Regulation of host health is intricately coordinated by a diverse interplay of immune cells detecting assaults from pathogens via recognition of pathogen associated molecular patterns (PAMPs) to mount an immune response, as well as detecting damage associated molecular patterns (DAMP) to indicate an area of damage and signal tissue repair. The gastrointestinal tract is a major signaling hub for such immune responses, as intestinal epithelial cells (IECs) compose the epithelial barrier, immune cells surveillance breached barriers to regulate the gut microbiome, and intestinal stem cells (ISCs) proliferate to replenish the IEC pool. One such method for regulating these cellular functions downstream of PAMPs/DAMPs within the gastrointestinal tract is via NF-κB signaling. This cellular signaling pathway is activated by one of two pathways: the well- defined canonical NF-κB pathway and the understudied noncanonical NF-κB pathway. The noncanonical NF-κB pathway is unique as it requires NIK, the NF-κB-inducing Kinase, to further elicit signal transduction of this pathway. Noncanonical NF-κB activation is critical to maintaining gut health, as signaling is regulated at a precise level to ensure a balance of pro-/anti-inflammatory signals to elicit a proper damage response. Any perturbations to NIK-activated signaling significantly predisposes the gastrointestinal niche towards chronic inflammatory conditions of the gastrointestinal tract. In this work, we explore the potential involvement of dysregulated noncanonical NF-κB signaling in inducing chronic inflammatory diseases of the gut, including Eosinophilic Esophagitis (upper GI tract), Celiac Disease/Non-Celiac Gluten Sensitivities (small intestine), Inflammatory Bowel Disease (entire intestine/large intestine), and an inflammatory subtype of colorectal cancer being Colitis-Associated Colorectal Cancer (large intestine). We study this pathway via the use of murine models bearing genetic deletions, cellular models, and the generation of miniature organs (i.e. "organoids") in petri dishes. Further, we assess varying levels of NF-κB signaling through the genetic deletions of NIK and RelA to inhibit noncanonical and canonical NF-κB pathways, respectively. Reciprocally, we also examine overactivated signaling via loss of the negative regulatory NLRs, which are proteins that function to impede NF-κB signaling. Clinical relevancy of this work is evaluated using biopsy samples collected from human patients with active disease states. Culminating our work, we find that noncanonical NF-κB signaling levels is both tissue- and cell-type specific in driving disease formation. Finally, we conclude our findings by suggesting the promise of NIK as a potential candidate for disease biomarkers and a target for future drug development. / Doctor of Philosophy / Redness, swelling, heat, pain, and loss of function – these are the five signs of inflammation. Under normal physiological conditions, inflammation is the body's conserved evolutionary response by serving as the first line of defense against infections propagated by foreign invaders like pathogens (i.e. bacteria, viruses, fungi), while also signaling to the immune system to resolve tissue damage. Therefore, properly maintained pro-inflammatory signaling is critical to ensuring a healthy state. However, an imbalance in pro- and anti-inflammatory signaling elicits a long-term, low-grade form of inflammation termed "chronic inflammation". Unresolved chronic inflammation can persist for several months or even years and further predisposes patients to various chronic inflammatory conditions and even inflammation-induced cancer. The NF-κB cellular signaling mechanism is a central regulator of inflammation and can be activated upon either the canonical NF-κB or noncanonical NF-κB pathways. In comparison to its canonical counterpart, the noncanonical NF-κB is vastly understudied, especially in regards to gastrointestinal health. A unique feature of the noncanonical NF-κB pathway is the required stabilization of the NF-κB-inducing Kinase (NIK) protein, which is required for further propagation of this signaling network. As evidenced by our culmination of works, we reveal that Noncanonical NF-κB signaling is critical to gut health, as it maintains a precise cellular signaling mechanism within the gut tract by properly maintaining pro- and anti-inflammatory signaling. Additional, downstream implications include regulation of cell division and activation of cell death to elicit a proper damage response. Within this dissertation, we evaluate the understudied noncanonical NF-κB pathway in various chronic inflammatory diseases of the gut including Eosinophilic Esophagitis (upper GI tract), Celiac Disease/Non-Celiac Gluten Sensitivities (small intestine), Inflammatory Bowel Disease (entire intestine/large intestine), and an inflammatory subtype of colorectal cancer Colitis-Associated Colorectal Cancer (large intestine). Through the use of murine models bearing deletions of genes related to noncanonical NF-κB signaling (esp. NIK), cell models, and the generation of "mini-organs" organoids from isolated intestinal stem cells, we are able to model the involvement of NIK and noncanonical NF-κB signaling in maintaining gastrointestinal health. Clinical relevancy of these findings was further evaluated by quantifying noncanonical NF-κB signaling levels in human biopsies. Culminating our work, we find noncanonical NF-κB signaling to be context-specific in driving disease formation. Finally, we conclude this work by suggesting the promise of NIK as a potential candidate for disease biomarkers and target for future drug development.

