Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A junção aderente (JA) é um dos principais componentes do complexo juncional apical. Esta juncão é um complexo multiprotéico em que a E-caderina, uma glicoproteína transmembrana, atua como principal mediadora da adesão célula-célula. Sua ancoragem ao citoesqueleto de actina ocorre via proteínas da família das cateninas. Modificações pós-traducionais da E-caderina, como fosforilação e glicosilação, podem modular a estabilidade e organização das JAs. Muitos estudos têm sugerido que no câncer a invasão e a metástase podem estar associadas a arranjos de glicanos na superfície celular. Em câncer colo-retal, o papel de alterações na expressão de glicanos sobre a estabilidade da adesão mediada por Ecaderina ainda não está claro. Neste estudo, investigamos a relação entre estas alterações e a estabilidade das JAs em células de câncer colo-retal. Nós utilizamos duas linhagens celulares com diferentes potenciais metastáticos, Caco-2 and HCT- 116, que constituem dois modelos de JAs: estáveis e instáveis, respectivamente. Ensaios de precipitação de lectinas e immunoblotting demonstraram que em HCT- 116, a linhagem mais invasiva, a E-caderina apresenta uma diminuição de glicanos reconhecidos pelas lectinas HPA e WGA, que reconhecem resíduos de Nacetilgalactosamina e, _-N-acetilhexosaminas e ácido siálico, respectivamente. Concomitantemente, em HCT-116 também foi observado um aumento de glicanos reconhecidos pelas lectinas L-PHA e E-PHA, que reconhecem respectivamente: acetilglicosamina_1,6-ligada formando N-glicanos tri- e tetraantenados e, Nacetilglicosamina bisectante _1,4-ligada formando N-glicanos biantenados. Ensaios de imunofluorescência mostraram que a presença desses glicanos reconhecidos por L-PHA, em HCT-116, é intensa na região de contato célula-célula quando comparada com células Caco-2, em que a marcação foi observada, predominantemente, na região apical. Além disso, a inibição completa da Nglicosilação por tunicamicina ou a inibição da síntese de N-glicanos complexos por swainsonina, aumentou a associação da E-caderina com o citoesquleto de actina em células HCT-116, mas não em Caco-2. Em células HCT-116, a inibição de Nglicosilação por tunicamicina produziu uma diminuição da atividade de ERK1/2 e a formação de adesão célula-célula foi mais evidente. Estes dados sugerem que alterações na expressão e localização subcelular de diferentes glicanos podem ser importantes eventos associados à perda da estabilidade das JAs em câncer coloretal. / The adherent junction (AJ) is one of the main components of the apical junctional complex. It is a multiprotein complex where E-cadherin, a transmembrane glycoprotein, acts as the main mediator of cell-cell adhesion in epithelium. This protein is anchored to the actin cytoskeleton via proteins of the catenin family. Posttranslational modifications of E-cadherin, such as phosphorylation and glycosylation, can modulate the assembly of AJs. Various studies have suggested that invasion and metastasis is associated to glycan patterns on the cell surface of tumor cells. In colorectal cancer the role of altered glycans expression and stability of E-cadherin-mediated adhesion is not clear. In this study we investigated the relation between changes of the glycans expression and AJs stability in colorectal cancer cells. We used two colon adenocarcinoma cell lines with different metastatic potential, Caco-2 and HCT-116, both models of stable and unstable AJs, respectively. Lectin binding assays demonstrated that in HCT-116, the more invasive cell line, E-cadherin presents a decrease of the glycans recognized by HPA and WGA lectins, which recognize N-acetylgalactosamine and, _-N-acetylhexosamines and sialic acid, respectively. Conversely, in HCT-116, there was an increase of glycans recognized by E-PHA and L-PHA lectins, which recognize bisecting _1,4- branched and _1,6-branched N-acetylglucosamine, respectively. Immunofluorescence assays showed a stronger L-PHA binding on cell-cell contact regions of HCT-116 cells when compared with Caco-2. Furthermore, in HCT-116 cells a complete inhibition of N-glycosylation by tunicamycin or inhibition of complex N-glycans synthesis by swainsonine increased the association of E-cadherin with the actin cytoskeleton. Finally, it was possible to observe that the inhibition of N-linked glycosylation by tunicamycin, leaded to a decreasing of ERK1/2 phosphorylation concomitantly with the formation of more intimate cell-cell contacts. These findings suggest that altered expression and subcellular localization of different glycans can be important events associated to loss of AJs stability in colorectal cancer.

