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HIV-1 is a lentivirus causing a serious immunodeficiency disease called AIDS to almost 38 million people in the world. Like all retroviruses, HIV has an ability to insert a copy of its genome into the host cell DNA thus having a lifelong effect on the host cell. Two identical copies of single-stranded RNA are first reverse transcribed, and then a protein called integrase inserts the DNA into the genome of T-cells of the human immune system. Studies have shown that in some retroviruses the integrase protein has a unique integration pattern favoring certain areas of the host cell genome. For example, active transcription units or promoter units are found to attract integration activity. On the other hand, with nuclear localization signals in its structure and an ability to attach to the human genome, it raises a question if the integrase could have additional functions. Regulating certain gene expression levels could support viral replication and the survival of the virus. The aim of the study was to determine whether integration sites differ from integrase genomic contact sites, which could be an indication of integrase’s additional role. Study was started by method optimization. Two chromatin fragmentation methods, sonication and endo-exonuclease treatment, were tested in order to achieve optimal sized DNA fragments for ChIP-sequencing. Two cell lines were infected with HIV-1. Genomic DNA was collected for integration site sequencing. Integrase genomic contact sites were studied with ChIP-sequencing. Next generation sequencing for both, integration sites and integrase genomic contact sites, was carried out by an outsource biotechnology company. Sequences were processed with bioinformatics platforms, such as Galaxy and Homer. Study of the effects of transfected integrase was also initiated by transfection method optimization. Treatment with endo-exonuclease resulted in correctly sized DNA fragments when DNA was first purified. In a cell lysate, a correct ratio of MNase and cells was not found, although several cell lysis methods and buffers were tried. This is why actual integrase genomic contact site samples were fragmented by sonication. In MRC-5 cell line, integration site sequencing resulted in over 2 M reads with 289 aligned, unique integration sites. Integrase genomic contact site sequencing resulted in 1.8E+6 Bowtie mapped reads. A brief analysis of the sequences including sequence visualization, comparison of localization in the genome and GO terms, showed that integrase genomic contact sites have certain patterns that differ from integration sites. Therefore, it was confirmed that the study is repeatable, as the results were in line with the study conducted several years earlier. These results also suggest that method optimization had been successful. As bioinformatics methods were used with default parameters only, more attention should be payed when sequences are analyzed more deeply. This will offer a more thorough understanding of the functions of integrase.

Kosteikot ovat biodiversiteetiltään yksi maailman rikkaimmista habitaateista. Kosteikkojen ja niillä elävien vesilintujen tuottamien ekosysteemipalvelujen laajuuteen ja tärkeyteen on havahduttu vasta viime aikoina. Kosteikkoja ja niiden linnustoa uhkaavat maailman laajuisesti maankäyttö sekä makeiden vesistöjen pilaantuminen. Viimeisen vuosisadan aikana maailman kosteikkojen määrä on vähentynyt puoleen ja niiden tila on heikentynyt. Kosteikkolinnuston elinolot ovat heikentyneet ja linnusto on taantunut. Kosteikkoja rakentamalla ja kunnostamalla on voitu parantaa linnuston elinolosuhteita. Etenkin keinotekoisten kosteikkojen merkitys on ollut suuri luontaisten kosteikkojen tilan heikkenemisen myötä. Vain vähän tiedetään miten luonnollisten kosteikkojen hoitotoimet vaikuttavat linnustoon paikallisella tasolla. Niukasti tietoa on myös saatavilla päättäjille hoitotoimien kustannustehokkuudesta. Tässä työssä tutkittiin Etelä-Suomen kosteikkojen hoitotoimien vaikutuksia levähtävään ja pesivään linnustoon. Työn tarkoituksena oli selvittää eri hoitotoimien vaikutuksia ravinnonhankinnaltaan erilaisiin lintukiltoihin ja -ryhmiin. Yhtenä tutkimuksen päätavoitteena