basin that partially freezes over every winter. If the salinity of parent water is higher than 0.6, the forming ice has a semi-solid structure with solid ice crystals and saline water (brine) channels. The brine channels offer habitats for small-sized organisms. Due to the low salinity of the Baltic Sea, the brine channels are small, and therefore, the Baltic Sea ice eukaryotic community is dominated by protists. Studies on Baltic sea-ice biology have been accumulating since the 1980 s, but there are still gaps on knowledge; for example, what protistan species and how many there are associated with sea ice. In this thesis, morphological, molecular and ecological information was combined to delineate species of a Baltic Sea cryptomonad, haptophyte and dinoflagellate. Protistan community composition in Baltic Sea ice was assessed with environmental sequencing, and diversity estimates were compared in different types of ice. The taxonomic and environmental-sequencing studies were linked using the gathered taxonomic information to evaluate the accuracy of the molecular diversity-measurement method. A new cryptomonad species, Rhinomonas nottbecki, was described based on morphological characters distinguished by light- and electron-microscopy together with molecular evidence from 18S rRNA gene and ITS region. The same approach was applied to the identification of the alternate stage Prymnesium polylepis (Haptophyta), which bloomed in the whole Baltic Proper during autumn spring 2007 2008. Also, a novel cold-water and sea-ice associated dinoflagellate subspecies, Heterocapsa arctica subsp. frigida, was described. Environmental 18S-rRNA-gene sequencing revealed that the richest eukaryotic lineages inhabiting the Baltic Sea ice were ciliates, cercozoa, dinoflagellates and diatoms. The different developmental stages and types of ice had different community composition. Protistan richness was higher in ice than water even though water included more divergent lineages. The Baltic fast ice had higher richness than pack and drift ice. The results of this thesis showed that there remains novelty to be described in the Baltic Sea, and what we know about the protistan community in Baltic Sea ice now is very incomplete. Although the environmental sequencing produced data that met the requirements of calculation of comparable diversity indices (all taxa defined at the same level), revealed cryptic taxa, and gave higher protistan richness than basic light microscopy of fixed samples, the lack of taxonomic detail was not restricted to the light-microscopic surveys but was also a result of the environmental-sequencing approach. This was shown when the environmental-sequencing approach was applied on the 18S-rRNA-gene data of the cryptomonad family Pyrenomonadaceae and the haptophyte genus Prymnesium. Only one Pyrenomonadaceae and two Prymnesium OTUs were found although both data sets included 15 distinct taxa. Errors in environmental sequencing and alignment make the use of high similarity levels in the OTU definition questionable, and the variability in the 18S rDNA is not equal within different eukaryotic lineages. Consequently, use of lower similarity level (97 %) is justifiable in the environmental-sequencing, but the approach used gave conservative estimates of the protistan richness in the Baltic Sea ice. The overall conclusion is that we need to apply all available techniques when assessing the diversity of protists, as each technique provides a biased perspective on nature. A labor intensive taxonomic approach that includes the study of live cells by light microscopy, detailed morphological description based on electron microscopy and phylogenetic analysis of suitable genetic markers gives us the best chance of finding out how many different species of protists live within the Baltic Sea ice or any other environment, and what they do.