Made available in DSpace on 2015-03-26T13:36:45Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2928675 bytes, checksum: b8457e10b48f36e111d09ef311ae3d5a (MD5) Previous issue date: 2010-02-18 / The begomovirus NSP (nuclear shuttle protein) facilitates the transport of viral DNA from the nucleus to the cytoplasm and cooperates with the movement protein MP to promote the translocation of viral DNA to the adjacent, uninfected cells through plasmodesmata. NSP interacts with members of the LRR-RLK ( leucine-rich repeat receptor like kinase ) family, designated NIKs ( NSP-Interacting Kinase ). Binding of NSP to the activation loop of NIK inhibits kinase activity and hence the viral protein suppresses receptor autophosphorylation and defense responses. Mutagenesis assays in the activation loop of NIK have demonstrated that the threonine 474 residue is phosphorylated in vitro and plays a crucial role in the kinase activity that is required for signaling. Replacement of Thr-474 with aspartate produces the T474 mutant, which exhibits constitutive activation, enhanced substrate phosphorylation activity and less inhibitory effect by NSP binding. The goal of this investigation was to analyse the NIK kinase domain in defense responses against begomovirus in tomato. The NIK mutant T474D cDNA was placed under the control of 35S promoter into a binary vector for plant transformation (35S-AtNIK-T474D). Primary transformants were selected by PCR and the expression of the transgene was confirmed by normal and quantitative RT- PCR in independently transformed lines. NIK and NIK-T474D overexpression in tomato plants affected the overall developmental performance of transgenic lines, which display elongated stems and a root system less developed. These phenotypes were consistent with a cross-communication between the NIK-mediated antiviral signaling and developmental signaling pathways. Infectivity assays were carried out in AtNIK- and AtNIK-T474D-overexpressing lines, with the virus ToYSV-[MG-Bi2]. Overexpression of super active AtNIK-T474D altered the infection rate by ToYSV, and interfered in symptom development. As compared to untransformed plants and NIK- overexpressing 35S-AtNIK1-6 transgenic lines, independent transgenic AtNIK-T474D lines displayed lower infection rate and attenuated symptoms. These results confirmed in planta the essential role for phosphorylation of the Thr-474 residue for NIK function and underlined the possibility for the development of more efficient tolerance strategies against geminiviruses. / A proteína NSP de begomovírus facilita o transporte do DNA viral do núcleo para o citoplasma e coopera com a proteína de movimento MP para promover o transporte do DNA viral às células adjacentes não infectadas através dos plasmodesmas. A proteína NSP interage com membros da família LRR-RLK ( leucine- rich repeat receptor like kinase ), designados NIK ( NSP-Interacting Kinase ). A ligação de NSP na alça de ativação de NIK inibe a atividade quinase, e conseqüentemente, a proteína viral inibe a atividade de autofosforilação desses receptores e sua atividade de defesa antiviral. Estudos de mutagênese na alça de ativação de NIK demonstraram que o resíduo Treonina 474 é fosforilado in vitro e exerce uma função crucial para atividade de quinase que é requerida para sinalização antiviral. Mutação no resíduo de Thr-474 para aspartato resulta no mutante T474D que exibe ativação constitutiva, atividade de fosforilaçao do substrato aumentada e menor efeito inibidor de NSP. Este trabalho teve como objetivo caracterizar o domínio quinase de NIK na resposta de defesa antiviral em tomateiros. Tomateiros foram transformados com a construção que codifica para NIK super ativa (35S-AtNIK-T474D). Os transformantes primários foram selecionados por PCR e a expressão do transgene em linhagens independentes foi confirmada por RT-PCR normal e em tempo real. A super expressão da NIK1 e NIK- T474D super ativa em tomateiros promoveu um alongamento de entrenós, mas afetou negativamente o desenvolvimento do sistema radicular, demonstrando uma possível comunicação cruzada