Diss. (sammanfattning) Umeå : Univ., 2004. / Härtill 5 uppsatser.

No description available.

No description available.

Wolbachia are maternally transmitted endosymbionts that infect a large proportion of arthropods and reduce vector competency of mosquitoes carrying deadly diseases such as Dengue, Zika, West Nile Virus, and Chikungunya. Wolbachia preferentially infect specific host cells, a phenomenon termed tissue tropism. Wolbachia tissue tropism has consequences in proper vertical and horizontal transmission. Although Wolbachia tropism to multiple Drosophila gonadal cell types has been identified, the mechanisms of tropism during development have not been characterized, in part because Wolbachia infected cell types are challenging to study during morphogenesis. Here we describe a novel Wolbachia tropism to polar cells (PCs) of the Drosophila ovary, a developmentally well characterized system. We show that Wolbachia intracellular accumulation is triggered by specific events of PC morphogenesis including differentiation from progenitors, and during stage 8 to 9 transition. Using genetic tools, we also demonstrate that induction of ectopic PC fate is sufficient to drive Wolbachia tropism. These findings implicate a tight coordination of host developmental events with Wolbachia tropism. Wolbachia tropism to multiple host cell types suggest that host pathways common to these would be conducive to intracellular Wolbachia growth. Indeed, we found that Wnt signaling, is active in gonadal cell types with Wolbachia tropism. Wnt signaling, first characterized in embryonic development and patterning, has novel functions in immunity and intracellular pathogen survival. Using RNAi mediated gene knockdowns, we studied the effect of Wnt signaling on Wolbachia in various infected cell types, including the testis stem cell niche, PCs and germline. Reduction of Wnt signaling caused a decrease in Wolbachia density and increased signaling led to higher density suggesting the reliance of Wolbachia on host Wnt signaling for its tropism. Moreover, expression of ectopic Wnt signaling was sufficient to drive Wolbachia tropism to previously uninfected tissues such as the D. melanogaster female germline stem cell niche. Finally, small molecule Wnt signaling agonists were sufficient to drive high Wolbachia titers in mosquito cell lines. These findings describe the effect of host signaling on Wolbachia tropism and provide an approach to affect Wolbachia levels in disease-causing vectors, thereby contributing to Wolbachia based vector control strategies.

A sequência de 35 aminoácidos da alça V3 da gp120 do gene env do HIV-1 é o principal determinante do tropismo viral pelos correceptores CCR5 ou CXCR4, utilizados pelo HIV-1 para a entrada na célula. O desenvolvimento de estratégias antirretrovirais baseadas no uso dos correceptores representa um avanço importante para o controle da progressão da infecção. Entretando, o uso clínico dos antagonistas de CCR5 implica na determinação do tropismo das cepas virais do indivíduo infectado e os programas preditores de bioinformática para a determinação do tropismo poderiam ser uma alternativa mais acessível para a triagem dos candidatos ao uso dos antagonistas de CCR5. Este estudo teve como objetivo utilizar ferramentas de bioinformática para a predição de tropismo e avaliar sua aplicabilidade na prática clínica. Foram coletadas amostras de sangue periférico de 101 indivíduos infectados pelo HIV-1 e sob acompanhamento clínico, dos quais foram extraídas amostras de DNA proveniente de PBMCs. As amostras de DNA foram amplificadas por PCR para a região da alça V3, das quais foram obtidas 94 sequências. Os sistemas preditivos foram avaliados utilizando 185 sequências com tropismo conhecido provenientes de banco de dados. Com base nesta análise foi possível elaborar um algoritmo para a predição do tropismo com 94% de confiabilidade. Assim, a predição das 94 amostras demonstrou uma prevalência de 80% (n=75) de cepas R5 e 20% (n=19) de cepas X4. Os sistemas preditivos de tropismo podem representar uma importante estratégia para a triagem dos candidatos ao uso dos antagonistas de coreceptor, porém, não são capazes de substituir completamente os ensaios padrão-ouro para a determinação do tropismo. / The 35 amino acids of the V3-gp120 of HIV-1 env gene is the main determinant of viral tropism by the coreceptors CCR5 and CXCR4 used for HIV-1 cell entry. The development of antiretroviral strategies based on the coreceptor usage represents an important step to control the infection progression. However, the clinical application of CCR5 antagonists involves the coreceptor usage determination of viral strains in the infected individual. The bioinformatics predictive programs for coreceptor usage determination could be a more available alternative for screening candidates to receive CCR5 antagonists. This study aimed to employ bioinformatics tools to predict tropism and assess its applicability in clinical practice. Peripheral blood samples were collected from 101 individuals infected with HIV-1 and under clinical follow-up. DNA samples were extracted from PBMCs. The DNA samples were amplified by PCR and 94 V3 sequences were obtained. The predictive systems were evaluated using 185 sequences of known tropism from a database. This analysis provides the construction of an algorithm showing 94% of reliability. Thus, the 94 sample prediction showed a prevalence of 80% (n=75) of R5 strain and 20% (n=19) of X4 strain. The predictive systems could be an important strategy in the screening of the tropism. Nonetheless, they are not able to fully replace the coreceptor usage gold-standard assays.