INTRODUÇÃO: O câncer de mama é uma importante causa de morbimortalidade, é conhecido por ser uma doença heterogênea. A caracterização clínica e molecular de seus subtipos é fundamental para nortear o prognóstico e o tratamento dessas pacientes. Novos estudos são necessários na melhor caracterização dos tumores triplo-negativos. O estudo da expressão das claudinas pode auxiliar na caracterização desses tumores. OBJETIVO: Investigar a associação da expressão das claudinas 1, 3, 4 e 7 e E-caderina com variáveis clínico-patológicas e fatores prognósticos, em uma série de tumores de mama triplo-negativos (RE-, RP- e HER2-). MÉTODOS: 80 tumores triplo-negativos foram analisados por imunoistoquímica automatizada para as claudinas 1, 3, 4, 7 e E-caderina. A expressão imunoistoquímica foi avaliada pelo escore H (intensidade multiplicada pela porcentagem de marcação). Foram avaliadas as associações entre características clínico-patólogicas e o escore H. Para a avaliação prognóstica das pacientes, curvas de Kaplan-Meier foram construídas a partir dos dados de seguimento das pacientes e do escore H. RESULTADOS: Foi encontrada associação significativa entre o alto escore H da CLDN-1 (HCLDN-1) e pacientes mais idosas e com a presença de necrose, alto escore H da E-caderina (HE-CAD) em pacientes mais jovens e baixo escore H da CLDN-7 (HCLDN-7) e Ki67 positivo. Além disso, pacientes com elevado HCLDN-1 tiveram menor sobrevida geral. Por outro lado, o elevado HCLDN-3 apresentou uma tendência à associação com maior sobrevida geral e sobrevida livre de doença. CONCLUSÕES: A expressão diferencial das claudinas e E-caderina podem auxiliar na caracterização clinico-patológica dos tumores triplo-negativos. Além disso, as claudinas 1 e 3 parecem ser fatores prognósticos para esses tumores. / INTRODUCTION: Breast cancer is a major cause of morbidity and mortality, is known to be a heterogeneous disease. The clinical and molecular characterization of its subtypes is critical to guide the prognosis and treatment of these patients. Further studies are needed for the best characterization of triple-negative tumors. The study of the expression of claudinas can aid in the characterization of these tumors. OBJECTIVE: To investigate the association of expression of claudinas 1, 3, 4 and 7 and E-cadherin with clinicopathological variables and prognosis in a series of triple-negative breast cancers (ER-, PR- and HER2-). METHODS: 80 triple negative tumors were analyzed by automated immunohistochemistry for the claudins 1, 3, 4, 7 and E-cadherin. The immunohistochemical expression was assessed by H-Score (intensity multiplied by the percentage of staining). We evaluated the associations between clinicopathological characteristics and H-Score. For the prognostic assessment of patients, Kaplan-Meier curves were constructed from the follow-up data of patients and H-Score. RESULTS: We found a significant association between high H-Score of CLDN-1 (HCLDN-1) and older patients and the presence of necrosis, high H-Score of E-cadherin (H-CAD) in younger patients and low H-Score CLDN-7-(7-HCLDN) and Ki67 positive. Furthermore, patients with high-HCLDN1 had a lower overall survival. On the other hand, the high HCLDN-3 showed a trend toward association with greater overall survival and disease-free survival. CONCLUSIONS: Differencial expression of claudins and E-cadherin can help in clinic-pathological characterization of triple-negative tumors. Futhermore, claudin 1 and 3 appear to be prognostic factor for these tumors.