entre a via de sinalização antiviral mediada por NIK e vias de sinalização de desenvolvimento. Experimentos de infectividade foram conduzidos em linhagens transgênicas superexpressando AtNIK ou AtNIK-T474D, utilizando o vírus ToYSV-[MG-Bi2]. Super expressão de NIK super ativa alterou a taxa de infecção por ToYSV e interferiu no desenvolvimento dos sintomas. Comparado com as plantas não transformadas e a linhagem transgênica 35S-AtNIK1-6 superexpressando NIK normal, a taxa de infecção foi inferior e os sintomas mais atenuados em linhagens transgênicas independentes superexpressando AtNIK-T474D. Estes resultados confirmam in planta o papel essencial da fosforilação do resíduo de Treonina 474 de NIK e indicam a possibilidade de se desenvolverem estratégias de tolerância a geminivirus mais eficientes.

Made available in DSpace on 2015-03-26T13:36:35Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2085482 bytes, checksum: 22303ee6597c676fbc153930781c403e (MD5) Previous issue date: 2007-09-28 / Geminivirus are plant virus packed as geminate semi- icosahedral particles and single-stranded circular DNA genomes. They can be either mono or bipartite and in the bipartite geminivirus, the genomic components are designated DNA-A and DNA-B. DNA-A encodes the proteins needed for replication, transactivation and encapsidation of the viral genome. DNA-B encodes two movement proteins, Movement Protein, MP, and Nuclear Shuttle Protein, NSP, both required for the establishment of a systemic infection. NSP transports the viral DNA between the nucleus and cytoplasm and acts cooperatively with MP to move the viral DNA cell-to-cell across the wall. The geminivirus are highly dependent on the host factors for proliferation and spread. NSP interacts with a plasma membrane receptor kinase, designated NIK (NSP- interacting kinase), from tomato, soybean and Arabidopsis. It has been demonstrated previously that inactivation of NIK gene enhances the susceptibility of the mutant to geminivirus infection. To elucidate the possible role of NIK in the protection mechanism to geminivirus infection, we investigated here the effects of NIK overexpression in the tomato response to geminivirus infection. The NIK gene from Arabidopsis thaliana was used to transform tomato. The primary transformants were confirmed by PCR, selected by RT-PCR according to the transgene expression, in vitro replicated and used for developmental analysis in vitro and in green house. NIK overexpression affected the overall developmental performance of transgenic lines which display delayed and stunted growth, with lower stem, less developed root and shoot systems and a leaf curly phenotype. These transgenic line phenotypes overexpression NIK were similar to that displayed by inhibition of Brassinosteroid synthesis or signaling. Infectivity assays in tomato with ToYSV-[MG-Bi2] revealed that overexpression of NIK caused attenuation of symptom, as the transgenic lines developed less severe symptoms in comparison with the untransformed lines. These results are consistent with a protective role of NIK against viral infection and suggest that the NIK-mediated signaling communicates to some level with the brassinosteroid signaling. / Geminivírus são vírus de plantas com partículas semi- icosaédricas geminadas e genoma de DNA circular de fita simples, podendo ser mono ou bissegmentados, sendo, no último caso, os dois componentes genômicos, denominados DNA-A e DNA-B. No DNA-A, encontram-se os genes que codificam as proteínas necessárias para a replicação viral e encapsidação. O DNA-B codifica duas proteínas de movimento, Movement Protein, MP e Nuclear Shuttle Protein, NSP, ambas requeridas para o estabelecimento de uma infecção sistêmica. NSP transporta o DNA viral entre o núcleo e o citoplasma e atua cooperativamente com MP para transportar o DNA viral de célula-a-célula através da parede celular. Os geminivírus são altamente dependentes de fatores do hospedeiro para sua proliferação. A proteína NSP interage com uma quinase receptora da membrana plasmática, designada NIK (NSP-Interacting Kinase) de tomate, soja e