Thèse (Ph. D.)--Université Laval, 2008. / Bibliogr. Publié aussi en version électronique dans la Collection Mémoires et thèses électroniques.

<p>Trichome formation in Arabidopsis lyrata is a naturally occurring trait with phenotypic polymorphisms within wild populations. In Swedish accessions of A. lyrata, three genetic polymorphisms situated in the coding region of GL1, an important transcription factor in trichome production, have been identified, and these are candidates for being the cause of a glabrous phenotype. In this study a complementation test has been performed to clarify which mutation/mutations that are detrimental for trichome formation. A set of constructs has been transformed into A. thaliana, a close relative to A. lyrata, and subsequent generations of plants were examined for phenotype, genotype and gene expression. A SNP (Single Nucleotide Polymorphism) in the R3 MYB domain of GL1, resulting in a change of an alanine to aspartic acid, was identified as the critical polymorphism. The other two mutations, two indels, were harmless to protein function. The inserted constructs were under control of the native GL1 promoter. Plants that, because of the SNP, lacked trichome production, became totally glabrous.</p>

Abstract Local adaptation is important evolutionary process leading to adaptive population differentiation. Currently, examining its genetic basis is a major goal of evolutionary and ecological genetics. In my thesis I studied local adaptation and its genetic basis in populations of a perennial outcrossing model plant Arabidopsis lyrata by combining common garden experiments at the native field sites and in controlled conditions with quantitative trait locus mapping. Estimates of fitness in the field – both at the level of multiple components as well as hierarchical total fitness – showed that populations of A. lyrata were locally adapted. The studied populations were also phenotypically differentiated in ecologically relevant traits. Different components of fitness were important for the advantage of the locals depending on the environment. Local alleles were associated with high fitness in the field, suggesting that differing directional selection pressures have been involved in phenotypic differentiation. Mostly different genomic regions governed local adaptation in different environments, but the results also suggested that some of these regions could involve rarely documented fitness tradeoffs (antagonistic pleiotropy). Loci governing flowering time differentiation differed between the studied environments, highlighting the need to conduct experiments both in the wild and in controlled conditions. In contrast to most existing studies, F2 hybrids in general had surprisingly high fitness at one study site, largely due to beneficial dominance effects at loci governing survival in that environment. In addition to nuclear genes, cytoplasmic genomes also were found to have a role in local adaptation. / Tiivistelmä Luonnonvalinta saa aikaan paikallista sopeutumista ja adaptiivista erilaistumista. Paikallisen sopeutumisen perinnöllisen taustan selvittäminen on tällä hetkellä yksi tärkeimpiä evolutiivisen ja ekologisen genetiikan tavoitteita. Tässä väitöskirjatyössä tutkin paikallista sopeutumista ja sen geneettistä taustaa monivuotisella, ristipölytteisellä mallikasvilla, idänpitkäpalolla (Arabidopsis lyrata). Käytin työssäni geenikartoitusta kasveilla joita kasvatettiin yhdenmukaisissa oloissa sekä populaatioiden luontaisilla kasvupaikoilla että kontrolloiduissa olosuhteissa. Kenttäolosuhteissa arvioitu kelpoisuus osoitti idänpitkäpalkopopulaatioiden olevan paikallisesti sopeutuneita sekä yksittäisten kelpoisuuteen vaikuttavien ominaisuuksien että hierarkkisen kokonaiskelpoisuuden tasolla. Tutkitut populaatiot olivat myös erilaistuneita ekologisesti tärkeissä ominaisuuksissa. Kelpoisuuteen vaikuttavat ominaisuudet myös poikkesivat ympäristöjen välillä. Paikalliset alleelit olivat yhteydessä korkeaan kelpoisuuteen luonnossa, minkä perusteella voitiin päätellä erisuuntaisen luonnonvalinnan vaikuttaneen populaatioden erilaistumiseen. Kromosomiston eri alueet olivat tärkeitä sopeutumisessa eri ympäristöihin, mutta myös joidenkin samojen genomin alueiden havaittiin mahdollisesti vaikuttavan vastakkaisesti kelpoisuuteen eri ympäristöissä. Myös kukkimisajan erilaistumiseen vaikuttavat genomin alueet poikkesivat eri ympäristöjen välillä erityisesti verrattaessa kenttäkokeita kasvatushuone- ja kasvihuonekokeisiin. Toisin kuin useimmissa tutkimuksissa on havaittu, F2-sukupolven jälkeläistön kelpoisuus oli yllättävän korkea yhdessä kenttäkoeympäristössä. Tähän vaikuttivat kelpoisuuden kannalta suotuisat dominoivat geenivaikutukset, jotka paransivat kasvien selviytymistä kyseisessä ympäristössä. Tumassa sijaitsevien geenien lisäksi myös soluelimien perimällä havaittiin olevan yhteys paikalliseen sopeutumiseen.