INTRODUÇÃO: O câncer de mama é uma importante causa de morbimortalidade, é conhecido por ser uma doença heterogênea. A caracterização clínica e molecular de seus subtipos é fundamental para nortear o prognóstico e o tratamento dessas pacientes. Novos estudos são necessários na melhor caracterização dos tumores triplo-negativos. O estudo da expressão das claudinas pode auxiliar na caracterização desses tumores. OBJETIVO: Investigar a associação da expressão das claudinas 1, 3, 4 e 7 e E-caderina com variáveis clínico-patológicas e fatores prognósticos, em uma série de tumores de mama triplo-negativos (RE-, RP- e HER2-). MÉTODOS: 80 tumores triplo-negativos foram analisados por imunoistoquímica automatizada para as claudinas 1, 3, 4, 7 e E-caderina. A expressão imunoistoquímica foi avaliada pelo escore H (intensidade multiplicada pela porcentagem de marcação). Foram avaliadas as associações entre características clínico-patólogicas e o escore H. Para a avaliação prognóstica das pacientes, curvas de Kaplan-Meier foram construídas a partir dos dados de seguimento das pacientes e do escore H. RESULTADOS: Foi encontrada associação significativa entre o alto escore H da CLDN-1 (HCLDN-1) e pacientes mais idosas e com a presença de necrose, alto escore H da E-caderina (HE-CAD) em pacientes mais jovens e baixo escore H da CLDN-7 (HCLDN-7) e Ki67 positivo. Além disso, pacientes com elevado HCLDN-1 tiveram menor sobrevida geral. Por outro lado, o elevado HCLDN-3 apresentou uma tendência à associação com maior sobrevida geral e sobrevida livre de doença. CONCLUSÕES: A expressão diferencial das claudinas e E-caderina podem auxiliar na caracterização clinico-patológica dos tumores triplo-negativos. Além disso, as claudinas 1 e 3 parecem ser fatores prognósticos para esses tumores. / INTRODUCTION: Breast cancer is a major cause of morbidity and mortality, is known to be a heterogeneous disease. The clinical and molecular characterization of its subtypes is critical to guide the prognosis and treatment of these patients. Further studies are needed for the best characterization of triple-negative tumors. The study of the expression of claudinas can aid in the characterization of these tumors. OBJECTIVE: To investigate the association of expression of claudinas 1, 3, 4 and 7 and E-cadherin with clinicopathological variables and prognosis in a series of triple-negative breast cancers (ER-, PR- and HER2-). METHODS: 80 triple negative tumors were analyzed by automated immunohistochemistry for the claudins 1, 3, 4, 7 and E-cadherin. The immunohistochemical expression was assessed by H-Score (intensity multiplied by the percentage of staining). We evaluated the associations between clinicopathological characteristics and H-Score. For the prognostic assessment of patients, Kaplan-Meier curves were constructed from the follow-up data of patients and H-Score. RESULTS: We found a significant association between high H-Score of CLDN-1 (HCLDN-1) and older patients and the presence of necrosis, high H-Score of E-cadherin (H-CAD) in younger patients and low H-Score CLDN-7-(7-HCLDN) and Ki67 positive. Furthermore, patients with high-HCLDN1 had a lower overall survival. On the other hand, the high HCLDN-3 showed a trend toward association with greater overall survival and disease-free survival. CONCLUSIONS: Differencial expression of claudins and E-cadherin can help in clinic-pathological characterization of triple-negative tumors. Futhermore, claudin 1 and 3 appear to be prognostic factor for these tumors.

O carcinoma epidermóide da mucosa oral (CEMO) é uma neoplasia maligna comum; no Brasil, são estimados, para 2016, 15.490 novos casos. A invasão óssea ocorre em casos avançados.; esta é classificada em erosiva e infiltrativa. Aparentemente, o processo de transição epitélio-mesenquimal, com o envolvimento da E-caderina, é implicado. Foi investigada a expressão de E-caderina, por meio da imunoistoquímica em 15 casos avançados de CEMO e avaliada sua correlação com as características clínicas e histológicas da invasão óssea. A imunoexpressão da E-caderina foi estudada nos 15 casos de CEMO com evidência histológica de invasão óssea. A maioria dos pacientes eram homens (10 pacientes) e apresentavam invasão em mandíbula (9 casos). A expressão de E-caderina foi negativa em CEMOs com invasão erosiva e positiva nos casos que apresentavam infiltração óssea. A expressão de E-caderina na invasão óssea sugere que a participação do fenômeno de transição epitélio-mesenquimal é um fator diretamente envolvido com o tipo de invasão óssea. / Oral squamous cell carcinoma (OSCC) is a common malignancy; in Brazil it is estimated, in 2016,15.490 new cases. Bone invasion occurs in advanced cases; it is classified in erosive and infiltrative patterns. Apparently, the epithelial-mesenchymal phenomenon, with important participation of E-cadherin is implicated. We investigated the expression of E-cadherin in advanced OSSC and correlated its expression with the clinical characteristics and histologic patterns of bone invasion. Immunoexpression of E-cadherin was studied in 15 cases of OSCC with histological evidence of bone invasion. Most patients were men (10 patients) and presented mandible invasion (9 cases). The expression of E-cadherin was negative in OSCC in erosive bone invasion and positive in the infiltrative bone invasion. E-cadherin expression in bone invasion suggests that participation of epithelial-mesenchymal phenomenon is dependent on the patterns of tumour bone invasion.