A sequência de 35 aminoácidos da alça V3 da gp120 do gene env do HIV-1 é o principal determinante do tropismo viral pelos correceptores CCR5 ou CXCR4, utilizados pelo HIV-1 para a entrada na célula. O desenvolvimento de estratégias antirretrovirais baseadas no uso dos correceptores representa um avanço importante para o controle da progressão da infecção. Entretando, o uso clínico dos antagonistas de CCR5 implica na determinação do tropismo das cepas virais do indivíduo infectado e os programas preditores de bioinformática para a determinação do tropismo poderiam ser uma alternativa mais acessível para a triagem dos candidatos ao uso dos antagonistas de CCR5. Este estudo teve como objetivo utilizar ferramentas de bioinformática para a predição de tropismo e avaliar sua aplicabilidade na prática clínica. Foram coletadas amostras de sangue periférico de 101 indivíduos infectados pelo HIV-1 e sob acompanhamento clínico, dos quais foram extraídas amostras de DNA proveniente de PBMCs. As amostras de DNA foram amplificadas por PCR para a região da alça V3, das quais foram obtidas 94 sequências. Os sistemas preditivos foram avaliados utilizando 185 sequências com tropismo conhecido provenientes de banco de dados. Com base nesta análise foi possível elaborar um algoritmo para a predição do tropismo com 94% de confiabilidade. Assim, a predição das 94 amostras demonstrou uma prevalência de 80% (n=75) de cepas R5 e 20% (n=19) de cepas X4. Os sistemas preditivos de tropismo podem representar uma importante estratégia para a triagem dos candidatos ao uso dos antagonistas de coreceptor, porém, não são capazes de substituir completamente os ensaios padrão-ouro para a determinação do tropismo. / The 35 amino acids of the V3-gp120 of HIV-1 env gene is the main determinant of viral tropism by the coreceptors CCR5 and CXCR4 used for HIV-1 cell entry. The development of antiretroviral strategies based on the coreceptor usage represents an important step to control the infection progression. However, the clinical application of CCR5 antagonists involves the coreceptor usage determination of viral strains in the infected individual. The bioinformatics predictive programs for coreceptor usage determination could be a more available alternative for screening candidates to receive CCR5 antagonists. This study aimed to employ bioinformatics tools to predict tropism and assess its applicability in clinical practice. Peripheral blood samples were collected from 101 individuals infected with HIV-1 and under clinical follow-up. DNA samples were extracted from PBMCs. The DNA samples were amplified by PCR and 94 V3 sequences were obtained. The predictive systems were evaluated using 185 sequences of known tropism from a database. This analysis provides the construction of an algorithm showing 94% of reliability. Thus, the 94 sample prediction showed a prevalence of 80% (n=75) of R5 strain and 20% (n=19) of X4 strain. The predictive systems could be an important strategy in the screening of the tropism. Nonetheless, they are not able to fully replace the coreceptor usage gold-standard assays.

No description available.

