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Genetic interactions patterning the Tribolium fate mapZhu, Xin January 1900 (has links)
Doctor of Philosophy / Division of Biology / Susan J. Brown / A segmented body plan is conserved in vertebrates and arthropods. Comparative studies implicate a conserved mode of A-P axis patterning and segmentation among vertebrates: Wnt signaling is involved in fate map patterning along the A-P axis and in segmentation in the posterior region of the embryo. On the other hand, comparative studies in arthropods have revealed distinct modes of development between long and short germ insects, which differ both morphologically and genetically. In the short germ insect Tribolium, a Wnt activity gradient contributes to A-P axis patterning and generates a posterior Tc-cad expression gradient that regulates segmentation through a pair-rule gene clock, forming segments sequentially as in vertebrates. In contrast, instead of Wnt activity, a hierarchy of regulatory genes regionalizes the blastoderm and defines segments simultaneously in the long germ insect Drosophila.
In Tribolium, Tc-zen-1, Tc-otd-1 and Tc-cad play key roles in patterning serosa, head and trunk regions, respectively, of the blastoderm fate map, which are impacted by changes in Wnt activity levels. However, interactions between these genetic factors have not been described. My work revealed that cross talk between the Wnt and Dpp signaling pathways regulates the expression of Tc-zen-1 to determine the serosa. Furthermore, mutually repression between Tcotd-1 and Tc-cad defines the head and trunk regions while mutual repression between Tc-zen-1 and cad determines the posterior extent of the dorsal serosa.
Analysis of target genes of the posterior Tc-cad gradient indicates that the Tc-cad gradient regulates the serial expression of gap genes, which are predominately regulators of Hox genes. Thus the posterior Tc-cad gradient determines segment formation through regulation of pair-rule genes in the insect segmentation clock, and defines segmental identity through regulation of gap genes. Given its effect on Tc-zen-1 and Tc-cad, the Wnt activity gradient is a key organizer of the Tribolium blastoderm fate map.
Since homologs of these genes as well as the Wnt signaling pathway have also been identified in several other short germ band insects, and affect cell fates along the A-P body axis, this work provides a new paradigm for the short germ mode of development to guide studies in other arthropods.
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Estudo da modulação da via Wnt pelo inibidor de Aurora-quinases AMG900 em linhagens celulares de meduloblastoma pediátrico / Study of Modulation of the Wnt pathway by Aurora kinases inhibitor AMG900 in pediatric medulloblastoma cell linesGeron, Lenisa 12 January 2016 (has links)
O meduloblastoma (MB) é o tumor cerebral maligno mais comum na infância. A formação/progressão desta neoplasia foi associada a alterações moleculares, que inclui a desregulação da via de sinalização Wingless (Wnt), responsável pelo desenvolvimento embrionário. Além disso, as proteínas da família Aurora-quinases (A, B e C) têm sido amplamente estudadas, uma vez que a Aurora A e B foram encontrados hiperexpressas em diversas neoplasias, como o MB. Estudos recentes mostraram que existe uma associação entre a Via Wnt e as Aurora-quinases. No entanto, poucos trabalhos foram realizados para confirmar essa associação. Ademais, não existem trabalhos que relatem os efeitos do AMG900, um pan-inibidor de aurora-quinases, em MB, dando enfoque na regulação da via Wnt. Assim, o objetivo deste trabalho foi avaliar a modulação da via Wnt pelo inibidor AMG900 nas linhagens celulares de meduloblastoma pediátrico. Foram realizados os ensaios de PCR convencional, sequenciamento, qRT-PCR, transfecção transiente, ensaio clonogênico, Western Blot e ciclo celular. As linhagens celulares UW402, UW473 e ONS-76 não apresentaram mutações no éxon 3 do gene CTNNB1 (?