Bio- ja ympäristötieteellinen tiedekunta

 

Recent Submissions

  • Immanen, Juha (Helsingin yliopisto, 2018)
    Life on Earth is carbon based and is largely enabled by plants. As autotrophic (self-sufficient) life-forms, they fix atmospheric carbon dioxide to sugar through photosynthesis, and atmospheric oxygen is a by-product of this process. Heterotrophic life-forms such as humans are fully dependent on energy and oxygen provided by plants and cyanobacteria. Plants are thus very important for the human economy. Agriculture and forestry are large business sectors, with a global annual worth of over 3 trillion US dollars. Vascular tissues develop from the vascular cambium and serve two main functions: they give physical structure and support and distribute the water, nutrients and other substances needed for growth. Phloem transports the nutrients, and xylem is mainly responsible for conducting water and providing structural support for the plant. Trees are crucially important organisms for various ecosystems: about 31% of global terrain, over 4 billion hectares, is covered by forests. Trees contribute to carbon fixation, oxygen production, the global water cycle and soil erosion prevention. Woody biomass represents a valuable renewable source of energy and raw material for pulping and for the construction industry. Trees provide an excellent system for studying secondary vascular development in high resolution due to their massive size, which results from a wide cambial meristem. Understanding the hormonal regulation of radial growth that underlies wood development is of great importance for improving the use of tree products as a renewable resource. Detailed knowledge of these regulatory mechanisms could provide powerful tools to help tree breeders boost lignocellulosic biomass production. In the future, improved forestry and agriculture could produce the required energy, food, feed and raw biological materials using much less land, enabling the preservation of large areas of natural forest. Cytokinins are important growth regulators, but there are many other important plant hormones and growth regulators. In the work included in this thesis, we provided the first comprehensive description of the gene families for cytokinin signaling and homeostasis in black cotton wood (Populus trichocarpa). We also analyzed the hormone distribution and genome-wide expression profiles across the Populus cambial zone at an unprecedentedly high resolution. We observed increased cambial auxin concentration and auxin-responsive gene expression in cytokinin over-producing transgenic hybrid aspen (Populus) trees. Our results indicate that a graded distribution of cytokinin signaling specifies meristematic activity by influencing the amplitude of the cambial auxin gradient. It seems that cytokinins and auxin together regulate cambial developmental fate and growth rate. The two major phytohormones clearly interact in a manner which calls for continuing research. Here, we show that cytokinin and auxin display different yet partially overlapping distribution profiles across the cambium. In contrast to auxin, which is most concentrated in the actively dividing cambial cells, the greatest concentration of cytokinins is in the developing phloem. By manipulating the cytokinin concentration in the cambial zone, we could increase and decrease cambial cell division activity. Stimulation of cambial growth dramatically increased the production of lignocellulosic trunk biomass in Populus trees under greenhouse conditions.
  • Eldfors, Samuli (Helsingin yliopisto, 2018)
    This thesis describes three studies on the identification and analysis of somatic mutations in three respective types of leukemia: large granular lymphocytic leukemia (LGL leukemia), B-cell precursor acute lymphoblastic leukemia (ALL) with TCF3-PBX1/t(1;19) fusion (TCF3-PBX1 BCP-ALL), and acute myeloid leukemia (AML). In the first study, we analyzed the somatic mutations in an LGL leukemia index patient and identified a mutation in STAT3. A subsequent cohort analysis found that the STAT3 gene was mutated in 40% of LGL leukemia patients. Functional validation showed that the identified STAT3 mutations are activating oncogenic mutations. Our results indicate that STAT3 mutations can be used as mutational biomarkers in the diagnosis of LGL leukemia. In the second study, we analyzed the somatic mutations, aberrant gene expression, and drug sensitivities in a relapsed TCF3-PBX1 BCP-ALL patient. We found that the patient’s cells were exceptionally sensitive to the phosphoinositide 3-kinase inhibitor idelalisib. Idelalisib has been previously approved for the treatment of chronic lymphocytic leukemia. The patient’s leukemia was highly sensitive to Idelalisib despite harboring loss-of-function mutations in TP53, which are associated with poor prognosis in ALL. Idelalisib sensitivity was confirmed in TCF3-PBX1 BCP-ALL cell lines. Our results indicate that idelalisib is a promising drug for the treatment of relapsed TCF3-PBX1 BCP-ALL. In the third study, we analyzed somatic mutations, copy number, and cytogenetic aberrations in newly diagnosed and relapsed AML patients. By comparing mutations frequencies in these two conditions, we found that mutations in CBL, PTPN11, NF1, and KRAS are both enriched and selected for in relapsed AML patients. Mutations in these genes have been shown to induce cytokine hypersensitivity which may confer cytokine protection and increased survival of leukemic cells during therapy. Our results show that mutations in these genes are common in relapsed AML patients and are under positive selection. We also found that mutations in TP53, WT1, PHF6 as well as trisomy 8 and monosomy 7/del(7q) are enriched and selected for in relapsed AML patients.