oli luoda käsitys hoitotoimien kustannustehokkuudesta sekä mihin resurssit kannattaa käyttää. Tutkimusalue käsitti 21 Etelä-Suomen linnustollisesti arvokasta kosteikkoa. Kosteikkoja kunnostettiin vuosina 2004–2012 kahdessa hoitojaksossa. Hoitojaksoja ennen ja niiden jälkeen kosteikkojen pesivä ja levähtävä linnusto selvitettiin. Suomen ympäristökeskus vastasi hoitotoimista ja linnustolaskennoista Uudenmaan ja Kymenlaakson ELY-keskusten kanssa. Kosteikkoja kunnostettiin pääasiassa avoimuutta lisäämällä laidunnuksen, puustonraivauksen, niiton ja äestyksen sekä ruoppauksen avulla. Muutolla levähtävä linnusto selvitettiin läpi muuttokauden noin viiden päivän välein suoritettujen laskentojen avulla ja pesivä linnusto selvitettiin viiden käyntikerran kartoitusmenetelmällä. Kohteiden linnut eriteltiin hoidettuihin osa-alueisiin sekä niiden ulkopuolelle jääviin hoitamattomiin osa-alueisiin. Kontrollialueina toimivat hoitamattomat kohteet ja osa-alueet. Lasketut linnut yhdistettiin ravinnonhankintansa perusteella kymmeneen kiltaan, jotka olivat puolisukeltajasorsat, sukeltajasorsat, kalansyöjävesilinnut, joutsenet, hanhet, kahlaajat, naurulokki, rantakanat ja kaulushaikara, avomaavarpuslinnut sekä pensaston ja ruovikon varpuslinnut. Kolmea viimeksi mainittua kiltaa tarkasteltiin vain pesimäaikana. Erikseen tarkasteltiin kansallisesti uhanalaisia sekä lintudirektiivin liitteen I lajeja. Hoitotoimien sekä niihin käytettyjen kokonaiskustannusten vaikutuksia kiltakohtaisiin lintumääriin tarkasteltiin lineaarisilla sekamalleilla. Syysmuuttoaineistossa kaikkien lintukiltojen määrät kasvoivat niitto- ja äestysalan myötä; laidunnuspinta-ala lisäsi hanhien ja kahlaajien lukumääriä. Kaikkien kiltojen lukumääriä lisäsi laidunala keväällä, lisäksi niitto- ja äestys - sekä ruoppausala lisäsivät kahlaajien lukumääriä. Kalansyöjien lukumäärät vähenivät ruoppausalan kasvun myötä, mutta yhteys oli tilastollisesti vain suuntaa antava. Pesimäaineistossa ruoppausala x kilta-interaktio osoittautui tilastollisesti merkitseväksi, mutta kiltakohtaisesti tarkasteltuna vaikutukset olivat suuntaa antavia; rantakanat ja kaulushaikara runsastuivat ja kalansyöjävesilinnut vähenivät. Sekä uhanalaisten että lintudirektiivin liitteen I lajien lukumääriä kasvatti laidunnus keväällä ja syksyllä. Hoitotoimiin käytettyjen kokonaiskustannusten suhteen runsastuivat syksyllä puolisukeltajasorsat, hanhet ja kahlaajat. Keväällä kokonaiskustannuksilla oli positiivinen yhteys puolisukeltajasorsien, kahlaajien ja naurulokin lukumääriin. Pesimäaineistossa kokonaiskustannukset kasvattivat naurulokin lukumääriä. Suuntaa-antavasti kokonaiskustannukset lisäsivät rantakanojen ja kaulushaikaran määriä sekä vähensivät kalansyöjävesilintujen määriä. Erityisesti huomioitavaa on, että hoitotoimilla havaittiin vain positiivisia merkitseviä tuloksia lintukiltoihin. Laidunnus oli tärkein yksittäinen hoitotoimi, mikä liittynee sen ympäristöä monipuolistavaan vaikutukseen. Hoitotoimista eniten hyötynyt kilta oli kahlaajat, mutta kaikki killat hyötyivät jostain hoitotoimesta. Hoitotoimien positiiviset vaikutukset liittynevät avoimuuden lisääntymiseen sekä karjan läsnäoloon, jotka voivat parantaa lintujen ruokailuolosuhteita sekä lisätä turvallisuutta. Kokonaiskustannukset paljastivat että naurulokki ja puolisukeltajasorsat hyötyivät hoitotoimista kokonaisuutena, sillä ne eivät poikenneet muista killoista yksittäisiä hoitotoimia tarkasteltaessa. Vähiten hyötyivät sukeltajasorsat sekä kalansyöjävesilinnut. Tutkimus osoittaa, että hoitotoimilla voidaan parantaa taantuneiden kosteikkolintujen elinolosuhteita. Koska hoitokohteet ovat Etelä-Suomen parhaimpia lintuvesiä, on hoitotoimien biologinen merkitys suuri. Vaikutukset eivät ole pelkästään kansalliset, vaan kunnostuksesta hyötyvät Suomenlahden muuttoreittiä pitkin pohjoiseen muuttavat kosteikkolinnut –aina Siperiaan asti. Yksi tärkeimmistä huomioista on, että hoidon loputtua kosteikkojen tila heikkenee. Hoitoa tulisi jatkaa, jottei saatuja hyötyjä menetettäisi. Tulevaisuudessa olisi tärkeää suunnitella mittavat hoitotoimet myös tutkimuksen kannalta, jolloin hoitotoimien vaikutuksia voitaisiin tutkia tarkemmin. Tärkeää olisi löytää keinot sukeltajasorsien ja kalansyöjien elinolojen parantamiseksi.