-catenina) e no éxon 15 do gene APC. Não foi observada uma expressão significativa de CTNNB1, confirmando que as linhagens não possuíam a via Wnt ativa. Com isso foi necessário a transfecção transiente com a ?- catenina. Após este ensaio, houve um aumento da expressão de CTNNB1, Ciclina D1 e CMyc nas três linhagens, o que não ocorreu com as Auroras A e B. No ensaio clonogênico foi observado uma redução do número de colônias nas linhagens UW473 e ONS-76. Observou-se um aumento da expressão proteica da ?-catenina, da Aurora A e B na UW473, o que ocorreu somente com a ?-catenina na linhagem ONS-76. Após o tratamento com o AMG900 ocorreu uma diminuição da expressão proteica de ?-catenina, da Aurora A e B em ambas as linhagens. A transfecção não alterou o percentil celular em G2/M na UW402 e UW473. Já na ONS-76 houve um aumento significativo em G2/M, e o AMG900 potencializou esse bloqueio apenas nessa linhagem. Os resultados sugerem que pode haver alguma relação entre a inibição das proteínas Aurora-quinases e a expressão de proteínas da via Wnt. / Medulloblastoma (MB) is the most common malignant brain tumor in childhood. Tumor formation/progression has been associated to molecular alterations that include dysregulation of signaling pathway Wingless (Wnt), responsible for embryonic development. In addition, cell cycle proteins Aurora-kinase (A, B and C) have been widely studied since Aurora A and B were found overexpressed in many cancers such as MB. Recent studies show that there is an association between Wnt pathway and Aurora kinase proteins. However, few studies have been conducted to confirm this association. Moreover, there are no studies reporting the effects of AMG900 in MB, by focusing on the regulation of the Wnt pathway. The aim of this study is to evaluate Wnt pathway modulation by Aurora kinases inhibitor AMG900 in pediatric medulloblastoma cell lines. Conventional PCR, sequencing, qRT-PCR, transient transfection, clonogenic assay, Western Blot and cell cycle assays were performed. UW402, UW473 and ONS-76 cell lines did not present mutations in exon 3 of CTNNB1 gene and exon 15 of APC gene. There was no significant expression of CTNNB1 and their target genes in these cell lines, confirming that they did not have Wnt pathway activated. Considering this, transient transfection was necessary. After this trial, there was an increase in expression of CTNNB1 gene and its target genes Cyclin D1 and C-Myc in the three cell lines, which was not observed in Aurora kinases. Furthermore, in the clonogenic assay, a reduction in the number of colonies in UW473 and ONS-76 cell lines was observed. It was also observed an increase in ?-catenin protein, Aurora A and B in UW473 cell line, but not in ONS-76 cell line. However, after treatment there was a decrease in protein expression of ?-catenin, Aurora A and B in both cells. Transfection did not change the cellular percentile in G2 / M in UW402 and UW473. In ONS-76 there was a significant increase in G2 / M, and the treatment with AMG900 potentiated this block only in this cell line. Results suggest that there may be some relation between the inhibition of Aurora kinase protein and protein expression in Wnt pathway.
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Functional Analysis of Two Major Sperm Tail Proteins Identifies ODF1 as Being Essential for the Tight Linkage of the Sperm Head to the Tail via SPAG4 and ODF2 as A Component of the β-catenin Destruction ComplexYang, Kefei 04 June 2014 (has links)
No description available.