  • Maaninka, Katariina (Helsingin yliopisto, 2018)
    Atherosclerosis is a slowly progressing disease characterized by the development of inflamed fatty deposits called atherosclerotic lesions or plaques within the innermost layer of the arterial wall, the intima. First subclinical atheromatous changes within the arterial intima begin early in childhood as a response to cholesterol accumulation and serve as precursors for more advanced lesions that develop during the following decades through complex series of cellular events. During the degenerative process of lesion formation, the affected arteries gradually lose their normal structure and function, and may become narrowed by the growing plaques, a process leading to impeded blood flow to end-organ. Eventually multiple subclinical cellular events may result in the development of unstable, vulnerable plaques that are prone to rupture culminating in the life threatening clinical manifestation of myocardial infarction or ischemic stroke. Atherosclerosis is strongly associated with lifestyle and age. Enhanced plasma level of low-density lipoprotein (LDL) is considered a definitive risk factor of atherosclerosis, emphasizing the crucial role of LDL in the process of lesion development. Circulating LDL particles are the major carriers of cholesterol to the arterial intima and their retention by subendothelial proteoglycans and subsequent modification within the intima drives intramural cholesterol accumulation and ensuing formation of atherosclerotic lesions. Another lipoprotein playing a central, yet opposite role in atherogenesis is the high density lipoprotein (HDL). HDL and its principal protein component, apolipoprotein A (apoA)-I, act against atherosclerotic lesion development by removing cholesterol from the arterial wall and delivering it to liver for excretion. In addition to this process referred to as reverse cholesterol transport, HDL and apoA-I possess various anti-inflammatory properties by which they may hinder the inflammatory processes involved in the plaque development, and thus to protect from the disease progression. However, similar to LDL, HDL within the arterial intima is subjected to various structural and compositional modifications that impair its normal physiological function leading to generation of HDL particles with reduced antiatherogenic properties. LDL and HDL modifications within the arterial intima can be induced by extracellular enzymes released from cells present in the intima. MCs are tissue-dwelling effector cells of innate and adaptive immunity that differentiate from committed circulating progenitor cells and are present in increased numbers in atherosclerotic lesions. Being capable of releasing large amounts of various neutral proteases upon activation, MCs are potential source of LDL and HDL proteolyzing enzymes. The aim of this thesis was to investigate the ability of various human MC neutral proteases to cleave the protein moieties of LDL and HDL particles, apoB-100 and apoA-I, respectively and thus to contribute to the pathogenic processes of LDL accumulation and inflammation both essential in development of atherosclerotic lesions. In the present study, human MCs expressing four tryptase isoforms, chymase, carboxypeptidase A3, cathepsin G, and granzyme B is described. Of these, chymase and cathepsin G efficiently proteolyzed apoA-I and apoB-100, respectively generating C-terminally truncated apolipoproteins. Proteolysis by chymase resulted in reduced anti-inflammatory properties of apoA-I and impaired the ability of apoA-I to induce cholesterol efflux from macrophage foam cells. Furthermore, proteolysis of apoB-100 by cathepsin G induced formation of fused LDL particles with increased binding to human aortic proteoglycans and atherosclerotic lesions of human carotid arteries. Taken together, the data presented in this thesis propose novel mechanisms by which human MC neutral proteases may promote lipid accumulation and inflammation within the arterial wall and thus promote lesion development and progression. .