The plant hormones gibberellins (GAs) regulate multiple processes of plant development. Most of this regulation occurs at the transcriptional level, through the activity of the DELLAs, which are nuclear-localized proteins subjected to GA-mediated proteolitic degradation. DELLAs are encoded by five genes, and genetic studies show that each DELLA displays specific, but also partially overlapping roles with respect to their paralogs. In this Thesis, we have addressed two issues: (1) the contribution of DELLA multiplication to the diversification of functions controlled by GAs; and (2) the identification of direct targets regulated by DELLAs in etiolated seedlings with special attention to those involved in differential growth processes. Using combinations of mutants and transgenic lines expressing two phylogenetically distant DELLA genes (RGA and RGL2), we have found that these two DELLA proteins can perform each other's role as long as they are expressed under the reciprocal promoters, indicating that DELLA subfunctionalization relies mainly on their differential expression patterns. To identify direct DELLA targets, we have performed transcriptomic analyses of dark-grown seedlings expressing an inducible version of gai-1, a stable, dominant allele of a DELLA gene. This approach rendered a list of over 150 genes differentially expressed after induction of gai-1. The presence of several auxin-related genes among the primary targets of DELLA proteins has allowed us to establish a new role for GAs in the modulation of hypocotyl gravitropism through the repression of IAA19/MASSUGU2 expression by DELLAs. Moreover, the repression of HOOKLESS1 and the auxin efflux carriers PIN3 and PIN7 by DELLAs, is proposed as the molecular mechanism to explain the already known physiological regulation of apical hook development by GAs. / Gallego Bartolomé, J. (2011). Della protein function during differential growth processes in arabidopsis [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/11403

A descoberta do tropismo do Vírus da Imunodeficiência Humana tipo 1 (HIV-1) pelos correceptores CCR5 e CXCR4 propiciou o desenvolvimento de novas estratégias terapêuticas com alvo nestes correceptores. Os antagonistas de CCR5 são fármacos que bloqueiam o correceptor CCR5, impedindo a entrada do (HIV) na célula, levando a uma diminuição significativa da viremia. Porém, a administração deste medicamento depende da determinação do tropismo dos vírus do indivíduo infectado antes e ao longo do uso desta estratégia terapêutica. A alça V3 da gp120 do envelope do HIV-1 é considerada o principal determinante do tropismo, mas outros fatores como diferenças intrínsecas entre as variantes do HIV-1, o número de sítios de N-glicosilação, variações no comprimento de determinadas regiões e a carga elétrica global da gp120 também podem afetar o tropismo viral. Este estudo apresenta a caracterização molecular da gp120 e a relação dos fatores moleculares com o tropismo pelos correceptores CCR5 e CXCR4. Foram incluídos no estudo 283 indivíduos infectados pelo HIV-1, dos quais foi possível obter 265 sequências da alça V3, resultados de tropismo fenotípico para 78 amostras e 20 sequências da gp120 completa. O tropismo fenotípico demonstrou a prevalência de vírus R5 (70,5%) e a presença de apenas uma amostra contendo exclusivamente vírus X4. O tropismo genotípico classificou 71,7% das sequências como vírus R5 e 28,2% como capazes de utilizar o correceptor CXCR4. A concordância entre os resultados de tropismo fenotípico e genotípico foi de 74,5%. Houve a prevalência do subtipo B (82%), sendo que a variante B\'(GWGR) esteve presente em 34,5% dos casos. Em média, as sequências da gp120 apresentaram 22 sítios de N-glicosilação, concentrados nas regiões C2 e C4. A carga elétrica líquida apresentou médias semelhantes entre vírus R5 e X4 tanto para a alça V3 quanto para a gp120. As características moleculares descritas neste estudo apresentaram distribuição semelhante entre vírus R5 e X4, não sendo possível estabelecer relações entre estes fatores a determinação do tropismo. / The tropism of Human Immunodeficiency Virus type 1 (HIV - 1) for CCR5 and CXCR4 coreceptors has provided the development of new therapeutic strategies targeting these coreceptors. CCR5 antagonists are able to prevent the HIV cell entry by blocking the CCR5 coreceptor, improving the viremia decreasing. However, this drug administration depends on viral tropism determination before and during the use of this therapeutic strategy. The V3 loop of the HIV-1 gp120 envelope is the major tropism determinant, but other factors such HIV-1 genetic variability, number of potential N-linked glycosylation sites, length variations in some protein domains and the gp120 global net charge may also affect the viral tropism. This study presents the molecular characterization of gp120 and the relationship between molecular factors and the tropism for CCR5 and CXCR4 coreceptors. A total of 283 HIV-1 infected subjects were included in this study. It was possible to obtain 265 V3 loop sequences, 78 results of phenotypic tropism and 20 sequences of the whole gp120. Phenotypic tropism showed the prevalence of R5 viruses (70.5%) while exclusively X4 viruses were found in only one sample. Genotypic tropism classified 71.7% of the sequences as R5 and 28.2% as virus able to use the CXCR4 correceptor. The agreement between phenotypic and genotypic tropism results was 74.5%. There was a prevalence of subtype B (82%), and the B\' (GWGR) variant was present in 34.5% of the cases. gp120 showed a mean of 22 N-linked glycosylation sites, which were more frequent in the C2 and C4 regions. The net charge showed similar means between R5 and X4 viruses for both V3 loop and gp120. The molecular characteristics described in this study showed similar distribution between R5 and X4 viruses and it was not possible to establish relationships between these factors the tropism determination.

