Molecular Mechanisms of Sexual Dimorphism in Threespine Stickleback

Sexual dimorphism is commonly understood as differences in external features, such as morphological features or coloration. However, it can more broadly encompass behavior and physiology and at the core of these differences is the genetic mechanism – mRNA and protein expression. How, and which, molecular mechanisms influence sexually dimorphic features is not well understood thus far. DNA, RNA and proteins are the template required to create the phenotype of an individual, and they are connected to each other via processes of transcription and translation. As the genome of males and females are almost identical with the exception of the few genes on the sex chromosome or the sex-determining alleles (in the case of organisms without sex chromosomes), it is likely that many of the downstream processes resulting in sexual dimorphism are produced by changes in gene regulation and result from a regulatory cascade and not from a vastly different gene composition. Thus, in this thesis a systems biology approach is used to understand sexual dimorphism at all molecular levels and how different genomic features, e.g. sex chromosome evolution, can affect the interplay of these molecules. The threespine stickleback, Gasterosteus aculeatus, is used as the model to investigate molecular mechanisms of sexual dimorphism. It has well-characterized ecology and behavior, especially in the breeding season when sexual dimorphism is high. Moreover, threespine stickleback has a recently evolved Y chromosome in the early stages of sex chromosome evolution, characterized by a lack of recombination leading to degeneration (i.e. gene loss). The aim of my thesis is to investigate how the genotype links to the molecular phenotype and relates to differences in molecular expression between males and females. Based on previous research on sex differences in mRNA expression, I investigated sex-biased protein expression in adult fish outside the breeding season to see if differences persisted after translation. As sex-biased expression also prevailed in the proteome and previous transcription expression seemed to be related to the sex chromosomes, I investigated the genome level with a particular focus on the sex-chromosomes. I characterized the status of Y chromosome degeneration in the threespine stickleback and its effects on gene function. Furthermore, since the degeneration process leaves genes in a single copy in males, I examined whether the resulting dosage difference of messenger RNA for hemizygous genes is compensated as it is in other organisms. In addition, threespine sticklebacks have wellcharacterized behavioral differences related to the male’s social status during the breeding season. To understand the connection between the genotype and behavior, I examined gene expression patterns related to breeding behavior using dominant and subordinate males as well as femaleSiirretty Doriast

Influence of interspecific interference competition on the genetic structure of Calopteryx splendens populations

Understanding the effects of interspecific competition on genetic diversity will deepen our knowledge on species evolution. In the case of Calopteryx splendens and C. virgo, sympatric damselfly species, interspecific interference competition by C. virgo has remarkable effects on territoriality of C. splendens resulting in reproductive character displacement. Since territoriality is correlated with phenotype and mating success, we investigated the effects of interspecific interference competition on genetic diversity of C. splendens populations. Using amplified fragment length polymorphisms (AFLP), we determined the population genetic structure of 12 C. splendens populations and used the genetic diversity information to relate heterozygosity of C. splendens to abundance of C. virgo in sympatric populations. We found that heterozygosity of C. splendens males decreased with increasing abundance of C. virgo males. This result most likely reflects changes in effective population size due to interspecific interference competition and shows an effect on genetic structure in damselfly populations.</p

Rapid changes in DNA methylation associated with the initiation of reproduction in a small songbird

Species with a circannual life cycle need to match the timing of their life history events to the environment to maximize fitness. However, our understanding of how circannual traits such as timing of reproduction are regulated on a molecular level remains limited. Recent studies have implicated that epigenetic mechanisms can be an important part in the processes that regulate circannual traits. Here, we explore the role of DNA methylation in mediating reproductive timing in a seasonally breeding bird species, the great tit (Parus major), using genome-wide DNA methylation data from individual females that were blood sampled repeatedly throughout the breeding season. We demonstrate rapid and directional changes in DNA methylation within the promoter region of several genes, including a key transcription factor (NR5A1) known from earlier studies to be involved in the initiation of timing of reproduction. Interestingly, the observed changes in DNA methylation at NR5A1 identified here are in line with earlier gene expression studies of reproduction in chicken, indicating that the observed shifts in DNA methylation at this gene can have a regulatory role. Our findings provide an important step towards elucidating the genomic mechanism that mediates seasonal timing of a key life history traits and provide support for the idea that epigenetic mechanisms may play an important role in circannual traits.Peer reviewe