  • Kettunen, Elina (Helsingin yliopisto, 2018)
    Fungi are one of the most diverse groups of organisms, but their fossil record is scarce compared with that of plants and animals. However, many fossils of microfungi have survived as inclusions in amber, which is fossilized resin produced by ancient trees millions of years ago. Some of these fossils were already discovered and described during the 19th century. There are important sources of Palaeogene amber in Europe: the Baltic and Bitterfeld deposits. Baltic amber is about 43–25 Ma old (Eocene), whereas Bitterfeld amber is slightly younger (approx. 24 Ma, Oligocene). The aim of this thesis was to increase our knowledge of the microfungi preserved in the Baltic and Bitterfeld ambers. The material studied included both historic collections and previously unstudied amber specimens. The approach led to several advances in the field of palaeomycology. The systematic affinities of the microfungi described by Robert Caspary and Richard Klebs over a century ago were reassessed. None of these historical specimens belong to the extant fungal genera they were originally assigned to. Amended descriptions were provided for the historical specimens, and several new types of fungi were described from novel amber specimens. These included the first fossils of lichen-associated filamentous fungi. The results demonstrate that relatively few fossil microfungi in amber can be identified accurately enough to be used as minimum age constraints in dating phylogenetic trees of different fungal lineages. All the fossil fungi studied grew either on or in the immediate vicinity of resin-producing trees, which made them likely candidates for preservation in amber.
  • Putkinen, Anuliina (Helsingin yliopisto, 2018)
    Boreal peatlands are highly important sinks for carbon (C). This function is enabled largely by one peat-forming plant, the Sphagnum moss. In addition to slowing the decomposition by gradually creating ombrotrophic conditions, it gives a shelter for the organisms mitigating the emissions of methane (CH4) – an effective greenhouse gas formed in submerged, anoxic peat layers. These organisms, methane oxidizing bacteria (methanotrophs, MOB), inhabit the dead, water-filled hyaline cells of the Sphagnum and provide the plant carbon dioxide (CO2) derived from the CH4 oxidation. While several studies have confirmed the presence of Sphagnum-associated methanotrophs (SAM), it is still unclear how dependent they are on the mosses and how environmental conditions affect their community composition and activity. This thesis evaluated SAM dynamics in the different stages of peatland development on both pristine and disturbed areas. Studies were based mainly on molecular methods, targeting the MOB-specific pmoA gene, and laboratory incubations, including stable isotope probing. In the first study, the connection between the SAM and the mosses was assessed by testing whether the SAM will disperse through the water phase. This trait, considered to represent a facultative symbiosis, was demonstrated in two experiments. In the field, mosses inactive in CH4 oxidation were transplanted next to active ones. Within a month, SAM communities of the neighboring mosses become more similar. The water-based colonization was further confirmed by bathing inactive mosses in flark pore water that showed high CH4 oxidation activity. Within just 11 h, activity was induced and the SAM abundance significantly increased in the treated mosses. The other two studies revealed similar SAM community composition patterns on a pristine chronosequence and on a gradient of re-vegetating cutover peatlands. Instead of the Sphagnum species, the general environmental conditions seemed to control the SAM community composition. Different types of SAM seemed to have their preferred environmental niches, with the type Ia MOB present and active especially in the young succession stages and the type II MOB in the older, hydrologically more stable stages. Despite the community differences, the potential CH4 oxidation did not differ along the gradients, suggesting functional redundancy. Only some drier bog samples did not show any detectable CH4 oxidation, demonstrating the regulatory role of the water table level on the SAM activity. The peat layers of the cutover gradient showed similar MOB community patterns but the potential CH4 oxidation increased with succession. The ability to disperse through the water provides a recovery mechanism from disturbances such as droughts, which are predicted to increase with climate warming. In addition, the diversity and functional redundancy of the SAM communities enhance the resilience of this important CH4 biofilter formed by the living Sphagnum mosses. The potential SAM activity in the mosses of the youngest cutover site promotes the Sphagnum transplantation practice as a tool to not only enhance the re-vegetation process, but also to mitigate the CH4 emissions formed in the rewetting and restoration of disturbed peatlands.