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Canonical Wg/Wnt pathway regulates Wolbachia intracellular density in DrosophilaHsia, Hsin-Yi 23 November 2016 (has links)
Wolbachia are widely spread, maternally transmitted insect endosymbiotic intracellular bacteria. They have been implicated in the control of several insect transmitted diseases, including dengue, yellow fever, Zika and malaria. Effective pathogen suppression in the insect host is shown to be proportional to the intracellular levels of bacteria. Therefore, understanding the molecular mechanisms underlying Wolbachia accumulation within organisms is extremely important for future epidemic control and research. Using Drosophila as a model insect, our lab has previously observed Wolbachia tropism to stem cell niches. Current work has identified polar cells as an additional site of Wolbachia tropism and demonstrated that Wg/Wnt signaling is important for Wolbachia intracellular accumulation in these somatic cells. In this thesis, we first observed that the Wg/Wnt pathway protein Armadillo also controls Wolbachia levels in the germline cells, indicating the possibility of having a conserved molecular mechanism controlling Wolbachia. Using RNAi and small molecule inhibitors of Shaggy, another component of the canonical Wg/Wnt pathway, we demonstrate that the canonical Wg/Wnt signaling is essential for Wolbachia intracellular accumulation. Our investigation provides fundamental insights into the mechanisms of Wolbachia intracellular accumulation. Furthermore, it offers novel strategies to modulate Wolbachia in non-model insect species, including various disease transmitting Anopheles, Culex, and Aedes. These findings potentially will increase the effectiveness of a Wolbachia-based vector transmitted disease suppression. / 2017-02-28
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Estudo da modulação da via Wnt pelo inibidor de Aurora-quinases AMG900 em linhagens celulares de meduloblastoma pediátrico / Study of Modulation of the Wnt pathway by Aurora kinases inhibitor AMG900 in pediatric medulloblastoma cell linesLenisa Geron 12 January 2016 (has links)
O meduloblastoma (MB) é o tumor cerebral maligno mais comum na infância. A formação/progressão desta neoplasia foi associada a alterações moleculares, que inclui a desregulação da via de sinalização Wingless (Wnt), responsável pelo desenvolvimento embrionário. Além disso, as proteínas da família Aurora-quinases (A, B e C) têm sido amplamente estudadas, uma vez que a Aurora A e B foram encontrados hiperexpressas em diversas neoplasias, como o MB. Estudos recentes mostraram que existe uma associação entre a Via Wnt e as Aurora-quinases. No entanto, poucos trabalhos foram realizados para confirmar essa associação. Ademais, não existem trabalhos que relatem os efeitos do AMG900, um pan-inibidor de aurora-quinases, em MB, dando enfoque na regulação da via Wnt. Assim, o objetivo deste trabalho foi avaliar a modulação da via Wnt pelo inibidor AMG900 nas linhagens celulares de meduloblastoma pediátrico. Foram realizados os ensaios de PCR convencional, sequenciamento, qRT-PCR, transfecção transiente, ensaio clonogênico, Western Blot e ciclo celular. As linhagens celulares UW402, UW473 e ONS-76 não apresentaram mutações no éxon 3 do gene CTNNB1 (?-catenina) e no éxon 15 do gene APC. Não foi observada uma expressão significativa de CTNNB1, confirmando que as linhagens não possuíam a via Wnt ativa. Com isso foi necessário a transfecção transiente com a ?- catenina. Após este ensaio, houve um aumento da expressão de CTNNB1, Ciclina D1 e CMyc nas três linhagens, o que não ocorreu com as Auroras A e B. No ensaio clonogênico foi observado uma redução do número de colônias nas linhagens UW473 e ONS-76. Observou-se um aumento da expressão proteica da ?-catenina, da Aurora A e B na UW473, o que ocorreu somente com a ?-catenina na linhagem ONS-76. Após o tratamento com o AMG900 ocorreu uma diminuição da expressão proteica de ?-catenina, da Aurora A e B em ambas as linhagens. A transfecção não alterou o percentil celular em G2/M na UW402 e UW473. Já na ONS-76 houve um aumento significativo em G2/M, e o AMG900 potencializou esse bloqueio apenas nessa linhagem. Os resultados sugerem que pode haver alguma relação entre a inibição das proteínas Aurora-quinases e a expressão de proteínas da via Wnt. / Medulloblastoma (MB) is the most common malignant brain tumor in childhood. Tumor formation/progression has been associated to molecular alterations that include dysregulation of signaling pathway Wingless (Wnt), responsible for embryonic development. In addition, cell cycle proteins Aurora-kinase (A, B and C) have been widely studied since Aurora A and B were found overexpressed in many cancers such as MB. Recent studies show that there is an association between Wnt pathway and Aurora kinase proteins. However, few studies have been conducted to confirm this association. Moreover, there are no studies reporting the effects of AMG900 in MB, by focusing on the regulation of the Wnt pathway. The aim of this study is to evaluate Wnt pathway modulation by Aurora kinases inhibitor AMG900 in pediatric medulloblastoma cell lines. Conventional PCR, sequencing, qRT-PCR, transient transfection, clonogenic assay, Western Blot and cell cycle assays were performed. UW402, UW473 and ONS-76 cell lines did not present mutations in exon 3 of CTNNB1 gene and exon 15 of APC gene. There was no significant expression of CTNNB1 and their target genes in these cell lines, confirming that they did not have Wnt pathway activated. Considering this, transient transfection was necessary. After this trial, there was an increase in expression of CTNNB1 gene and its target genes Cyclin D1 and C-Myc in the three cell lines, which was not observed in Aurora kinases. Furthermore, in the clonogenic assay, a reduction in the number of colonies in UW473 and ONS-76 cell lines was observed. It was also observed an increase in ?-catenin protein, Aurora A and B in UW473 cell line, but not in ONS-76 cell line. However, after treatment there was a decrease in protein expression of ?-catenin, Aurora A and B in both cells. Transfection did not change the cellular percentile in G2 / M in UW402 and UW473. In ONS-76 there was a significant increase in G2 / M, and the treatment with AMG900 potentiated this block only in this cell line. Results suggest that there may be some relation between the inhibition of Aurora kinase protein and protein expression in Wnt pathway.