Antimicrobial resistance is an emerging concern at the global scale, threatening the effectiveness of antibiotics in treating bacterial infections. Among anthropogenically impacted environments, wastewater treatment plants have been indicated as possible reservoirs of antibiotic resistance genes, putative hotspots for their horizontal gene transfer, and a source of their dissemination to the environment. Generally, the abundance of antibiotic resistance genes is reduced during the wastewater treatment process. However, some genes were shown to be enriched in purified effluent water and dried sludge, which are then released to the environment, compared to influent water. Also, the taxonomy of the hosts carrying antibiotic resistance genes could change as a result of horizontal gene transfer events. The aim of this study was to analyse and compare the host range of a series of antibiotic resistance genes in influent water, effluent water and dried sludge collected from the Viikinmäki wastewater treatment plant in Helsinki, Finland, by applying Emulsion, Paired Isolation and Concatenation PCR (epicPCR). EpicPCR is a method that can link a gene of interest to the 16S rRNA gene from the genome of the host bacterium, without any cultivation step. The abundance of the hosts was also evaluated by sequencing the 16S rRNA gene from the whole bacterial community. In several cases, the target antibiotic resistance genes (blaIMP, blaNDM, ermB, ermF, sul1 and strB) were carried in effluent water and dried sludge by taxa that were not hosting them in influent water, suggesting that horizontal gene transfer events might have occurred during the treatment. All the examined genes were detected both in abundant and in rare taxa, including genera that also comprise pathogenic species, such as Arcobacter and Acinetobacter. Some of the detected hosts were not previously known to show resistant phenotypes, namely members of the family Methylophilaceae. These results corroborate the idea that wastewater treatment plants might be hotspots for the horizontal gene transfer of resistance determinants, and potentially disseminate antibiotic resistant pathogens to the environment. However, in order to ensure the accuracy of the results, the limits of epicPCR as a method need to be identified and addressed.

During this study I examined the roles that resource diversity and trophic complexity play in the emergence and maintenance of biodiversity (ecological complexity) via the use of the evolutionary software Avida. I found a positive relationship between resource diversity and ecological complexity as would be expected from previous studies. However, a trade off was observed between ecological complexity and resource diversity, following the principle of diminishing returns. I also showed that trophic complexity exhibited a positive relationship with ecological complexity echoing what was described in previous studies. However, this increase in ecological complexity resulted in an increase in competition which slowed down the accumulation of higher resources which in-turn affected the emergence of higher-level functions. This study further suggests that an increase in ecological complexity results in an increase in ecosystem stability, however more rigorous measures are needed to fully establish this link. Despite, the inherent abstractions that are present in the Avida system, it has provided valuable insights into the relationships between these ecological factors. The findings suggest that there is need for more future studies possibly looking at the resilience and recovery of these ecosystems as this would be critical for understanding and addressing the current biodiversity crisis.