  • Perälä, Tommi (Helsingin yliopisto, 2018)
    Considering the uncertainties about model parameters and structure, observations, and the resulting predictions is crucial when managing natural resources. This thesis is compiled of four research articles that deal with uncertainties and change in marine fish populations. In all the articles, the Bayesian statistical modelling framework is adopted to account for various types of uncertainty. When modelling complex systems, one must recognize that all models are only approximations of reality and are based on the limited understanding about the system at that moment and on previously observed behaviour. Even if a model explains the recent behaviour and predicts how the system will respond in the near future, there is always a possibility that the system changes thus rendering the old model useless. Approaches to respond to sudden changes in the system’s dynamics are vital for successful resource management. Fish populations' renewal ability is mainly determined by their reproductive success. Thus, it is of utmost importance to be able to understand and model the reproductive dynamics of marine fish populations. In this thesis, fish reproduction is studied using models that link together the number of new individuals entering the adult population (recruits) and the size of the reproducing component of the population (spawning stock), namely, the stock-recruitment relationship. The focus in this work is on temporal and density-dependent variability in the stock-recruitment relationship. The temporal variability is studied using Bayesian change-point detection methods applied to detecting changes in the per capita reproductive output of four Atlantic cod populations by analysing recruit-per-spawner ratios, and in the parameters of the stock-recruitment relationship of four fish species in the southern Gulf of St. Lawrence. In this work, novel Bayesian methods are utilized to improve the handling of uncertainties about the parameters, the timings of the changes, and in short term predictions. This thesis presents computational methods for analysing temporal variability in linear-Gaussian problems where analytical solutions are available, and extend the methodology to analytically intractable non-linear and non-Gaussian problems by utilizing numerical approximate solutions. Traditionally, compensatory population dynamics have been assumed in marine fish populations. The validity of the assumption of compensatory stock-recruitment relationship is examined by studying the density-dependence of nine Atlantic herring populations at low population sizes using models that better capture the uncertainty related to low-abundance dynamics and allow for depensatory dynamics caused by Allee effects. The Allee effect has been largely ignored in marine fish population models. Here methodology for detecting Allee effects is developed, and it is concluded that Allee effects might be more common than previously thought. In Bayesian models, expert elicitation is used to construct prior distributions for the model parameters. When observational data is scare or not available, informative prior distributions are crucial for conducting inference. However, experts might not be calibrated and can have significant biases in their assessments about key parameters of the models. Statistical models are developed for the use of cacalibration data to infer and to correct for experts’ biases in their assessments.
  • Viljanen, Martta (Helsingin yliopisto, )
    Adaptation to environmental light conditions at different time scales and biological levels was studied using the visual system of opossum shrimps (genus Mysis) as model. The focus of the study was on two aspects of visual adaptation: 1) mechanisms behind spectral tuning which enables effective photon catch in different light environments and 2) photo-induced damage and protective mechanisms in the eyes arising as a trade-off from tuning the visual system to be highly sensitive. For spectral adaptation studies mysids representing 12 species were collected from different water bodies around circumpolar and Caspian areas. Their opsin genes were sequenced and compared with phylogenetic relationships. Spectral sensitivities were determined for 15 populations representing four species by recording single-rhabdom absorption spectra with microspectrophotometer. Water transmission spectra were measured and the wavelength of maximal transmission of light and the attenuation coeffcient was determined to quantify the light conditions in the respective habitats. Animals originating from different environments were also bred in carefully controlled laboratory conditions to observe possible effects of ambient salinity on spectral sensitivity. The photoprotective mechanisms were studied by subjecting animals from populations with intrinsically different vulnerability to light-induced damage to an ultra-slow light acclimation procedure before exposing them to a bright light. The effects of this procedure were examined structurally by transmission electron microscopy and functionally by electroretinography. The equilibrium between rhodopsin and metarhodopsin was studied by microspectrophotometry. The acclimation protocol was conducted at different speeds to investigate the time scale of light acclimation. The studies of spectral tuning show that the spectral sensitivity of different Mysis populations generally correlates in an adaptive manner with the light conditions in their living environment. However, neither opsin gene sequence nor water light transmission could fully explain the observed differences in spectral sensitivities between study populations. Neither were there differences in chromophore use. The findings indicate that there are two opsin genes which are expressed in different proportions following a reaction norm triggered by an yet unidentified environmental factor. This hypothesis still requires more investigation. The study on light damage shows that very slow light acclimation can prevent structural and functional deterioration of photoreceptors caused by bright light exposure. The time scale of successful acclimation corresponds to the tempo of seasonal changes of light levels in the natural habitat. One key player in this phenomenon seems to be the amount of native visual pigment.