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Database for the Study of Biological Pathways, with Wnt Signaling Pathway Use CaseMailavaram, Sravanthi 17 April 2009 (has links)
No description available.
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Wnt/ß-catenin signaling pathway in non-myocyte lineages in the heartFang, Ming 26 May 2016 (has links)
No description available.
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Análise comparativa da expressão dos genes Vangl1 e Vangl2 durante a ontogênese da galinha (Gallus gallus) / Comparative analysis of Vangl1 and Vangl2 gene expression during chicken ontogenesis (Gallus gallus)Pedrosa, Angelica Vasconcelos, 1986- 24 August 2018 (has links)
Orientador: Lúcia Elvira Alvares / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-24T17:37:00Z (GMT). No. of bitstreams: 1
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Previous issue date: 2013 / Resumo: A correta padronização do corpo do embrião requer a atividade de diferentes vias de sinalização. Dentre elas, uma que se destaca é via de sinalização Wnt de polaridade celular planar (Wnt/PCP), que é responsável pelo controle da polaridade celular e pela organização celular de diversos tecidos nos animais. Uma vez interrompida, a via Wnt/PCP pode causar falhas no fechamento do tubo neural, provocando defeitos congênitos. Em seres humanos, mutações em componentes-chave da via Wnt/PCP como as proteínas codificadas pelos genes Vangl1 e Vangl2 têm sido associadas à graves malformações geradas por falhas no fechamento do tubo neural. Estruturalmente, ambos os genes Vangl1 e Vangl2 codificam proteínas de superfície transmembranares, essenciais para o desenvolvimento apropriado do embrião. O presente trabalho teve como objetivo a caracterização do padrão de expressão dos genes Vangl1 e Vangl2 durante a embriogênese de Gallus gallus. Ensaios de hibridação in situ em embrião inteiro (whole mount) e cortes em vibratómo foram realizados com a finalidade de estabelecer temporal e espacialmente o padrão de expressão dos genes Vangl1 e Vangl2. Como resultado, observou-se que estes genes são expressos durante as etapas de gastrulação, neurulação e no início da organogênese do desenvolvimento embrionário de Gallus gallus. No início da gastrulação, os genes Vangl1 e Vangl2 possuem domínios de expressão comuns nos embriões de galinha, uma vez que ambos são expressos na linha primitiva, nódulo de Hensen e crescente cardiogênico. Contudo, nossos dados revelaram particularidades na expressão destes genes, uma vez que há uma predominância dos transcritos de Vangl1 na região posterior da linha primitiva, enquanto Vangl2 apresenta uma expressão uniforme ao longo desta estrutura. Em adição, enquanto Vangl1 é expresso na notocorda e em toda a extensão do nódulo de Hensen, Vangl2 é expresso no entorno desta estrutura. Ao longo da neurulação e na organogênese inicial, ambos os genes Vangl são expressos de maneira similar, em domínios que abrangem a placa, as pregas e o tubo neural. Outros importantes domínios de expressão dos Vangl correspondem às vesículas ópticas e óticas, às vesículas encefálicas particularmente na região das flexuras encefálicas, aos diferentes tipos de mesoderma (paraxial, intermediário e lateral) e ao assoalho da faringe. Ao comparar os resultados obtidos por hibridação in situ em galinha ao um levantamento bibliográfico sobre outros vertebrados, observou-se uma sobreposição dos domínios-chave de expressão nos diferentes organismos, demonstrando a conservação filogenética da atividade destes genes e sugerindo uma possível conservação funcional. Desta forma, nossos dados sugerem que os genes Vangl desempenham um importante papel no desenvolvimento embrionário de aves, possivelmente coordenando os movimentos morfogenéticos durante a gastrulação, bem como a formação da placa neural e posterior dobramento e fechamento do tubo neural, além de outros processos da embriogênese de aves / Abstract: The correct patterning of the embryo's body requires the activity of different signaling pathways. Among them, one that stands out is the Wnt Planar Cell Polarity Signaling Pathway (Wnt/PCP), which is responsible for controlling the cell polarity and cellular organization of