Regulatory Architecture of Gene Expression Variation in the Threespine Stickleback Gasterosteus aculeatus

Much adaptive evolutionary change is underlain by mutational variation in regions of the genome that regulate gene expression rather than in the coding regions of the genes themselves. An understanding of the role of gene expression variation in facilitating local adaptation will be aided by an understanding of underlying regulatory networks. Here, we characterize the genetic architecture of gene expression variation in the threespine stickleback (Gasterosteus aculeatus), an important model in the study of adaptive evolution. We collected transcriptomic and genomic data from 60 half-sib families using an expression microarray and genotyping-by-sequencing, and located expression quantitative trait loci (eQTL) underlying the variation in gene expression in liver tissue using an interval mapping approach. We identified eQTL for several thousand expression traits. Expression was influenced by polymorphism in both cis- and trans-regulatory regions. Transe-QTL clustered into hotspots. We did not identify master transcriptional regulators in hotspot locations: rather, the presence of hotspots may be driven by complex interactions between multiple transcription factors. One observed hotspot colocated with a QTL recently found to underlie salinity tolerance in the threespine stickleback. However, most other observed hotspots did not colocate with regions of the genome known to be involved in adaptive divergence between marine and freshwater habitats.Peer reviewe

Avian ecological epigenetics : pitfalls and promises

Epigenetic mechanisms can alter gene expression without a change in the nucleotide sequence and are increasingly recognized as important mechanisms that can generate phenotypic diversity. Most of our current knowledge regarding the origin and role of epigenetic variation comes from research on plants or mammals, often in controlled rearing conditions. Epigenetic research on birds in their natural habitats is still in its infancy, but is needed to answer questions regarding the origin of epigenetic marks and their role in phenotypic variation and evolution. Here we review the potential for studying epigenetic variation in natural bird systems. We aim to provide insights into (1) the origin of epigenetic variation, (2) the relationship between epigenetic variation and trait variation, and (3) the possible role of epigenetic variation in adaptation to changing environments. As there is currently little research on epigenetics in wild birds, we examine how findings on other taxa such as plants and mammals relate to birds. We also examine some of the pros and cons of the most commonly used methods to study patterns of DNA methylation in birds, and suggest some topics we believe need to be addressed to develop the field of wild avian epigenetics further.Peer reviewe

Seasonal Variation in Genome-Wide DNA Methylation Patterns and the Onset of Seasonal Timing of Reproduction in Great Tits

In seasonal environments, timing of reproduction is a trait with important fitness consequences, but we know little about the molecular mechanisms that underlie the variation in this trait. Recently, several studies put forward DNA methylation as a mechanism regulating seasonal timing of reproduction in both plants and animals. To understand the involvement of DNA methylation in seasonal timing of reproduction, it is necessary to examine within-individual temporal changes in DNA methylation, but such studies are very rare. Here, we use a temporal sampling approach to examine changes in DNA methylation throughout the breeding season in female great tits (Parus major) that were artificially selected for early timing of breeding. These females were housed in climate-controlled aviaries and subjected to two contrasting temperature treatments. Reduced representation bisulfite sequencing on red blood cell derived DNA showed genome-wide temporal changes in more than 40,000 out of the 522,643 CpG sites examined. Although most of these changes were relatively small (mean within-individual change of 6%), the sites that showed a temporal and treatment-specific response in DNA methylation are candidate sites of interest for future studies trying to understand the link between DNA methylation patterns and timing of reproduction.Peer reviewe

Temporal changes in DNA methylation and RNA expression in a small song bird: within- and between-tissue comparisons