Abstract Adaptation to local environment is important for all organisms to guarantee survival and to maximize reproduction. Populations of the same species may live in environments that differ markedly. Due to differential selection pressures this can lead to population differentiation, which can be studied both at the phenotypic and at the gene level. The growing season cued by long days is typically short in the north, whereas southern populations have long growing seasons and are adapted to short days. Seasonal fluctuations in temperature also differ between northern and southern environments. Daylength and temperature regulate the timing of flowering in plants. Environmental regulation of flowering and its genetic basis has been extensively studied in the annual model species Arabidopsis thaliana. The perennial growth and flowering habit has been studied especially in trees, but studies on herbaceous plants species have been lacking. In this thesis, I have studied adaptation to growing season length in a perennial herbaceous model species Arabidopsis lyrata. Individuals from populations adapted to northern and southern environments in Europe were grown in same conditions in the growth chambers and in the field. Differentiation between the populations was studied by observing their flowering phenotypes and by studying the expression of genes that are candidates for governing the phenotypic differentiation. The main result in the thesis was that adaptation to short growing season in north can be seen as long daylength requirement for flowering and as fast developmental rate. Critical daylength for flowering likely regulates especially the timing of flowering cessation in the end of the growing season. Flowering time of individuals from northern populations also responded more strongly to cold treatment (representing winter) than that of the southern population. The cold requirement for flowering guarantees that the plants only flower after the winter in the spring with suitable conditions. Expression studies indicated that population differentiation in flowering could be at least partly governed by the expression variation in a few candidate genes. The results in this thesis are valuable for instance for understanding perennial species in general, including tree and crop species, and for predicting how plants response to changing climate. / Tiivistelmä Ympäröiviin oloihin sopeutuminen on tärkeää kaikille organismeille selviytymisen ja jälkeläistuoton kannalta. Saman lajin eri populaatiot saattavat elää ympäristöissä, joiden olosuhteet poikkeavat toisistaan huomattavasti. Tällöin populaatioihin kohdistuvat erilaiset valintapaineet ja populaatiot erilaistuvat. Erilaistuminen havaitaan tarkastelemalla yksilöiden ilmiasuja ja geenejä. Kasvukauden pituus määrittää eteläisten ja pohjoisten kasvuympäristöjen valintapaineita. Pohjoisessa kasvukausi on lyhyt ja sen alkamisesta ja loppumisesta kertoo pitkä päivänpituus. Etelässä on pitkä kasvukausi ja siellä elävät populaatiot ovat sopeutuneet lyhyeen päivänpituuteen. Myös vuodenaikaiset lämpötilavaihtelut eroavat pohjoisten ja eteläisten alueiden välillä. Muutokset päivänpituudessa ja lämpötilassa säätelevät kasveilla kukkimisen ajankohtaa. Kukkimiseen vaikuttavia ympäristötekijöitä ja kukkimista sääteleviä geenejä on tutkittu paljon yksivuotisella mallilajilla lituruoholla (Arabidopsis thaliana). Monivuotisten kasvien kasvun ja kukkimisen säätelyä on tutkittu etenkin puilla, mutta hyvin vähän ruohovartisilla kasveilla. Tässä väitöskirjatyössä tutkin kasvukauden pituuteen sopeutumista monivuotisella ruohovartisella lajilla, idänpitkäpalolla (Arabidopsis lyrata). Pohjoisiin ja eteläisiin ympäristöoloihin sopeutuneiden eurooppalaisten populaatioiden yksilöitä kasvatettiin samanlaisissa olosuhteissa kontrolloiduissa kasvatushuoneissa ja kenttäolosuhteissa. Populaatioiden erilaistumista tarkkailtiin kukkimiseen liittyvissä ominaisuuksissa sekä eroja selittävien kandidaattigeenien ekspressiossa. Tutkimuksen päätulos oli, että pohjoisen populaation lyhyeen kasvukauteen sopeutuminen voidaan havaita pitkän päivän vaatimuksena kukkimiselle ja nopeana kehityksenä. Kenttäolosuhteissa päivänpituus sääteli etenkin kukkimisen lopetusta kasvukauden lopussa. Pitkä kylmäkäsittely nopeutti kukkimista etenkin pohjoisessa populaatiossa. Kukkimista edeltävä kylmävaatimus takaa, etteivät kasvit kuki syksyllä epäsuotuisissa olosuhteissa, vaan vasta keväällä talven jälkeen. Populaatioiden erilaistuminen kukkimisen päivänpituus- ja kylmävaatimuksessa selittyy todennäköisesti ainakin osittain kandidaattigeenien ekspressioeroilla. Tämän väitöskirjatyön tuloksia voidaan soveltaa monivuotisten viljelykasvien jalostuksessa tai ennustettaessa kasvipopulaatioiden sopeutumista ilmastonmuutokseen.