many tissues in animals. Failures in the Wnt/PCP signaling can cause neural tube birth defects. In humans, mutations in key components of the Wnt/PCP as the Vangl1 and Vangl2 molecules were identified in patients with neural tube defects. Structurally, both Vangl1 and Vangl2 genes encode transmembrane surface proteins similar, which are essential to proper development. The present study aimed to characterize the expression pattern of Vangl1 and Vangl2 genes during embryogenesis in Gallus gallus. Whole-mount in situ hybridization assays and vibratome sectioning of embryos were conducted in order to establish the spatial and temporal expression pattern of Vangl1 and Vangl2 genes. Our results showed that these genes are expressed during gastrulation, neurulation and early organogenesis in Gallus gallus. At the onset of Gastrulation, Vangl1 and Vangl2 genes have common areas of expression in chicken embryos, since both are expressed in the primitive streak, Hensen's node and cardiogenic crescent. However, our data showed particularities in the expression of these genes, since there is a predominance of Vangl1 transcripts in the posterior region of the primitive streak while Vangl2 has a uniform expression throughout that structure. In addition, while Vangl1 is expressed in the notochord and in the full length of the Hensen's node, Vangl2 is expressed only around this structure. Throughout neurulation and early organogenesis, both Vangl genes are expressed in a similar manner on the neural plate, neural groove, neural folds and in the neural tube. Other important areas of Vangl expression correspond to optical and otic vesicles, the brain vesicles, the different types of mesoderm (paraxial, intermediate and lateral) and the floor of the pharynx. By comparing the chicken expression of Vangl genes with other vertebrates, we notice that there are overlapping expression patterns among key areas among different organisms, showing a phylogenetic conservation of expression domains and suggesting a possible functional conservation. Overall, our data suggests that Vangl genes play an important role in embryonic development of bird, possibly by coordinating the morphogenetic movements during gastrulation, as well as the formation of neural tube, among other processes during the birds embriogenesis / Mestrado / Biologia Celular / Mestra em Biologia Celular e Estrutural
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Estudos sobre os genes da família Dapper = origem, evolução e análise da expressão durante a ontogênese dos membros de galinha = Studies on the Dapper gene family : origin, evolution and expression analysis during chicken limb development / Studies on the Dapper gene family : origin, evolution and expression analysis during chicken limb developmentSobreira, Debora Rodrigues, 1981- 07 November 2013 (has links)
Orientadores: Lúcia Elvira Alvares, José Xavier Neto / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T10:52:38Z (GMT). No. of bitstreams: 1
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Previous issue date: 2013 / Resumo: Os genes da família Dapper (Dpr) codificam proteínas adaptadoras capazes de ligar-se fisicamente a diferentes moléculas e modular as vias de sinalização Wnt e TGF-?. Diferentes análises funcionais revelaram que os Dpr atuam na especificação do eixo corporal e do tecido neural, nos movimentos morfogenéticos, no desenvolvimento do olho, na indução da cardiogênese, adipogênse e cicatrização. Diversos estudos foram realizados a fim de compreender o papel desempenhado pelos Dpr durante a embriogenêse dos vertebrados e na homeostase de tecidos adultos. Contudo, muitas questões ainda necessitam ser elucidadas. Este projeto de Doutorado teve como objetivo (1) descrever os sítios de expressão da família gênica Dpr durante a ontogênese dos membros de galinha, associando-a as vias de sinalização Wnt e TGF-? e (2) investigar a origem e a evolução desses genes durante a filogenia dos metazoários. Nossos resultados confirmaram que os genes Dpr são dinamicamente expressos durante o desenvolvimento dos membros de galinha, provavelmente, modulando os sinais Wnt e TGF-?. Os genes Dpr são encontrados no mesênquima indiferenciado dos membros em formação