BackgroundDNA methylation is likely a key mechanism regulating changes in gene transcription in traits that show temporal fluctuations in response to environmental conditions. To understand the transcriptional role of DNA methylation we need simultaneous within-individual assessment of methylation changes and gene expression changes over time. Within-individual repeated sampling of tissues, which are essential for trait expression is, however, unfeasible (e.g. specific brain regions, liver and ovary for reproductive timing). Here, we explore to what extend between-individual changes in DNA methylation in a tissue accessible for repeated sampling (red blood cells (RBCs)) reflect such patterns in a tissue unavailable for repeated sampling (liver) and how these DNA methylation patterns are associated with gene expression in such inaccessible tissues (hypothalamus, ovary and liver). For this, 18 great tit (Parus major) females were sacrificed at three time points (n=6 per time point) throughout the pre-laying and egg-laying period and their blood, hypothalamus, ovary and liver were sampled.ResultsWe simultaneously assessed DNA methylation changes (via reduced representation bisulfite sequencing) and changes in gene expression (via RNA-seq and qPCR) over time. In general, we found a positive correlation between changes in CpG site methylation in RBCs and liver across timepoints. For CpG sites in close proximity to the transcription start site, an increase in RBC methylation over time was associated with a decrease in the expression of the associated gene in the ovary. In contrast, no such association with gene expression was found for CpG site methylation within the gene body or the 10kb up- and downstream regions adjacent to the gene body.ConclusionTemporal changes in DNA methylation are largely tissue-general, indicating that changes in RBC methylation can reflect changes in DNA methylation in other, often less accessible, tissues such as the liver in our case. However, associations between temporal changes in DNA methylation with changes in gene expression are mostly tissue- and genomic location-dependent. The observation that temporal changes in DNA methylation within RBCs can relate to changes in gene expression in less accessible tissues is important for a better understanding of how environmental conditions shape traits that temporally change in expression in wild populations.</div

Fine-tuning of seasonal timing of breeding is regulated downstream in the underlying neuro-endocrine system in a small songbird

The timing of breeding is under selection in wild populations as a result of climate change, and understanding the underlying physiological processes mediating this timing provides insight into the potential rate of adaptation. Current knowledge on this variation in physiology is, however, mostly limited to males. We assessed whether individual differences in the timing of breeding in females are reflected in differences in candidate gene expression and, if so, whether these differences occur in the upstream (hypothalamus) or downstream (ovary and liver) parts of the neuroendocrine system. We used 72 female great tits from two generations of lines artificially selected for early and late egg laying, which were housed in climate-controlled aviaries and went through two breeding cycles within 1 year. In the first breeding season we obtained individual egg-laying dates, while in the second breeding season, using the same individuals, we sampled several tissues at three time points based on the timing of the first breeding attempt. For each tissue, mRNA expression levels were measured using qPCR for a set of candidate genes associated with the timing of reproduction and subsequently analysed for differences between generations, time points and individual timing of breeding. We found differences in gene expression between generations in all tissues, with the most pronounced differences in the hypothalamus. Differences between time points, and early- and late-laying females, were found exclusively in the ovary and liver. Altogether, we show that fine-tuning of the seasonal timing of breeding, and thereby the opportunity for adaptation in the neuroendocrine system, is regulated mostly downstream in the neuro-endocrine system.Peer reviewe