  • Kauppi, Laura (Helsingin yliopisto, 2018)
    Introductions of non-native species are changing the composition of plant and animal communities worldwide. In order to assess their ecological and socio-economic impacts it is imperative to know their effects on the surrounding ecosystem. Basic knowledge of the non-native species’ biology, ecology and effects on ecosystem functions in their new environment is therefore needed but often lacking. Naturally low species richness and frequent disturbances make the Baltic Sea one of the most heavily invaded seas in the world. One of the most successful invaders has been the spionid polychaete genus Marenzelleria, three species of which now occur in the Baltic Sea, M. viridis, M. neglecta and M. arctia. Their differing burrowing and ventilation behavior compared to the native species suggest an impact on nutrient cycling. A combination of monitoring data, field surveys and laboratory experiments were used to investigate the ecosystem effects of Marenzelleria spp. The genus occurs in the entire Baltic Sea with highest densities in deeper (over 30 m) areas. M. arctia prefers deeper sites, all three species co-occur at muddy sites up to 20 m depth, and M. viridis and M. neglecta occur together and hybridize at sandy sites. Population dynamics and productivity of the three species differ spatially. Seasonal differences in biotic and abiotic factors lead to variation in the relative importance of Marenzelleria spp. on an important ecosystem function, nutrient cycling. The contribution of Marenzelleria spp. to nutrient cycling was highest during spring when food input to the benthos is high. M. arctia, M. neglecta and M. viridis had density-dependent effects on bioturbation metrics and solute fluxes, implying spatial and temporal variation in their impact on nutrient cycling following seasonal and spatial changes in their densities and biomass. The impact could be modified by the composition of the surrounding macrofauna community and the variation in abiotic factors. Combining the observational and experimental results implies a possible enhancement of phosphorus binding capacity by Marenzelleria spp. in deeper areas especially during summer when oxygen conditions deteriorate and densities increase, but an enhancement of phosphate effluxes in normoxic areas through enhanced remineralization of organic matter. Through density-dependent effects on bioirrigation and directly on ammonium fluxes, the genus also has an impact on nitrogen cycling. The results from this thesis imply spatial and seasonal differences in the impact of Marenzelleria spp. on nutrient cycling related to the environmental conditions and to the densities and biomasses of Marenzelleria spp. and other macrofauna. At disturbed sites Marenzelleria spp. could possibly enhance phosphorus burial and thus remove nutrients from primary production, whereas at undisturbed, normoxic sites they could enhance organic matter remineralization thus preventing deposition of large quantities of organic matter on the sea floor. The results also highlight the need to study the effects on non-native species in the natural environment incorporating the spatial and seasonal variability, and natural community composition in order to accurately estimate their contribution to ecosystem function.
  • Wang, Cui (Helsingin yliopisto, 2018)
    Pleistocene glaciations have profoundly influenced the genetic diversity of organsims in the Northern Hemisphere. Large ice sheets covered vast areas of the Eurasian continent, driving species southward to different isolated refugia, often resulting in deep divergences within species. Phylogeographic studies carried out on Pungitius species based on mitochondrial DNA (mtDNA) support profound intraspecific genetic divergence in refugia during glaciation cycles. However, compared to species distributed at lower latitudes, those distributed at higher latitudes may have also occurred in cryptic refugia in periglacial areas during glaciations, complicating the inferences of the phylogeographic patterns of the fish species with a circumpolar distribution, such as the Pungitius sticklebacks. Moreover, comprehensive phylogenetic studies of Pungitius species have been lacking in the sense that not all extant species have been included into analyses. In this dissertation, I carried out phylogeographic studies on seven Pungitius species using both mtDNA and genome-wide nuclear SNP markers, with worldwide sampling of populations to shed light on intra- and interspecific divergence in this genus, as well as to study their historical demography and interspecific hybridization. By sequencing five mtDNA regions, I found six highly divergent Pungitius lineages including those corresponding to P. pungitius, P. platygaster, P. tymensis and P. kaibarae, and two independent monophyletic lineages of P. laevis. I also found a third lineage of P. laevis that clustered together with P. pungitius. To understand whether this clustering of the P. laevis lineage III and P. pungitius mtDNA was a result of convergence or interspecific introgression, I conducted phylogeographic and population genetic analyses using both mtDNA and nuclear gene sequences. The results indicated asymmetric mitochondrial introgression from P. pungitius to P. laevis and genetic admixture of these species. Hence, the results suggest that the P. laevis lineage III has experienced historical hybridization. Deep intraspecific mitochondrial divergence was found within P. laevis in central and southern France, coinciding with major drainages, suggesting that these areas correspond to distinct glacial refugia for the species explaining the observed intraspecific divergence. To further clarify evolutionary relationships between different Pungitius species and populations, as well as to study the prevalence and extent of introgression among recognized species, phylogenomic datasets were constructed from restriction-site associated DNA in combination with mitochondrial genomes. All divergences in the Western Palearctic were estimated to have occurred during the Pleistocene (≤ 2.6 Ma). The phylogenetic patterns suggest a major split in Pungitius genus occurred early in history, resulting in an East Asian group (P. kaibarae, P. tymensis, P. sinensis) and European - North America group (P. hellenicus, P. platygaster, P. laevis and P. pungitius). The genus probably originated from the Western Pacific and spread to Europe and North America through the Arctic Ocean in several waves after the opening of the Bering Strait. Four cases of incongruence between nuclear and mtDNA-based trees revealed evidence for frequent hybridizations and mitogenome capture during the evolutionary history of this genus. Further analyses of these four cases of cytonuclear incongruence also revealed evidence for nuclear introgression, but the estimated levels of autosomal introgression were low.