Arabidopsis. Foi demonstrado, em estudos anteriores, que a inativação do gene NIK aumenta a suscetibilidade dos mutantes à infecção viral. Visando elucidar o possível papel de NIK no mecanismo de proteção à infecção por geminivírus, este estudo propôs uma avaliação dos efeitos da superexpressão de NIK na resposta de tomate à infecção por geminivírus. Assim sendo, o gene NIK de Arabidopsis thaliana foi utilizado para transformar tomateiro. Os transformantes primários foram confirmados por PCR, selecionados por RT- PCR, de acordo com a expressão do transgene, repicados in vitro e utilizados em experimentos de análise de desenvolvimento in vitro e em casa-de-vegetação. Análises fenotípicas demonstraram que a superexpressão da proteína promoveu retardo de crescimento dos transformantes, os quais apresentaram baixa estatura, menor comprimento caulinar, menor desenvolvimento de sistema radicular e aéreo, folhas enrugadas/encarquilhadas. O fenótipo observado nas linhagens transgênicas superexpressando NIK é muito similar àquele resultante da inibição da via de biossíntese ou sinalização de brassinosteróides. Os ensaios de infectividade em tomateiro com o vírus ToYSV-[MG-Bi2] revelaram que a superexpresão de NIK causou atenuação dos sintomas uma vez que os transformantes apresentaram menor severidade de sintomas em relação ás linhagens não transformadas. Estes resultados são consistentes com o papel protetor de NIK contra a infecção viral e sugere que a sinalização mediada por NIK comunica-se, em algum nível, com a via de sinalização de brassinosteróides.

Tumor-necrosis factor (TNF)-receptor (R) associated factor 3 (TRAF3) is an important adaptor protein that plays a variety of context-dependent regulatory roles in all types of immune cells. In B cells, TRAF3 mediates signaling downstream of CD40, B cell activating factor (BAFF)-R, and toll-like receptors (TLR)s to restrain B cell survival and function. Downstream of CD40 and BAFF-R, TRAF3 negatively regulates NF-κB2 activation through NF-κB inducing kinase (NIK) stabilization. NF-κB2 activation is important for B cell-homeostatic survival. However, the constitutively active NF-κB2 in other TRAF3 deficient immune cell types does not lead to increased cell survival. More importantly, loss-of-function mutations of the TRAF3 gene are found at relatively high frequencies in B cell malignancies such as multiple myeloma and B cell lymphoma. Therefore, TRAF3 plays a critical and unique role in B cells to restrain cell survival and differentiation that contributes to B cell malignancies. In this study, we aim to identify TRAF3 modulated survival pathways that contribute to homeostatic B-cell survival and B-cell differentiation. We found that TRAF3 degradation was not sufficient or necessary to induce NF-κB2 activation. We also showed that TRAF3 degradation is dependent on association with TRAF2 and cytoplasmic tail of CD40 or BAFF-R. TRAF3 regulation of NIK is important for mature B cell development; however, NIK only partially contributes to TRAF3-mediated B cell survival. TRAF3 also regulates the protein level of proviral integrations of Moloney virus (Pim2), a pro-survival serine/threonine protein kinase encoded by the Pim2 gene, to restrain B cell survival; this regulation can operate independently of the NF-κB2 pathway. Furthermore, we showed that TRAF3 negatively regulates IL-6R signaling, a pathway that contributes to expansion of the plasma cell compartment and to the pathogenesis of multiple myeloma, a plasma cell malignancy. We found that TRAF3 facilitates recruitment of PTPN22, a tyrosine phosphatase, to associate with Jak1 following IL-6 binding to the IL-6R complex. This regulation by TRAF3 restrains plasma cell differentiation, and also provides the first demonstration that PTPN22 regulates cytokine receptor signaling. Collectively, these findings highlight the importance of TRAF3 in the regulation of B cell-specific survival and differentiation pathways. This information could be exploited for more precise and effective therapeutic choices in treatment of B cell malignancies with TRAF3 deficiencies.