  • Kontiainen, Pekka (Helsingin yliopisto, 2010)
    Individuals face variable environmental conditions during their life. This may be due to migration, dispersion, environmental changes or, for example, annual variation in weather conditions. Genetic adaptation to a novel environment happens through natural selection. Phenotypic plasticity allows, however, a quick individual response to a new environment. Phenotypic plasticity may also be beneficial for individual if the environment is highly variable. For example, eggs are costly to produce. If the food conditions vary significantly between breeding seasons it is useful to be able to adjust the clutch and egg size according to the food abundance. In this thesis I use Ural owl vole system to study phenotypic plasticity and natural selection using a number of reproduction related traits. The Ural owl (Strix uralensis) is a long-lived and sedentary species. The reproduction and survival of the Ural owl, in fact their whole life, is tied to the dramatically fluctuating vole densities. Ural owls do not cause vole cycles but they have to adjust their behaviour to the rather predictable population fluctuations of these small mammals. Earlier work with this system has shown that Ural owl laying date and clutch size are plastic in relation to vole abundance. Further, individual laying date clutch size reaction norms have been shown to vary in the amount of plasticity. My work extends the knowledge of natural selection and phenotypic plasticity in traits related to reproduction. I show that egg size, timing of the onset of incubation and nest defense aggressiveness are plastic traits with fitness consequences for the Ural owl. Although egg size is in general thought to be a fixed characteristic of an individual, this highly heritable trait in the Ural owl is also remarkably plastic in relation to the changes in vole numbers, Ural owls are laying the largest eggs when their prey is most abundant. Timing of the onset of incubation is an individual-specific property and plastic in relation to clutch size. Timing of incubation is an important underlying cause for asynchronous hatching in birds. Asynchronous hatching is beneficial to offspring survival in Ural owl. Hence, timing of the onset of incubation may also be under natural selection. Ural owl females also adjust their nest defense aggressiveness according to the vole dynamics, being most aggressive in years when they produce the largest broods. Individual females show different levels of nest defense aggressiveness. Aggressiveness is positively correlated with the phenotypic plasticity of aggressiveness. As elevated nest defense aggressiveness is selected for, it may promote the plasticity of aggressive nest defense behaviour. All the studied traits are repeatable or heritable on individual level, and their expression is either directly or indirectly sensitive to changes in vole numbers. My work considers a number of important fitness-related traits showing phenotypic plasticity in all of them. Further, in two chapters I show that there is individual variation in the amount of plasticity exhibited. These findings on plasticity in reproduction related traits suggest that variable environments indeed promote plasticity.
  • Sokka, Laura (Helsingin yliopisto, 2011)
    Human activities extract and displace different substances and materials from the earth s crust, thus causing various environmental problems, such as climate change, acidification and eutrophication. As problems have become more complicated, more holistic measures that consider the origins and sources of pollutants have been called for. Industrial ecology is a field of science that forms a comprehensive framework for studying the interactions between the modern technological society and the environment. Industrial ecology considers humans and their technologies to be part of the natural environment, not separate from it. Industrial operations form natural systems that must also function as such within the constraints set by the biosphere. Industrial symbiosis (IS) is a central concept of industrial ecology. Industrial symbiosis studies look at the physical flows of materials and energy in local industrial systems. In an ideal IS, waste material and energy are exchanged by the actors of the system, thereby reducing the consumption of virgin material and energy inputs and the generation of waste and emissions. Companies are seen as part of the chains of suppliers and consumers that resemble those of natural ecosystems. The aim of this study was to analyse the environmental performance of an industrial symbiosis based on pulp and paper production, taking into account life cycle impacts as well. Life Cycle Assessment (LCA) is a tool for quantitatively and systematically evaluating the environmental aspects of a product, technology or service throughout its whole life cycle. Moreover, the Natural Step Sustainability Principles formed a conceptual framework for assessing the environmental performance of the case study symbiosis (Paper I). The environmental performance of the case study symbiosis was compared to four counterfactual reference scenarios in which the actors of the symbiosis operated on their own. The research methods used were process-based life cycle assessment (LCA) (Papers II and III) and hybrid LCA, which combines both process and input-output LCA (Paper IV). The results showed that the environmental impacts caused by the extraction and processing of the materials and the energy used by the symbiosis were considerable. If only the direct emissions and resource use of the symbiosis had been considered, less than half of the total environmental impacts of the system would have been taken into account. When the results were compared with the counterfactual reference scenarios, the net environmental impacts of the symbiosis were smaller than those of the reference scenarios. The reduction in environmental impacts was mainly due to changes in the way energy was produced. However, the results are sensitive to the way the reference scenarios are defined. LCA is a useful tool for assessing the overall environmental performance of industrial symbioses. It is recommended that in addition to the direct effects, the upstream impacts should be taken into account as well when assessing the environmental performance of industrial symbioses. Industrial symbiosis should be seen as part of the process of improving the environmental performance of a system. In some cases, it may be more efficient, from an environmental point of view, to focus on supply chain management instead.  