The relationship between leaf and its environment has resulted in a tremendous diversification of leaf shape within and between plants species, which is important to cope with the differing environmental conditions. Arabidopsis lyrata is a prime model plant that shows leaf shape variation within species and between related species. In Cardamine and Capsella, the RCO genes (homeodomain leucine zipper family transcription factors) are involved in shaping leaves, yielding more complex shaped leaves (leaflets). In A. thaliana, over the course of evolution, the RCO-A and RCO-B paralogous genes have been deleted that led to the loss of lobes (leaf simplification). Based on previous quantitative trait locus (QTL) mapping results, these gene family members are thought to control leaf shape variation also in A. lyrata. Since the paralog involved in leaf shape variation is unknown, both copies of the RCO genes (AL6G13350 and AL6G13360) were sequenced. The study aimed to identify whether DNA sequence variation exists in the two paralogous RCO genes, which could explain the phenotype variation both within population and between A. lyrata populations, along with related species A. arenosa. The results showed limited variation in the coding regions in the form of indels, single nucleotide polymorphisms (SNPs) and amino acid substitutions resulting in no significant difference in phenotype between genotype (p&gt;0.139). The most variants were rare and increasing the number of individuals within the populations, to cover the full phenotypic spectrum, may lead to different results. Not being able to obtain the nucleotide sequence of the promotor region, further analysis is required since it is an important region for gene expression, which could explain phenotype variation for leaf shape in Arabidopsis lyrata.

Trichome formation in Arabidopsis lyrata is a naturally occurring trait with phenotypic polymorphisms within wild populations. In Swedish accessions of A. lyrata, three genetic polymorphisms situated in the coding region of GL1, an important transcription factor in trichome production, have been identified, and these are candidates for being the cause of a glabrous phenotype. In this study a complementation test has been performed to clarify which mutation/mutations that are detrimental for trichome formation. A set of constructs has been transformed into A. thaliana, a close relative to A. lyrata, and subsequent generations of plants were examined for phenotype, genotype and gene expression. A SNP (Single Nucleotide Polymorphism) in the R3 MYB domain of GL1, resulting in a change of an alanine to aspartic acid, was identified as the critical polymorphism. The other two mutations, two indels, were harmless to protein function. The inserted constructs were under control of the native GL1 promoter. Plants that, because of the SNP, lacked trichome production, became totally glabrous.