  • Kovac, Bianca (Helsingin yliopisto, 2018)
    Actin cytoskeleton is essential in generating mechanical forces together with the associated adhesions and transmitting signals that impact processes such as cell migration. Cell migration is necessary for numerous biological processes including wound healing and embryonic development. Moreover, aberrant cell migration promotes cancer invasion and metastasis. Cell migration events require dramatic spatial and temporal reorganization of the actin cytoskeleton that involves coordinated formation and regulation of multiple structures such as actin stress fibers. Actin stress fibers are dynamic structures, which differ in their subcellular localization, connection to substratum and their dynamics. However, these actin stress fibers are less characterized in terms of the molecules required for assembly-disassembly and the signaling pathways involved in regulating their functions in mesenchymal and epithelial cells. This thesis focuses on characterizing key players and signaling pathways involved in regulating actin stress fiber assembly-disassembly, cell adhesion and contractility. Understanding these cell plasticity changes is essential as in cancer context they are likely to be deregulated thus leading to increased migratory and invasive potential of the cells. During this thesis study, NUAK2 a novel serine-threonine kinase was identified to associate with myosin phosphatase Rho-interacting protein (MRIP) on actin stress fibers. Association between NUAK2 and MRIP increases cells contractility and promotes formation of actin stress fibers through phosphorylation of myosin light chain (MLC). The identified NUAK2-MRIP association reveals a novel mechanism for the maintenance of actin stress fibers. Our findings implicate NUAK2 as an important regulator of cell contractility and actin stress fiber assembly. Thus providing further knowledge of how actin stress fibers and cell contractility can be regulated in mesenchymal cells. To further characterize the specificity of molecules required for the assembly of actin stress fibers, we studied the function of most abundant actin crosslinking proteins in non-muscle cells, α-actinin-1 and α-actinin-4. Our findings reveal that specifically α-actinin-1 and not α-actinin-4, is required to assemble dorsal stress fibers found at the leading edge of mesenchymal cells. In addition, loss of α-actinin-1 modulates cell-matrix adhesions leading to decreased cell migration without altering cells contractility. Contrary to traditional views, dorsal stress fibers assembled by α-actinin-1 are non-contractile and are induced by Rac1 signaling. Rac1 is essential in regulating polymerization of actin filaments. Thus suggesting that force required for cell migration is at least partially generated through actin polymerization. Interestingly, we found α-actinin-1 to be upregulated in various cancers and especially associates with decreased survival in estrogen receptor (ER) negative breast cancer patients. In mammary epithelial cells, α-actinin-1 levels regulate epithelial cell plasticity, reorganize actin stress fibers and destabilize cell-cell adhesions accompanied with increased cell migration. This thesis extends the knowledge of especially α-actinin-1 in regulating actin stress fiber assembly and cell plasticity in both epithelial and mesenchymal cells. Furthermore, identifying α-actinin-1 as a candidate prognostic biomarker in ER negative breast cancer patients.