  • Paloniemi, Riikka (Helsingin yliopisto, 2008)
    Nature conservation in everyday life: Private landowners perceptions of and experiences on temporary nature conservation and its renewing process This study explores the legitimacy of official nature conservation of private lands. It describes how temporary nature conservation became a part of Finnish nature conservation policy and how forest owners perceived this tool and its usage. In addition, the study analyses forest owners' attitudes on official nature conservation. The study combines individual and official perspectives, and presents a nature conservation politics of everyday life. The theoretical background of the study is learning processes of environmentally responsible participation, and especially empowerment. Main methods in gathering the material for the study have been interviews and a survey. In the 1990 s, Finnish landowners opposed the implementation of nature conservation, especially conservation of shores and the establishment of the European-wide network of conservation areas (Natura 2000). After negative experiences on these conservation efforts, some private landowners were disempowered and some even rejected conservation completely. The Nature Conservation Act of 1996 launched an option to conserve nature officially for fixed time-periods. Use of such a policy tool did not immediately become a widely-used and appreciated conservation practice. During the following decade, however, it attracted remarkable attention within Finnish nature conservation policy discussions. The perspective of landowners began to be emphasised when local and regional organisations for nature conservation and forestry together defined the ideas of natural values trading. Later, the national governance process of the committee defining the Forest Biodiversity Programme for Southern Finland (METSO) institutionalised these ideas in one of its pilot projects. Landowners participated in the project of natural values trading by offering their forests for conservation, which reflects the increased acceptance and legitimacy of nature conservation on private forests. The central elements producing the legitimacy of natural values trading have been voluntariness, temporariness, and dialogue between nature conservation and forestry. Natural values trading analysed in the study is an example of new environmental policy instruments and its creation process represents governance in the implementation of nature conservation. It has increased the legitimacy of nature conservation policy of private forests in Finland. The results reveal the importance of participation and learning processes in the implementation of nature conservation policy, and the need to also pay attention to these processes in the future.
  • Kärpänen, Terhi (Helsingin yliopisto, 2006)
    The circulatory system consists of two vessel types, which act in concert but significantly differ from each other in several structural and functional aspects as well as in mechanisms governing their development. The blood vasculature transports oxygen, nutrients and cells to tissues whereas the lymphatic vessels collect extravasated fluid, macromolecules and cells of the immune system and return them back to the blood circulation. Understanding the molecular mechanisms behind the developmental and functional regulation of the lymphatic system long lagged behind that of the blood vasculature. Identification of several markers specific for the lymphatic endothelium, and the discovery of key factors controlling the development and function of the lymphatic vessels have greatly facilitated research in lymphatic biology over the past few years. Recognition of the crucial importance of lymphatic vessels in certain pathological conditions, most importantly in tumor metastasis, lymphedema and inflammation, has increased interest in this vessel type, for so long overshadowed by its blood vascular cousin. VEGF-C (Vascular Endothelial Growth Factor C) and its receptor VEGFR-3 are essential for the development and maintenance of embryonic lymphatic vasculature. Furthermore, VEGF-C has been shown to be upregulated in many tumors and its expression found to positively correlate with lymphatic metastasis. Mutations in the transcription factor FOXC2 result in lymphedema-distichiasis (LD), which suggests a role for FOXC2 in the regulation of lymphatic development or function. This study was undertaken to obtain more information about the role of the VEGF-C/VEGFR-3 pathway and FOXC2 in regulating lymphatic development, growth, function and survival in physiological as well as in pathological conditions. We found that the silk-like carboxyterminal propeptide is not necessary for the lymphangiogenic activity of VEGF-C, but enhances it, and that the aminoterminal propeptide mediates binding of VEGF-C to the neuropilin-2 