In 2001, Argentina suffered an economic collapse that almost wiped out the middle-class, leaving thousands homeless. This resulted in the growth of the informal settlements and informal sectors of employment in and around the formal city of Buenos Aires, the “Capital Federal.” The urgency for survival led to the rise of many waste-pickers or, as they became known the cartoneros. Today they are the primary recycling program in the city with many cooperatives helping their movement. The city has enforced a ‘zero waste’ policy for 2020 in dealing with their serious trash problem and the cartoneros will play a large role in this transformation. This thesis aims to address two complex social issues (disparity and waste) with a long-term architectural initiative which hopes to ultimately blur the social boundaries that exist in the “Paris of the South” via the cartoneros.

Le genre Entérovirus (EV) (famille des picornaviridae) est composé de petits virus à ARN non enveloppées classés en 12 espèces dont 7 sont pathogènes pour l'homme : 4 espèces (A-D) d'enterovirus humains (HEV) et trois espèces (A-C) de rhinovirus humains (HRV). Dans le genre enterovirus, les HRV et HEV sont reconnus comme des pathogènes respiratoires fréquemment responsables d'infections des voies aériennes supérieures et inférieures chez l'enfant et l'adulte. Entre 2009 et 2012, de nouveaux génotypes d'HEV à tropisme respiratoire (HEV-68, 104, 109, et le CVA-21) ont été décrits dans des cas isolés ou épidémiques démontrant la capacité des espèces A à D à induire des infections respiratoires basses humaines.La première phase de ce travail de thèse a eu pour objectifs de préciser le rôle étiologique des infections à EVs; d'identifier les génotypes potentiellement responsables des pathologies respiratoires pédiatriques nécessitant une hospitalisation, mais aussi d'analyser et de comparer les caractéristiques cliniques et épidémiologiques entre les différents groupes de génotypes identifiés. Nous avons réalisé une étude rétrospective sur une cohorte de 309 enfants hospitalisés au CHU de Reims entre septembre 2009 et juin 2010 pour une infection respiratoire aiguë non documentée microbiologiquement par la réalisation des tests virologiques et bactériologiques conventionnels. Nos résultats montrent que le génome des EVs (HEV et HRV) est retrouvé dans 60,5% (187/309) des aspirations naso-pharyngées des enfants hospitalisés, distinguant 15 infections à HEV (dont 10 souches HEV-68) et 172 à HRV. Les cas de bronchiolite et d'exacerbation de l'asthme (133/187) positifs pour la détection des souches HEV (12/133) étaient plus âgés (P=0,003) et plus fréquemment associés avec une détresse respiratoire (P=0,01) et un besoin en oxygénothérapie au moment de leur hospitalisation (P=0,01) que les cas infectés par un HRV. De plus, nous avons mis en évidence pour la première fois en France la circulation épidémique de souches d'HEV-68 (10/15 des souches d'HEV détectées) isolées au cours de l'automne 2009 chez des enfants hospitalisés pour une infection respiratoire aiguë. Nos résultats fournissent de nouvelles informations sur ce génotype ré-émergent qui semble présenter un tropisme respiratoire spécifique des voies respiratoires inférieures.La seconde phase de ce travail de thèse s'est intéressée à étudier les mécanismes liés au développent des processus inflammatoires de la muqueuse au cours de l'infection des voies respiratoires basses par HEV. A l'aide d'un modèle in vitro de cellules respiratoires humaines (A549) infectées par HEV-B (CVB5, Mitchell), nous avons observé que l'infection réplicative des HEV dans les cellules A549 induisait une augmentation dose et temps-dépendante des ARNm, et des protéines IL-8, MCP-1 et RANTES.En conclusion, nos résultats obtenus à partir de prélèvements respiratoires dans le cadre de notre étude de cohorte suggèrent que les EVs représentent une cause étiologique fréquente d'infections respiratoires basses chez