  • Tauscher, Petra (Helsingin yliopisto, 2018)
    The variety of species in the animal kingdom notwithstanding, early embryogenesis frequently relies on the same set of molecules. These highly conserved molecules are considered genetic toolkit genes; a set of genes that determines the body plan in various species. For example, the determination of the dorsal-ventral (D/V) axis often relies on bone morphogenetic proteins (BMPs). Studies in various organisms, vertebrates as well as invertebrates, established that the BMP signaling network is crucial for localizing the position of the central nervous system. Given that the same set of molecules is used repeatedly in different species, the question of how animal diversity could evolve arises. In this study, I focused on BMP signaling in the fruit fly Drosophila melanogaster. In Drosophila, BMP signaling takes place at various stages of development. Amongst others, it is crucial for D/V patterning of the early embryo, for growth and patterning of the larval wing imaginal disc, as well as for formation of longitudinal and posterior crossveins (PCV) in the pupal wing. The Drosophila genome contains three BMP-type ligands, Decapentaplegic (Dpp), a BMP2/4 type ligand, and the paralogs Screw (Scw) and Glass bottom boat (Gbb), both of which are BMP5-8-type ligands. Interestingly, Scw is exclusively found in higher Dipterans such as Drosophila, and is expressed only in the early embryo, whereas Gbb is repeatedly used during fly development. Previous studies revealed that Scw can replace Gbb in PCV formation in the pupal wing. On the other hand, Gbb cannot function in the context of early embryogenesis. Based on these facts, Gbb and Scw constitute a highly suitable model to study protein divergence and evolution. Here, I examined how post-translational modifications of BMP-type ligands may affect the signaling outcome, and hence, animal development; the focus was on proteolytic processing and N-glycosylation. BMP-type ligands are produced as inactive pro-proteins and need to be proteolytically processed in order to become mature ligands. Scw has two cleavage sites crucial for fly viability. I showed in a cell–based signaling assay that proteolytic processing of both cleavage sites is needed for rapid, peak-level BMP signaling. Furthermore, there exists one highly conserved N-glycosylation motif in the mature BMP ligand domain. In addition, Scw exhibits a unique N-glycosylation site which is not present in other BMP-type ligands. Cell culture-based signaling assays revealed that both N-glycosylation motifs of Scw ligand are needed for peak level signaling. In addition, lack of N-glycosylation motifs negatively affects the BMP signaling outcome in the early embryo, as well as fly viability. In contrast, fewer N-glycosylations appear to be beneficial in the context of PCV formation. These findings suggest that post-translational modifications of BMP ligands play a role in signal transduction and therefore in animal development. Furthermore, I propose that post-translational modifications can act as flexible modules to allow adaption of conserved molecules to different developmental contexts. Besides, understanding how post-translational modifications affect BMP signaling outcome may improve the chances to develop more signaling-efficient BMPs needed in clinical applications.
  • Mätlik, Kärt (Helsingin yliopisto, 2018)
    The human genome contains approximately 20,000 protein-coding genes and tens of thousands of genes for non-coding RNA. Understanding the function of these genes in a living organism is crucial for learning about key processes in early development, mechanisms of organ function and human behavior. Importantly, many human diseases are caused by abnormal gene expression. Therefore, understanding the causes and pathological processes behind these disorders could provide means for early diagnosis, prevention or treatment. The main aim of this thesis was to develop new strategies for studying gene function and regulation in experimental animals. The techniques described in this work allow increasing gene expression specifically in those cell types, which naturally express the gene. The results presented in this thesis suggest that increasing gene expression in correct physiological context may potentially have widespread applications in basic biological studies and for understanding human disease.
  • Gui, Jinghua (Helsingin yliopisto, 2018)
    In this thesis, I mainly investigate how BMP/Dpp signaling is involved in development of the early pupal wing of Drosophila, and the mechanisms coupling Dpp signaling with morphogenesis.
  • Lume, Maria (Helsingin yliopisto, 2018)
    Neurotrophic factors are small secretory proteins with important functions both in the nervous system and in peripheral tissues. Glial cell line-derived neurotrophic factor (GDNF) is best known for its ability to support the survival of midbrain dopaminergic neurons and enteric neurons. Also, GDNF is essential for the development of kidney and testis. It has been shown that both the absence and excessive amounts of GDNF protein negatively regulate kidney morphogenesis, highlighting the importance of proper spatiotemporal regulation of GDNF. Despite the wealth of knowledge regarding GDNF functions both in and outside the nervous system, relatively little is known about the trafficking mechanisms of GDNF. GDNF is synthesized as a precursor protein, proGDNF. In this thesis, we characterized the cellular localization and secretion of two GDNF splice variants, pre-(α)pro-GDNF and pre-(β)pro-GDNF, that differ in their pro-regions. Both precursor forms were shown to be secreted from cell lines. However, while (α)pro-GDNF co-localized mainly with the Golgi markers, the (β)pro-GDNF was found primarily in the secretogranin-II