Les travaux effectués dans le cadre de cette thèse ont permis de documenter un nouveau système modèle par la caractérisation et l'utilisation à.''Arabidopsis lyrata, une espèce apparentée à la plante modèle Arabidopsis thaliana. L'intérêt de ce nouveau système modèle tient au fait qu'il pourra contribuer à la compréhension de l'évolution des génomes et de la spéciation chez les plantes. Dans cette étude, la stratégie d'un croisement pseudotestcross réciproque {A. lyrata ssp. lyrata x A. lyrata ssp. petraea) a été employée afin de construire une carte génétique pour chacune des sous-espèces. Les marqueurs AFLP se sont avérés efficaces pour révéler la couverture des cartes tfA. lyrata mais ont surtout permis d'augmenter la densité des marqueurs sur les groupes de liaisons. Ensuite, nous avons développé un système alternatif pour l'étude de la polyploïdie chez Arabidopsis. Un processus en deux étapes comprenant un croisement A. thaliana x A. lyrata ssp. petraea suivi d'un événement spontané de doublement chromosomique a permis d'obtenir des allopolyploïdes. Plusieurs phénomènes observés chez ces allopolyploïdes suggèrent la présence d'un choc génomique. Des changements génétiques et épigénétiques ont été décrits avec ce système modèle. A. lyrata possède un ensemble de caractéristiques intéressantes pour devenir un nouvel outil de travail en génétique moléculaire des plantes. / The work carried out within this thesis resulted in the description of a novel model system by characterizing and using Arabidopsis lyrata, a close relative species of the model plant Arabidopsis thaliana. This new model system could contribute to a better understanding of genome evolution and speciation in plants. In this study, a 2-way pseudo-testcross strategy {A. lyrata subsp. lyrata x A. lyrata subsp. petraea) was used to construct a genetic map for each subspecies. The AFLP markers proved to be effective to reveal the coverage of A. lyrata maps, but mostly increased marker density on the linkage groups. Then, we developed an alternative system for the study of polyploidy in Arabidopsis. A two-step process involving the cross A. thaliana x A. lyrata subsp. petraea followed by a spontaneous chromosomal doubling generated allopolyploids. Several phenomena observed with these allopolyploids suggest the presence of genomic shock. Genetic and epigenetic changes were described with this model system. A. lyrata has several interesting characteristics to become a new working tool in plant molecular genetics.

<p>In this thesis, I combined comparative and experimental approaches to examine selection on reproductive traits, and population differentiation in the insect-pollinated, outcrossing, perennial herb <i>Arabidopsis lyrata</i>. More specifically, I (1) determined whether selection on flowering phenology and floral display can be attributed to interactions with pollinators and herbivores, (2) examined whether population differentiation in flowering phenology and floral display is correlated with current selection on these traits, and (3) tested for local adaptation from contrasting environments in Europe.</p><p>A field experiment conducted in a Swedish population demonstrated, that interactions with pollinators may markedly affect selection on both floral display and phenology of flowering. In an alpine population in Norway, grazing damage to inflorescences strongly influenced selection on floral display. The results suggest that variation in the abundance of pollinators and herbivores should contribute to spatio-temporal variation in selection on flowering phenology and floral display in <i>A. lyrata</i>. </p><p>A common-garden experiment showed that flowering phenology and floral display vary among Scandinavian populations of <i>A. lyrata</i>. For some traits patterns of population differentiation were consistent with differences in the direction and strength of phenotypic selection determined in comparisons (a) between an alpine population in Norway and a coastal population in Sweden, and (b) among coastal populations in Sweden. This suggests that current selection contributes to the maintenance of genetic differentiation in these traits.</p><p>Adaptive differentiation among populations was examined in a reciprocal transplant experiment that included populations from three contrasting environments, alpine Norway, coastal Sweden and lowland, continental Germany. The experiment provided evidence for local adaptation, and indicated that populations have diverged in traits affecting plant establishment and early growth.</p>

In this thesis, I combined comparative and experimental approaches to examine selection on reproductive traits, and population differentiation in the insect-pollinated, outcrossing, perennial herb Arabidopsis lyrata. More specifically, I (1) determined whether selection on flowering phenology and floral display can be attributed to interactions with pollinators and herbivores, (2) examined whether population differentiation in flowering phenology and floral display is correlated with current selection on these traits, and (3) tested for local adaptation from contrasting environments in Europe. A field experiment conducted in a Swedish population demonstrated, that interactions with pollinators may markedly affect selection on both floral display and phenology of flowering. In an alpine population in Norway, grazing damage to inflorescences strongly influenced selection on floral display. The results suggest that variation in the abundance of pollinators and herbivores should contribute to spatio-temporal variation in selection on flowering phenology and floral display in A. lyrata. A common-garden experiment showed that flowering phenology and floral display vary among Scandinavian populations of A. lyrata. For some traits patterns of population differentiation were consistent with differences in the direction and strength of phenotypic selection determined in comparisons (a) between an alpine population in Norway and a coastal population in Sweden, and (b) among coastal populations in Sweden. This suggests that current selection contributes to the maintenance of genetic differentiation in these traits. Adaptive differentiation among populations was examined in a reciprocal transplant experiment that included populations from three contrasting environments, alpine Norway, coastal Sweden and lowland, continental Germany. The experiment provided evidence for local adaptation, and indicated that populations have diverged in traits affecting plant establishment and early growth.