After drastic declines in large carnivores’ populations globally, conservation efforts have been successful, and predators’ populations are in recovery. However, their comeback has led to new interactions with locals, generating different conflicts. Two main approaches have been considered to mitigate these conflicts, those being the land sparing and land sharing models, however, the land sparing model requires great extents of protected areas, areas that in Europe are missing, therefore forcing a call for the land sharing model. In Finland, this approach has generated debates among different stakeholders, the outcomes of this debate shaping the fate for the four species: brown bear (Ursus arctos), grey wolf (Canis lupus), Eurasian lynx (Lynx lynx) and wolverine (Gulo gulo). Attitudes towards those species can be used to explore the drivers of the conflicts, however, only few studies have explored this context, considering the ecological and social dimension separately. In addition, the ecotourism industry has been recently recognized as a new stakeholder in the Finnish large carnivore’s context, but the effects of its activities were assessed only ecologically. Therefore, with this study I aimed to explore the attitudes of locals from a specific region of Finland towards the four large carnivores’ species, and to assess the different drivers of those species, through a combination of field questionnaires, social variables and large carnivores’ population data. I explored potential correlates of the differences in attitudes, adding also the spatial effect of ecotourism over the socio-ecological factors. I predicted attitudes to vary among species, having on one side the brown bear with positive attitudes, in contrast the wolf with negative attitudes, while neutral attitudes towards the lynx and wolverine. I also expected to find more negative attitudes in smaller localities rather than in localities with a greater human population density. Also, I explored whether the ecotourism activities have a positive or negative effect over the locals’ attitude towards carnivores, expecting the ecotourism industry to bring positive attitudes in nearby localities. As result, attitudes towards the four different species varied significantly, the attitudes towards each different species having different drivers, with the human population size being important for wolverines and wolves, while the status for bear and lynx populations. The ecotourism had an effect only on bear attitudes, being positively correlated (closer the ecotourism activities were, more positive the attitudes are). To mitigate the large carnivores-human conflict in Finland, a community approach is not the solution, since the different origins of the attitudes ‘drivers. However, the attitudes among species are positively correlated, consequently, by ameliorate the attitudes towards one species, also the others will benefit. Finally, by inducing a proper management within the ecotourism industry and promoting more the respective activity on a national level, the ecotourism can have a positive impact and get a positive role in the Finnish conflict.

Abstract: Northern boreal peatlands form one of the biggest carbon pools in the biosphere, thus having great potential to cause major changes to the global carbon cycle. The ongoing recent warming may affect the carbon dynamics though factors, such as, vegetation, hydrology and permafrost balance. As the future is still uncertain there are no definitive answers on how the peatlands will react in the future. Fortunately, moisture sensitive organisms such as, bryophytes and testate amoeba is preserved in the peat and can therefore be used to reconstruct past climatic shifts. In this thesis I studied palaeohydrology and peat accumulation over the last two millennia, from three peat cores originating in a permafrost peatland in Rogovaya, Russia. I used testate amoeba as a proxy of past moisture conditions and plotted the taxa composition of each core against 14C and 210Pb dated samples, to reconstruct past moisture shifts. The results were also supplemented by plant macrofossil and carbon accumulation data for more robust results. Of the three cores, Rog11 provided the oldest testate amoeba dataset by reaching the Dark Ages Cold Period. During this period there were indications of dry moisture conditions followed by a wet Medieval Warm Period. The Little Ice Age gave indications of a drying trend, while toward the end of the LIA Rog8 indicated opposite moisture conditions. From the end of the LIA onwards a general trend of drying and increased carbon accumulated is noted. Yet, during the last decade the trend has turned. The wet shift might indicate that the threshold for the peatland has been reached and the amount of melting permafrost has exceeded the evapotranspiration rate. As a conclusion my result indicates that the dynamics of both hydrology and carbon are complicated processes affected by both autogenic and allogenic factors, therefore causing large variability even on a local scale. The absence of widely spread observations of the most recent wet shift also indicates that the response of the peatland to the recent warming might be unequal. To rectify this situation, continued research is crucial, so that we can increase our understanding of climate-peatland interactions.