l'enfant avec une pathogénicité supérieure des HEVs (principalement dans notre étude HEV-68) par rapport aux souches HRVs. De plus, nos résultats obtenus à partir d'expérimentation in vitro démontrent que les HEVs du groupe B sont capables d'induire au cours de l'infection des cellules épithéliales alvéolaires humaines (A459) une sécrétion spécifique d'IL-8, MCP-1 et RANTES. La production de ces chimiokines correspond à une réponse innée de la cellule épithéliale humaine infectée par les HEVs: nous avons montré pour la première fois que ce mécanisme était en partie régulé par l'activation de la voie non canonique de NF-kB via la protéine NIK dans la cellule épithéliale respiratoire humaine. / The Enterovirus (EV) genus (picornaviridae family) consists of small non-enveloped positive RNA viruses classified in 12 species of which 7 are pathogenic for humans: four species (A-D) of human enterovirus (HEV) and three species (A-C) of human rhinovirus (HRV). Among the EV genus, HEV and HRV are recognized as leading causes of acute respiratory tract infections (ARTIs) in human. Between 2009 and 2012, new HEV respiratory genotypes (e. g. HEV-68, 104, 109, 117 and CVA-21) have been described in isolated cases or outbreaks supporting the ability HEV species A to D to induce lower respiratory tract infections. This supports the hypothesis of an underestimation of the prevalence and etiological role of EVs in pediatric acute respiratory tract infections (ARTIs) (more specifically bronchitis, bronchiolitis and asthma exacerbation).To assess the etiological role and the clinical characteristics of HRV and HEV infections in pediatric patients hospitalized for ARTIs, we conducted a retrospective study of 309 hospitalized pediatric patients in University Hospital Centre of Reims with microbiologically unexplained ARTIs from September 2009 to June 2010. Among the 309 ARTIs, 15 HEV and 172 HRV strains were identified. Among bronchiolitis and asthma exacerbation cases (n=133), HEV infected cases were older (P=0.003) and were more frequently associated with a respiratory distress (P=0.01) and a need for oxygen therapy at the time of admission (P=0.01) than cases infected by HRV strains. Interestingly, during this retrospective study, we provided evidence that during the fall 2009 in France, HEV-D68 strains were responsible for a low proportion of pediatric cases hospitalized for acute airway diseases including bronchiolitis and asthma exacerbation.To identify the mechanisms that can regulate the development of airway mucosa inflammation during HEV respiratory lower tract infections, we investigated the production of chemokines by HEV infected human alveolar epithelial cells (A549). Using in vitro model A549 cells infected by HEV-B (CVB5, Mitchell), we demonstrated that HEV-B strains isolated from upper respiratory tract of child with bronchiolitis could actively replicate in various human airway epithelial cells, and that this replicative infection induced specific dose and time-dependent increases in mRNA and protein secretion of IL-8, MCP-1 and RANTES, but not of all other CC and CXC human chemokines tested. The protein secretion of these chemokines appeared to be significantly increased at 48 and 72 hours post-infection in culture treated by low-doses of IFN-γ in comparison with mock-infected cells (P <0.001), and was correlated to the viral replication activity. In second time, we explore the pathogenic mechanisms that can regulate inflammatory responses to HEV in lower respiratory airways. We show that HEV infection induced a time-dependent increase of NIK protein accumulation that peaks at 16 hours post-infection (H P-I). NIK protein accumulation mediated the processing of p100 in p52, which association with Rel B was evidenced in nuclear compartment between 16 and 48 H P-I.In conclusion, our findings indicate that EVs are a common cause of lower respiratory tract infections in pediatric patients with a potential higher pathogenicity of HEV strains (mainly HEV-68) by comparison to HRV strains. Moreover, our in vitro results demonstrated that HEV are capable to induce the release of specific chemokines (IL -8, MCP-1 and RANTES) by alveolar epithelial cells during a replicative infection. Finally, we demonstrated for the first time that this innate airway epithelial cell response against HEV infection was partly regulated by the activation of the non-canonical NF-kB via NIK protein.