coreceptor, which we suggest to be involved in VEGF-C signalling via VEGFR-3. Furthermore, we found that overexpression of VEGF-C increases tumor lymphangiogenesis and intralymphatic tumor growth, both of which could be inhibited by a soluble form of VEGFR-3. These results suggest that blocking VEGFR-3 signalling could be used for prevention of lymphatic tumor metastasis. This might prove to be a safe treatment method for human cancer patients, since inhibition of VEGFR-3 activity had no effect on the normal lymphatic vasculature in adult mice, though it did lead to regression of lymphatic vessels in the postnatal period. Interestingly, in contrast to VEGF-C, which induces lymphangiogenesis already during embryonic development, we found that the related VEGF-D promotes lymphatic vessel growth only after birth. These results suggest, that the lymphatic vasculature undergoes postnatal maturation, which renders it independent of ligand induced VEGFR-3 signalling for survival but responsive to VEGF-D for growth. Finally, we show that FOXC2 is necessary for the later stages of lymphatic development by regulating the morphogenesis of lymphatic valves, as well as interactions of the lymphatic endothelium with vascular mural cells, in which it cooperates with VEGFR-3. Furthermore, our study indicates that the absence of lymphatic valves, abnormal association of lymphatic capillaries with mural cells and an increased amount of basement membrane underlie the pathogenesis of LD. These findings have given new insight into the mechanisms of normal lymphatic development, as well as into the pathogenesis of diseases involving the lymphatic vasculature. They also reveal new therapeutic targets for the prevention and treatment of tumor metastasis and lymphatic vascular failure in certain forms of lymphedema. Several interesting questions were posed that still need to be addressed. Most importantly, the mechanism of VEGF-C promoted tumor metastasis and the molecular nature of the postnatal lymphatic vessel maturation remain to be elucidated.
  • Pykälä, Juha (Helsingin yliopisto, 2007)
    Semi-natural grasslands are the most important agricultural areas for biodiversity. The present study investigates the effects of traditional livestock grazing and mowing on plant species richness, the main emphasis being on cattle grazing in mesic semi-natural grasslands. The two reviews provide a thorough assessment of the multifaceted impacts and importance of grazing and mowing management to plant species richness. It is emphasized that livestock grazing and mowing have partially compensated the suppression of major natural disturbances by humans and mitigated the negative effects of eutrophication. This hypothesis has important consequences for nature conservation: A large proportion of European species originally adapted to natural disturbances may be at present dependent on livestock grazing and / or mowing. Furthermore, grazing and mowing are key management methods to mitigate effects of nutrient-enrichment. The species composition and richness in old (continuously grazed), new (grazing restarting 3-8 years ago) and abandoned (over 10 years) pastures differed consistently across a range of spatial scales, and was intermediate in new pastures compared to old and abandoned pastures. In mesic grasslands most plant species were shown to benefit from cattle grazing. Indicator species of biologically valuable grasslands and rare species were more abundant in grazed than in abandoned grasslands. Steep S-SW-facing slopes are the most suitable sites for many grassland plants and should be prioritized in grassland restoration. The proportion of species trait groups benefiting from grazing was higher in mesic semi-natural grasslands than in dry and wet grasslands. Consequently, species trait responses to grazing and the effectiveness of the natural factors limiting plant growth may be intimately linked High plant species richness of traditionally mowed and grazed areas is explained by numerous factors which operate on different spatial scales. Particularly important for maintaining large scale plant species richness are evolutionary and mitigation factors. Grazing and mowing cause a shift towards the conditions that have occurred during the evolutionary history of European plant species by modifying key ecological factors (nutrients, pH and light). The results of this Dissertation suggest that restoration of semi-natural grasslands by private farmers is potentially a useful method to manage biodiversity in the agricultural landscape. However, the quality of management is commonly improper, particularly due to financial constraints. For enhanced success of restoration, management regulations in the agri-environment scheme need to be defined more explicitly and the scheme should be revised to encourage management of biodiversity.