Eläinten hyvinvoinnin parantamiseen tarvitaan paljon tieteellistä tutkimusta tunteista, älykkyydestä ja lajikohtaisista tarpeista. Tiedon pohjalta pystytään vertaamaan ja kehittämään eläinten tarpeisiin soveltuvia olosuhteita, kuten esimerkiksi virikkeitä ja hoitomuotoja eläintarhoissa. Eläintarhaolosuhteissa eläimen hyvinvoinnin saavuttaminen vaatii kompromisseja tilan ja resurssien rajallisuuden vuoksi. Parhaimman kompromissin löytämiseen vaaditaan tieteellistä tietoa lajikohtaisesta hyvinvoinnista. Maisterintutkielman tarkoituksena oli tuottaa tieteelliseen tietoon perustuva arvio Korkeasaaren eläintarhassa valittujen lajien hyvinvoinnista, jotka olivat huulipekari (Tayassu pecari), pikkumarmosetti (Cebuella pygmaea), punaniskakenguru (Macropus rufogriseus) ja sinikurkkuara (Ara glaucogularis). Tavoitteena oli tunnistaa tekijät, jotka vaikuttavat kyseisten eläinten hyvinvointiin luonnossa, ja verrata niitä eläintarhassa olevien yksilöiden olosuhteisiin. Tämän avulla pystyttiin arvioimaan lajikohtaiset tarpeet, joita tarvitaan eläinten hyvinvoinnin saavuttamiseksi. Vertasin hyvinvointiarvioitani tutkittavien lajien hoitajien tekemiin arvioihin, minkä tarkoituksena oli ymmärtää, riippuuko hyvinvoinnin arvio arvioijan näkökulmasta, ja onko lajien välillä eroja siinä, mihin seikkoihin arvioitsijat kiinnittävät huomiota. Arviointeja vertailemalla oli myös tarkoitus löytää tekijöitä, joilla on vaikutusta lajikohtaiseen hyvinvointiin Korkeasaaressa, minkä pohjalta voidaan ymmärtää ja kehittää mahdollisia hyvinvointia edistäviä toimenpiteitä. Toteutin tutkielman eri osioissa. Ensin suoritin kirjallisuuskatsauksen, jonka tarkoituksena oli määrittää tutkittavien lajien käyttäytyminen ja hyvinvoinnin tarpeet tieteellisten tutkimuksien avulla. Seuraavaksi selvitin eläinten olosuhteet Korkeasaaressa aitauksien kuvaamisen, tarkastelun ja hoitajien haastattelujen avulla. Tämän jälkeen tein hyvinvointikartoitukset, joita verrattiin tutkittavien eläinten hoitajien vastaaviin tilastollisesti. Hoitajat arvioivat lajien hyvinvoinnin aina paremmaksi kuin minä, ja erot arvioinneissa olivat suuria. Johtopäätöksenä voidaan pitää, että suora vertailu luonnonympäristöön voi johtaa huomattavasti alhaisempiin hyvinvointiarvioihin. Tällainen on esimerkiksi trooppisen ilmaston vertailu Suomen vuodenaikaisvaihteluun, vaikka eläimillä olisi aina mahdollisuus suotuisiin lämpö- ja valaistusolosuhteisiin. Hyvinvoinnin osa-alueiden välillä ei ollut merkittäviä eroja. Huomattavaa on, että hoitajat arvioivat samalla myös paitsi eläimen olosuhteita, myös omaa onnistumistaan niiden hoidossa. Tämä voi vaikuttaa siihen, että yksikään laji ei saanut hoitajilta yhtään 1-arviota, kun taas minulla oli objektiivisempi ote arviointiin, sillä kritiikki ei kohdistunut minuun edes välillisesti.

Protein kinases are signaling molecules that regulate vital cellular and biological processes by phosphorylating cellular proteins. Kinases are linked to variety of diseases such as cancer, immune deficiencies and degenerative diseases. This thesis work aimed to identify direct substrates for protein kinases in the CMGC family, which consists of the cyclin-depended kinases (CDK), mitogen activated protein kinases (MAPK), glycogen synthase kinase-3 (GSK3) and CDC-like kinases (CLK). CMGC kinases have been identified as cancer hubs in interactome studies, but large-scale identification of direct substrates has been difficult due to the lack of efficient methods. Here, we present a heavy-labeled 18O-ATP-based kinase assay combined with LC-MS/MS analysis for direct substrate identification. In the assay, HEK and HeLa cell lysates are treated with a pan-kinase inhibitor FSBA which irreversibly blocks endogenous kinases. After the removal of FSBA, cell lysates are incubated with the kinase of interest and a heavy-labeled ATP, which contains 18O isotope at the γ-phosphate position. Resulting phosphopeptides are enriched with Ti4+- IMAC before the LC-MS/MS analysis, which distinguishes the desired phosphorylation events based on a mass shift caused by the heavy 18O. With this pipeline of methods, we managed to quantify and identify direct substrates for 26 members of CMGC kinase family. A total of 1345 substrates and 3841 interacting kinase-substrate pairs were identified in cytosolic cell lysates, from which 165 were annotated in the PhosphoSitePlus® database. To identify substrates for kinases with nuclear localization, ten kinases were tested with nuclear HEK cell lysate. We identified 194 kinase-substrate pairs, 141 of which were unique to the nuclear fraction and 27 annotated in the PhosphoSitePlus® database. Finally, kinases with outstandingly high amounts of novel substrates were subjected to gene ontology analysis. We were able to link the gene ontology classifications of novel substrates to the biological processes regulated by the kinase of interest. These results indicate that heavy-labeled 18O-ATP-based kinase assay linked LC-MS/MS is a useful tool for large-scale direct kinase substrate identification.