LAPSIPERHEIDEN PALVELUIDEN TOIMIVUUS JA TUNNETTAVUUS POHJOIS-SATAKUNNAN PERUSPALVELUKUNTAYHTYMÄSSÄ

Tämän tutkimuksen tarkoituksena oli selvittää, mitä palveluita Pohjois-Satakunnan peruspalvelukuntayhtymän alueella tarjotaan lapsiperheille, kuinka hyvin kuntayhtymän alueella asuvat perheet tuntevat kuntayhtymän lapsiperheille tarjoamat palvelut ja miten he käyttävät niitä. Lisäksi tavoitteena oli saada kehittämisideoita palveluiden parantamiseksi. Tutkimusongelmina oli selvittää 1) miten PoSa:n lapsiperheille tarjoamat palvelut tunnetaan, 2) kuinka lapsiperheet löytävät ja käyttävät PoSa:n tarjo-amia palveluita ja 3) miten perhepalveluita ja niiden tunnettavuutta voisi kehittää. Tutkimus suoritettiin kyselytutkimuksena vuoden 2015 aikana. Tutkimuksen teoreettisessa osuudessa käsiteltiin kuntayhtymän alueeseen kuuluvia kuntia, peruspalveluita määritteleviä lakeja ja säädöksiä, PoSa:n lapsiperheille tarjoamia palveluita sekä PoSa:n nettisivujen sisältöä. Lisäksi teoreettisessa osuudessa käsiteltiin tutkimuksellisen opinnäytetyön metodiikkaa, kyselytutkimuksen laatimisen ohjeita sekä neuvoja. Kyselytutkimuksen laatimista käsittelevässä osiossa käytiin läpi kyselytutkimuksen rakentamista sekä toteutusta. Tutkimuksessa selvisi kuntalaisten tyytyväisyys palveluiden toimimiseen. Tunnetuimpia palveluita käytettiin kyllä, mutta kuntayhtymän tarjoamat lisäpalvelut olivat kuntalaisille tuntemattomia ja vähän käytettyjä palveluita. Tutkimuksesta kävi ilmi tiettyjen palveluiden käytön vähäisyys ja heikko tunnettavuus. Vastausprosentti palveluiden arvioinnissa jäi matalaksi, mutta annettujen vastausten perusteella palveluiden voidaan katsoa toimivan melko hyvin. Perhepalveluiden toimivuutta voidaan tutkimustulosten perusteella parantaa tuomalla palveluita lähemmäs kuntalaisia sekä lisäämällä työvoimaa erityisesti pienemmissä toimipisteissä. Tutkimustuloksien perusteella pääteltiin, että palveluiden tunnettavuutta voidaan lisätä esimerkiksi esitteiden ja selkeämpien nettisivujen avulla.The purpose of this research was to determine which basic services the healthcare consortium of municipalities in the North-Satakunta region (referred to as PoSa) provides for families with children, how well aware those families are of the availability of these services and how much the families avail themselves of them. Another objective of this research was to find new development and improvement ideas for the services provided. The research problems were to find out the following: 1) How well the services provided to families with children in the PoSa area are known among the target group? 2) In which ways families with children discover and utilize the services offered by the PoSa area? 3) How family services and the awareness of their functions could be developed? The research was conducted as a survey during the year 2015. The theoretical part of the research covers the municipalities that belong to the region, the laws and regulations that define the basic healthcare services, the services offered to families with children within the North-Satakunta region and the contents of the website of the PoSa service area. The methods of a researchbased thesis and instructions and advice for elaborating a survey were also discussed in the theoretical part. The planning and execution phase is elaborated in the part that discusses the planning and carrying out of a survey-based research project. As a result, the satisfaction level of the target group regarding the provided services was determined. The most common services were well utilized and recognized, but many respondents were not aware of lesser known additional services and those services were considerably less used. The research revealed those services that had lower level of use and awareness within the target group. The percentage of answers was moderate, but according to the answers given, the health care services can be considered reasonably successful. According to the survey the functionality of the family services could be improved by bringing the services closer to the citizens of the municipalities and by more personnel especially in the smaller service points and facilities. Also conclusions based on the survey results suggested that awareness of the services provided can be raised for example via handouts and by making their website clearer

Leikki varhaiskasvatuksessa : leikin havainnoinnin sekä leikkiympäristöjen kehittäminen Preitilän päiväkodissa

Tämän kehittämistyön tarkoituksena oli kehittää Preitilän päiväkotiin uudet, ajatuksella mietityt leikkiympäristöt. Kehittämistyön tavoitteena oli kehittää leikkiympäristöjä siten, että leikkiympäristöt ovat toimivat, viihtyisät ja leikkiin kutsuvammat. Lisäksi kehittämistyön tarkoituksena oli luoda varhaiskasvattajien käyttöön uudet leikin havainnointilomakkeet tukemaan leikin havainnointia. Kehittämistyön teoreettinen viitekehys koostuu leikin merkityksestä ja teoriasuuntauksista, leikin kehityksestä lapsen eri ikävaiheissa sekä leikkiympäristöjen teoriaperustasta. Teoreettisessa viitekehyksessä käsitellään myös havainnointia varhaiskasvatuksessa sekä leikin havainnointia. Toteutetut leikkiympäristöjen muutostyöt perustuvat teoriaperustaan leikkiympäristöjen merkityksestä sekä lapsen leikin kehityksestä. Leikin havainnointilomakkeen pohjana toimi teoriatieto leikin havainnoinnista sekä lapsen leikin kehitysvaiheista ja leikin muodoista. Kehittämistyön menetelminä käytettiin dialogista keskustelua, havainnointia sekä kyselylomaketta. Kehittämistyön tuotoksena syntyi Preitilän päiväkotiin kasvattajien ja lasten toiveiden mukaiset uudet leikkiympäristöt sekä kasvattajien käyttöön leikin havainnointilomakkeet 1-3 ja 3-6 vuotiaiden lasten leikin havainnointiin