A descoberta do tropismo do Vírus da Imunodeficiência Humana tipo 1 (HIV-1) pelos correceptores CCR5 e CXCR4 propiciou o desenvolvimento de novas estratégias terapêuticas com alvo nestes correceptores. Os antagonistas de CCR5 são fármacos que bloqueiam o correceptor CCR5, impedindo a entrada do (HIV) na célula, levando a uma diminuição significativa da viremia. Porém, a administração deste medicamento depende da determinação do tropismo dos vírus do indivíduo infectado antes e ao longo do uso desta estratégia terapêutica. A alça V3 da gp120 do envelope do HIV-1 é considerada o principal determinante do tropismo, mas outros fatores como diferenças intrínsecas entre as variantes do HIV-1, o número de sítios de N-glicosilação, variações no comprimento de determinadas regiões e a carga elétrica global da gp120 também podem afetar o tropismo viral. Este estudo apresenta a caracterização molecular da gp120 e a relação dos fatores moleculares com o tropismo pelos correceptores CCR5 e CXCR4. Foram incluídos no estudo 283 indivíduos infectados pelo HIV-1, dos quais foi possível obter 265 sequências da alça V3, resultados de tropismo fenotípico para 78 amostras e 20 sequências da gp120 completa. O tropismo fenotípico demonstrou a prevalência de vírus R5 (70,5%) e a presença de apenas uma amostra contendo exclusivamente vírus X4. O tropismo genotípico classificou 71,7% das sequências como vírus R5 e 28,2% como capazes de utilizar o correceptor CXCR4. A concordância entre os resultados de tropismo fenotípico e genotípico foi de 74,5%. Houve a prevalência do subtipo B (82%), sendo que a variante B\'(GWGR) esteve presente em 34,5% dos casos. Em média, as sequências da gp120 apresentaram 22 sítios de N-glicosilação, concentrados nas regiões C2 e C4. A carga elétrica líquida apresentou médias semelhantes entre vírus R5 e X4 tanto para a alça V3 quanto para a gp120. As características moleculares descritas neste estudo apresentaram distribuição semelhante entre vírus R5 e X4, não sendo possível estabelecer relações entre estes fatores a determinação do tropismo. / The tropism of Human Immunodeficiency Virus type 1 (HIV - 1) for CCR5 and CXCR4 coreceptors has provided the development of new therapeutic strategies targeting these coreceptors. CCR5 antagonists are able to prevent the HIV cell entry by blocking the CCR5 coreceptor, improving the viremia decreasing. However, this drug administration depends on viral tropism determination before and during the use of this therapeutic strategy. The V3 loop of the HIV-1 gp120 envelope is the major tropism determinant, but other factors such HIV-1 genetic variability, number of potential N-linked glycosylation sites, length variations in some protein domains and the gp120 global net charge may also affect the viral tropism. This study presents the molecular characterization of gp120 and the relationship between molecular factors and the tropism for CCR5 and CXCR4 coreceptors. A total of 283 HIV-1 infected subjects were included in this study. It was possible to obtain 265 V3 loop sequences, 78 results of phenotypic tropism and 20 sequences of the whole gp120. Phenotypic tropism showed the prevalence of R5 viruses (70.5%) while exclusively X4 viruses were found in only one sample. Genotypic tropism classified 71.7% of the sequences as R5 and 28.2% as virus able to use the CXCR4 correceptor. The agreement between phenotypic and genotypic tropism results was 74.5%. There was a prevalence of subtype B (82%), and the B\' (GWGR) variant was present in 34.5% of the cases. gp120 showed a mean of 22 N-linked glycosylation sites, which were more frequent in the C2 and C4 regions. The net charge showed similar means between R5 and X4 viruses for both V3 loop and gp120. The molecular characteristics described in this study showed similar distribution between R5 and X4 viruses and it was not possible to establish relationships between these factors the tropism determination.