e em células progenitoras de condrócitos, pericôndrio e tendões. Esses resultados sugerem as moléculas Dpr como um novo grupo de marcadores do desenvolvimento dos membros em galinha. Já nossas análises filogenéticas revelaram que os Dprs surgiram durante a evolução dos organismos deuterostômios e um novo ortólogo dessa família de proteínas, denominado Dpr4, foi descrito. Acreditamos que o nosso trabalho irá fornecer bases para estudos moleculares com o intuito de estabelecer a função individual de cada membro da família Dpr, bem como auxiliar no entendimento sobre como estas proteínas podem interagir e cooperar entre si para modular diferentes vias de sinalização molecular em diferentes contextos celulares / Abstract: The Dapper (Dpr) genes form a small gene family of adaptor proteins important to several processes of vertebrates development, such as the specification of the body axis and neural tissue, morphogenetic movements, eye development, induction of cardiogenesis, adipogenesis and wound healing, by modulating the Wnt and TGF-? signaling pathways using specific conserved domains/motifs. Three Dpr genes have been identified in human and mouse, two in chicken, one in frog and two in zebrafish genome. Since the discovery of Dpr proteins, several assays have been performed in order to understand the role of this family during embryogenesis, although many questions still need to be elucidated. Thus, this PhD project aimed to (1) describe the possible role of Dpr genes during ontogeny of chicken regarding the regulation of Wnt and TGF-? signaling pathways and (2) investigate the origin and evolution of Dpr family over the course of metazoan evolution. Our results demonstrated that Dpr genes are involved in chicken limb development, probably, by modulating Wnt and TGF-? signals. Dpr genes were found in the undifferentiated limb mesenchyme, progenitor of chondrocytes, perichondrium and tendons. These results suggest that Dpr genes are good candidates to a new set of markers in chicken limb development. Furthermore, our phylogenetic analysis revealed that the Dprs arose late during the deuterostomes evolution and allowed the identification of a new Dpr paralog (Dpr4), meaning that a repertoire of four Dact genes is found in vertebrates. Thus, our work will provide the basis for molecular studies in order to establish the role of each individual member of this family as well as how the set of Dpr proteins can interact and cooperate to modulate different molecular signaling pathways in different cellular contexts / Doutorado / Biologia Tecidual / Doutora em Biologia Celular e Estrutural
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Adjusting Wnt signaling, nové regulační mechanismy signální dráhy Wnt / Adjusting Wnt signaling, new regulatory mechanisms of the Wnt pathwayFafílek, Bohumil January 2012 (has links)
4 Abstract The Wnt pathway is one of the major signaling cascades contributing to multiple cellular processes during embryogenesis, and adult tissue homeostasis and regeneration. Moreover, aberrant activation of the Wnt signaling pathway is connected with development of neoplasia, notably colorectal cancer. The aim of the thesis was to identify new ways of the Wnt pathway regulation to understand better physiological as well as non-physiological mechanisms of Wnt signaling. The results are summarized in four publications. The first article deals with TROY, a member of tumor necrosis factor receptor family. We identified TROY as a Wnt target gene during our search for Wnt responsive genes in colorectal cancer cell lines. Additionally, we detected expression of Troy in tumors of two mouse models of intestinal cancer. In the healthy gut, Troy is produced in fast cycling intestinal stem cells where negatively regulates the Wnt pathway. The second study focuses on processing and posttranslational modification of murine Wnt1 and Wnt3a. Wnts are glycosylated and double acetylated by lipid adducts and our results revealed that O-linked acylation of serine is required for the subsequent S-palmitoylation of cysteine. Moreover, acylation of Wnts is connected with their signaling activity which is related to Wnt1 and...
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