Plants control the exchange of gases through the stomatal pores. Stomata are formed by guard cells and the closure of stomata are regulated via a complex signaling network in response to various biotic and abiotic stimuli, such as pathogens, elevated levels of CO2 and darkness. The leucine-rich repeat receptor-like kinase (LRR-RLK) GUARD CELL HYDROGEN PEROXIDE-RESISTANT1 (GHR1) is part of the network regulating stomatal closure. GHR1 is an inaktive pseudokinase that can activate SLOW ANION CHANNEL-ASSOCIATED1 (SLAC1), an anion channel that is crucial for stomatal closure, via interacting proteins. The exact role of GHR1 is still partly unknown, however, it has been suggested that GHR1 could function as a scaffold or as an allosteric regulator of additional components required for stomatal closure. The aim of this study was to identify novel interactors of GHR1. First stable plant lines expressing fusion proteins GHR1-YFP, GHR1W799*-YFP and plain YFP as a negative control were generated and from these lines fusion protein expression levels and the subcellular localization were studied. Next the plant lines were used for purifying GHR1 interacting proteins with the use of co-immunoprecipitation and identification of the proteins with mass spectrometry. The unlikely GHR1 interactor candidates were then filtered from the mass spectrometry data. The subcellular localization and the protein expression of the interacting proteins were studied with the use of internet databases. Literature of the GHR1 interacting proteins were studied in order to make possible connections with GHR1 and stomatal closure. In this study 38 GHR1 interactors were identified. Literature search revealed that many of the identified interactors had a known role in stomatal movements. These included proteins such as PLASMA MEMBRANE INTRINSIC PROTEIN2-1 (PIP2-1) and BETA CARBONIC ANHYDRASE 4 (BCA4), that are known to have a role in stomatal closure. Future work includes confirming the interactions with independent methods and studying the molecular mechanisms related to stomatal movements. The GHR1 interactome identified here for the first time reveals novel parts of the network regulating stomatal movements and thus increases our understanding of molecular mechanisms behind stomatal functions.

This study aimed to identify conservation landscapes with potential to be mutually beneficial for people and African savanna elephants (Loxodonta africana) in South Africa through spatial conservation planning analyses that integrate ecological and socioeconomic data. The research questions were: (i) what are the most ecologically suitable areas for the reintroduction of elephants, and (ii) which of these areas provide the best opportunities for also sustaining socioeconomic development of local people. The first question was answered with an ecological model that predicts habitat suitability for elephants, developed by a combination of literature review, expert opinion, and GIS-based methods. The second question was answered by combining the ecological model with socioeconomic criteria in Zonation spatial conservation planning software. The results show that the central part of South Africa holds most potential for elephant conservation as it has the largest uniform area of high-quality habitat, while the area also meets the socioeconomic criteria. The priority areas for the conservation of elephants were classified into top priority classes of 1%, 2%, 5%, 10% and 20%. The identified areas hold an unrealized opportunity in the wildlife and ecotourism sectors, and the reintroduction of elephants to those areas could provide the foundation for long-term economic activity of local communities e.g. in the form of elephant-based ecotourism, while contributing to the conservation of elephants. Conserving just the top 5% priority areas would grow South African protected area estate by approximately three million hectares and increase the current elephant range by approximately 75%. Ideally, the results of this study could be used to inform the on-going decision-making process on where to allocate resources for elephant conservation in South Africa.