Abstract Plants and animals colonized Northern Europe after the last Ice Age from different refugia, not covered by the ice sheet. Many plants, such as the northern rock cress (Arabidopsis lyrata ssp. petraea) adapted to the short growing season in the North. We thus expect that colonization of the new environment was accompanied by directional selection for traits conferring this adaptation. In this thesis I studied whether recent directional selection can be detected in two important genes, PHYTOCHROME A (PHYA) and FLOWERING LOCUS C1 (FLC1), related to the flowering time pathway. To detect directional selection, I compared DNA sequence variation from the samples of a southern (Plech, Germany) and a northern (Spiterstulen, Norway) population. I also studied the current response potential to changing conditions in the marginal Spiterstulen population. Adaptation potential was characterized by assessing plasticity and amount of additive genetic variation, focusing on flowering traits. In addition, associations of 21 flowering time candidate genes for phenological and fitness traits were studied. There were several lines of evidence for recent directional selection in both candidate genes, PHYA and FLC1, in the northern Spiterstulen population Variation was strongly reduced around both genes and in addition they were highly differentiated between populations. In the Spiterstulen population there was a remarkable reduction in additive genetic variation for flowering traits, for instance when compared with morphological traits. On the other hand, phenological traits showed high plasticity. Some of the photoperiodic pathway genes showed association to flowering or reproductive fitness. The results suggest that directional selection during the colonization of the northern areas has impacted the two studied genes. Genetic changes were likely involved in altered photoperiodic and vernalization responses which might be adaptive for a short growing season. Further, directional selection was probably responsible for reducing additive genetic variation in flowering traits. Because there was only little genetic variation, adaptation to future environmental change of the marginal Spiterstulen population is likely to rely largely on plastic reactions to environmental signals, or tracking the environment by dispersal. / Tiivistelmä Kasvit ja eläimet levittäytyivät Pohjois-Eurooppaan viimeisen jääkauden jälkeen mannerjäätikön ulkopuolella jääneistä refugioista. Useat kasvit, kuten idänpitkäpalko (Arabidopsis lyrata ssp. petraea) sopeutuivat pohjoisen lyhyeen kasvukauteen. On syytä olettaa, että suuntaava valinta vaikutti sopeutumisessa tärkeisiin ominaisuuksiin. Tässä väitöskirjassa tutkin voidaanko suuntaavan valinnan aiheuttamia jalanjälkiä löytää kahdesta tärkeästä kukkimisaikageenistä, FYTOKROMI A (PHYA) ja FLOWERING LOCUS C1 (FLC1) geeneistä. Tätä varten vertasin DNA sekvenssimuuntelua pohjoisessa (Norja) ja eteläisessä (Saksa) populaatiossa, kiinnittäen erityisesti huomiota geneettisen muuntelun määrään ja erilaistumiseen. Lisäksi tutkin miten Spiterstulenin reunapopulaatio voi vastata tulevaisuudessa muuttuvaan ympäristöön. Sopeutumispotentiaalia arvioitiin sekä fenotyyppisen plastisuuden että additiivisen geneettisen muuntelun määrällä. Lisäksi tutkin miten vaihtelu 21 kukkimisaikageenissä liittyy fenologisiin ja kelpoisuusominaisuuksiin. Useat merkit viittasivat siihen, että suuntaava valinta oli vaikuttanut kummassakin tutkitussa geenissä. Muuntelu oli vähentynyt voimakkaasti kumpaakin geeniä ympäröiviltä kromosomialueilta, jotka olivat myös selkeästi erilaistuneet. Additiivinen geneettinen muuntelu oli selvästi vähentynyt kukkimisominaisuuksissa verrattuna morfologisiin ominaisuuksiin, mahdollisesti suuntaavan valinnan johdosta. Kukkimisominaisuudet olivat kuitenkin plastisia. Jotkin valojaksoreitin geenit vaikuttivat sekä kukkimiseen että lisääntymiskykyyn. Nämä tulokset osoittavat että suuntaava valinta vaikutti kahteen tutkittuun geeniin pohjoiseen levittäytymisen aikana. Geneettiset muutokset liittyivät todennäköisesti muuttuneisiin valojakso-, ja vernalisaatiovasteisiin, jotka saattoivat edistää sopeutumista lyhyeen kasvukauteen. Koska geneettistä muuntelua oli vain hyvin vähän, fenotyyppisellä plastisuudella on todennäköisesti tärkeä rooli sopeutumisessa muuttuvaan ympäristöön Spiterstulenin reunapopulaatiossa.