positive vesicles of the regulated secretory pathway. In accordance, the two splice isoforms responded differently to KCl-induced depolarization that is known to trigger the secretion of neurotrophin family members in neuronal cells. Only (β)pro-GDNF and corresponding mature GDNF were secreted activity-dependently, whereas (α)pro-GDNF and its corresponding mature GDNF were secreted via the constitutive secretory pathway. In addition, we determined which enzymes are responsible for the proteolytic cleavage of proGDNF into mature GDNF. To elucidate, whether secreted proGDNF has any biological activity, the recombinant cleavage-resistant proGDNF mutant protein was expressed in mammalian CHO cells and next purified from the media. Our results demonstrate that proGDNF is biologically active. Furthermore, similarly to mature GDNF, proGDNF can signal via the GDNF receptor α1/RET receptor tyrosine kinase complex and activate downstream MAPK and AKT pathways. Interestingly, proGDNF is not able to activate RET via the GFRα2 receptor. Finally, we identified a novel sorting receptor for GDNF and its receptors. Our results show that SorLA, a member of the vacuolar protein sorting 10-p domain receptor family, can internalize GDNF and GFRα1. While GDNF is subsequently degraded in lysosomes, GFRα1 is recycled back to the cell membrane. In the presence of SorLA and GFRα1, also RET is internalized and directed to early endosomes. By regulating the availability of GDNF and its co-receptors, SorLA can inhibit GDNF-induced neurotrophic activity in SY5Y cells. Moreover, SorLA seems to regulate intracellular localization of GFRα1 in hippocampal neurons. In summary, results of this thesis characterize the cellular regulation of GDNF regarding its secretion, processing, internalization and subsequent degradation. Furthermore, this is the first time that biological functions of the GDNF precursor protein proGDNF are described. Our findings indicate that the trafficking of GDNF is very different from that of other neurotrophic factors, and in contrast to apoptotic proneurotrophins, proGDNF is a trophic protein with increased specificity to GDNF receptor complex GFRα1-RET.
  • Rosa, Elena (Helsingin yliopisto, 2018)
    Variation in biotic and abiotic ecological factors unavoidably shapes the life history of living organisms. The ability to integrate environmental cues and respond adequately can mark the difference between life and death. Phenotypic plasticity consists of a rapid response to variation in environmental variables, and it entails the ability of one genotype to generate multiple phenotypes over an environmental gradient. However, the degree of plasticity a phenotypic trait can display is limited by physiological and ecological constraints, which may impose trade-offs on other traits. Insects are an excellent system to explore phenotypic plasticity, because of their presence at all ecosystem levels making them important bio-indicators, and their occurrence in large numbers with a fast generation time making them optimal study subjects. The aim of this thesis is to uncover the degree of phenotypic plasticity in immunity and life-history traits displayed by a butterfly occurring in the temperate zone, the Finnish Glanville fritillary (Melitaea cinxia), in response to variation in the following ecological factors: food quality, larval density, susceptibility to a pupal parasitoid, food limitation and exposure to cold spells. The ecological factors were tested on different life stages of the butterfly, based on their ecological relevance. Particular importance is given to potential trade-offs among life-history traits, with special attention to the immune response. The immune system entails a complex of processes and structures that are activated any time insects interact with their environment, and hence its regulation and maintenance are costly. Therefore, extremely plastic immune responses may not be optimal. The study approach involved a combination of laboratory manipulation of the ecological factors tested, collection of observational data of adult life history and mobility in a semi-natural enclosure, and three different immune assays (encapsulation rate, phenoloxidase activity and immune gene expression). Interactions with two components of the natural community were also included: the host plant pathogen powdery mildew (Podosphaera plantaginis), and the generalist pupal parasitoid wasp Pteromalus apum. Key findings include a poorer performance of larvae fed on a diet including powdery mildew, and the upregulation of two immune genes on the same diet. This suggests that larvae feeding on infected host plants may pay a twofold cost of poor nutrient intake and unnecessary immune activation, potentially due to interaction with the plant pathogen. Moreover, larvae reared in high density showed a better performance and a greater ability to kill brood members of a pupal parasitoid wasp, which, however, did not help butterfly survival. This finding indicates that larvae of this species benefit from group living and do not suffer from stressful interactions in absence of food limitation. Further, a trade-off between pupal investment in the encapsulation immune response and adult mobility was detected, indicating potential costs of immunity in regard to expensive activities such as flight. Finally, cold spells during the pupal stage induced an increased melanisation of adult wings. In males, this response also led to a strong immune reaction to a bacterial challenge and markedly reduced lifespan, suggesting a condition-dependent cost of immunity. The findings indicate that the immune system is affected to some extent by most of the factors tested, but the degree of plasticity displayed may be constrained by costs arising from indiscriminate immune activation.