Polyploidization (whole-genome duplication) is widely considered one of the most important evolutionary forces driving the diversification of flowering plants. Polyploids tend to originate recurrently and many plant species retain individuals of two or more different ploidy levels in certain parts of their distributional range of even within their populations. The main aim of this thesis was to address the understudied aspects of polyploid speciation by employing new, convenient methods and/or studying plant model systems with unique features. Difference in monoploid genome size of Tripleurospermum inodorum (Asteraceae) cytotypes provided a unique opportunity for addressing the rate of spontaneous polyploidization in natural populations by enabling the easy distinction of neopolyploid mutants from long-established polyploids in routine flow-cytometric analyses. Repeated ploidy screening in mixed-ploidy populations of annual T. inodorum have been, to our knowledge, the very first attempt to document temporal changes in cytotype composition in situ. In spite of considerable between- year oscillations in cytotype frequencies, both diploids and tetraploids usually persisted locally for several consecutive years. The common incidence of such ploidy mixtures along with a partial fertility of triploid...

Indiana University-Purdue University Indianapolis (IUPUI)

Evolutionary algorithms belongs among the youngest and the most progressive methods of solving difficult optimization tasks. They received huge popularity mainly due to good experimental results in optimization, a simplicity of the implementation and a high modularity, which is an ability to be modified for different problems. Among the most frequently used Evolutionary algorithms belongs Genetic Algorithm, Differential Evolution and Evolutionary Strategy. It is able to apply these algorithms and theirs variants to both continuous, discrete and mixed optimization tasks. A subject of this theses is to compare three main types of algorithms on the catalyst optimization task with mixed variables, linear constraints and experimentally evaluated fitness function.

Adaptive population shifts in response to climate change Ing. Michaela Horníková, Doctoral thesis Abstract Understanding of species' reactions to past climate and environmental changes is a hot topic in many fields of biology as it is relevant also for addressing species' future under the contemporary climate change. Using an emerging model species, the bank vole, I combine genomic phylogeographic data with information on known intraspecific functional variability and environmental niche modelling and aim to elucidate the particular role of intraspecific variation and ultimately selection in shaping the species' response to the climatic and environmental changes after the end of the last glaciation. Based on the mtDNA markers, bank voles exhibit a complex phylogeographic pattern suggesting population replacement events during the postglacial recolonization of Europe and thus possible involvement of selection in the process. An extensive dataset of more than 6000 SNPs was used to search for signs of population replacement in the bank vole genomic DNA and to investigate the species' postglacial recolonization history throughout its European distribution range. The genomic data revealed even more complex population history than previously detected with mtDNA markers, including not only admixture but also...