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / COX-2 e Caderina-E participam de forma fundamental na manutenÃÃo do estado fisiolÃgico da mucosa gÃstrica e tÃm papel essencial na reaÃÃo inflamatÃria e reparo, e no cÃncer. O objetivo deste trabalho à avaliar a expressÃo das duas proteÃnas no carcinoma gÃstrico e metÃstases linfonodais e suas possÃveis participaÃÃes na progressÃo tumoral. Foram utilizados 97 casos de gastrectomias por carcinoma gÃstrico, 36 dos quais com linfonodos disponÃveis, dos arquivos do Hospital do CÃncer do CearÃ. Os casos foram classificados nos tipos intestinal (40 casos), difuso (34), mistos (16) e nÃo-classificados (7 casos) de acordo com a classificaÃÃo de Lauren (1965). Utilizou-se tÃcnica de tissue microarray associada à imunohistoquÃmica com anticorpo monoclonal anti-COX-2 e anti-Caderina-E e sistema de detecÃÃo universal estreptavidina-biotina-peroxidase. A expressÃo de COX-2 foi avaliada de acordo com os seguintes escores: Intensidade (I): 0=negativa; 1=discreta; 2= moderada; 3= acentuada; ExtensÃo (E) de cÃlulas coradas: 1= 0 a 25%; 2= >25 a 50%; 3= >50 a 75%; 4= >75 a 100%. Escore final: I x E, sendo considerados escores < 6 como COX-2 de baixa expressÃo e escores &#8805;6 de alta expressÃo. Classificou-se a expressÃo de Caderina-E nos escores: 0=negativo; 1=citoplasmÃtica; 2=citoplasmÃtica + membranar; 3= membranar-normal (Jawhari et al., 1997a). Foram comparadas expressÃo normal e anormal e membranar e nÃo membranar em cada histotipo de carcinoma, na sede primÃria e linfonodos. ExpressÃo positiva para COX-2 e anormal de Caderina-E predominaram nos diversos histotipos de carcinoma gÃstrico primÃrio, principalmente difusos e mistos. Observou-se maior expressÃo de COX-2 nas metÃstases linfonodais, em relaÃÃo Ãs lesÃes primÃrias, sobretudo nos carcinomas difusos. Carcinomas intestinais estavam associados à expressÃo membranar de Caderina-E enquanto tumores difusos se relacionaram com ausÃncia de expressÃo membranar, o que mostra a importÃncia da Caderina-E na diferenciaÃÃo do cÃncer gÃstrico. Carcinomas gÃstricos apresentam dois padrÃes de imunomarcaÃÃo citoplasmÃtica: granular (paranuclear), associado à expressÃo citoplasmÃtica exclusiva, que prevalece no componente difuso dos tumores mistos, e homogÃneo, em todo o citoplasma, correlacionado com expressÃo citoplasmÃtica-membranar, predominante nos outros histotipos. Carcinomas difusos apresentam expressÃo membranar de Caderina-E, que se expressa com maior freqÃÃncia nas metÃstases linfonodais do que nas lesÃes primÃrias e està presente em grupos celulares infiltrantes e cÃlulas isoladas, nas duas sedes anatÃmicas. Os dados sugerem que o carcinoma misto representa histotipo distinto de carcinoma gÃstrico, baseado nos aspectos peculiares da expressÃo citoplasmÃtica de Caderina-E aqui mostrados e em outros achados da literatura. NÃo houve associaÃÃo estatisticamente significativa entre expressÃo de COX-2 e de Caderina-E e demais parÃmetros clÃnico-patolÃgicos nesta amostra. Os dados aqui observados sugerem que COX-2 e Caderina-E sÃo importantes proteÃnas relacionadas com a progressÃo tumoral-linfonodal no cÃncer gÃstrico / Both COX-2 and E-Cadherin play important roles in physiological and pathological processes in the stomach, such as control of acid secretion, inflammation and cancer. The aim of this study was to analyze the relationship between COX-2 and E-Cadherin immunoexpression in human gastric adenocarcinomas and respective lymph node metastases and their possible action in tumoral progression. Tissue microarrays were prepared from paraffin embedded samples of 97 primary gastric cancers, included 36 with respective nodal metastases. Cases were classified according to Laurenâs classification as intestinal (n=40), diffuse (n=34), mixed (n=16) and undetermined (n=7). Immunoexpression of COX-2 was evaluated regarding intensity (0-absent; 1-mild; 2-moderate; 3-strong) and extension (0-negative or rare cells; 1-<25%; 2-25-50%; 3-50-75%; 4->75% immunoreactive neoplastic cells). A combined score was calculated (intensity x extension): 0-12. A cut-off of 6 was considered to classify COX-2 expression as low (<6) or high (&#8805;6). E-Cadherin expression was evaluated according to the system proposed by Jawhari et al. (Gastroenterology, 1997) as abnormal patterns of expression: 0-no expression; 1-cytoplasmic expression; 2-heterogeneous expression, both membranous and cytoplasmic) and normal membranous pattern (3). Membranous (scores 2 and 3) and Non-membranous (scores 0 and 1) were too compared. Overall, COX-2 positive and abnormal E-Cadherin expression predominate in all types of primary gastric carcinomas. COX-2 expression was higher in lymph node metastases than in primary tumors, with a significant difference for diffuse carcinoma. A positive relationship was observed between E-Cadherin membranous expression and intestinal tumors, and absence of membranous expression and diffuse ones, which indicates the importance of E-Cadherin to gastric cancer differentiation. Granular (paranuclear) cytoplasmic immunostaining pattern was basically associated with cytoplasmic E-Cadherin expression while homogeneous pattern is frequently seen in cytoplasmic-membranous expression. Diffuse carcinomas show membranar expression more frequently in lymph nodes metastases than in gastric primary tumors in both isolated and grouped cells. The data suggest that mixed carcinoma is a distinct hystotype, based on its peculiar cytoplasmic expression of E-Cadherin shown here and other features of literature. There was no significant association linking COX-2 and E-Cadherin expression to other clinicopathological parameters. The data show that COX-2 and E-Cadherin are important proteins related to tumoral progression in gastric cancer

Slug est un membre de la famille des protéines Snail impliquées au cours du développement dans le contrôle de la forme et la différenciation cellulaire. Nous avons localisé Slug au cours du développement des glandes mammaires de souris dans les cellules qui participent aux mécanismes de croissance. Seule une sous-population de cellules épithéliales mammaires située dans le compartiment basal de la glande exprime Slug. Cette expression est maintenue pendant les différentes étapes du développement de la glande mammaire, de la puberté jusqu'au début de la gestation. Nous avons observé une perte d'expression dans lobules se différenciant en alvéoles sécrétoires, ensuite une re-expression au stade d' involution. La population exprimant Slug est positive pour la cytokératine 5, décrit comme un marqueur des cellules basales et myoépithéliales, également considéré comme un marqueur de cellules souches ou progénitrices, et elle est incluse dans la population CD24 positive et surexprimant le CD49, connue pour contenir les cellules souches de l'épithélium mammaire. Nous avons constaté que canaux epithéliaux des glandes mammaires de souris Slug knock-out envahissent moins le coussinet adipeux mammaire. En outre, Ils montrent un phénotype de branchements latéraux, ce qui suggère une différenciation précoce. Ce phénotype ressemble le phénotype des glandes mammaires de souris Knock-out pour la P-cadhérine. Nous avons également constaté une diminution de l'expression de la P-cadhérine in vivo dans les glandes mammaires des souris SlugLaZ, et in vitro, dans les cellules épithéliales mammaires transfectées avec un siARN ciblant Slug. Ces cellules montrent un retard de migration cellulaire. Ces observations valident à notre hypothèse que le facteur de transcription Slug contrôle la différenciation des cellules épithéliales au cours de la croissance physiologique de la glande mammaire murine. / Slug is a member of the Snail protein family involved during development in the control of cell shape and differentiation. We located Slug during mammary gland development in mouse in cells participating to the growth mechanisms. Only a distinct sub-population of mammary epithelial cells located in the basal compartment was found to express Slug. This expression is maintained during the various stages of mammary gland development, from puberty until the beginning of gestation. We observed a loss of expression in lobules differentiating into secreting alveoli, followed by re-expression at involution stage. The Slug expressing population was positive for Cytokeratin 5, described as a basal and myoepithelial cell marker, also considered as a stem/progenitor marker, and was included into the (CD24+ CD49++) population, known to contain the mammary epithelial progenitor cells. We found that mammary gland from Slug-deprived mice were slower to invade the mammary fat pad. In addition, they displayed increased lateral branching, suggesting precocious differentiation. This phenotype resembles the phenotype of mammary glands of P-cadherin Knock-out mice. We also found that P-cadherin is down regulated in vivo in SlugLaZ mice mammary gland, and in vitro, in mammary epithelial cells transfected with an siRNA targeting Slug. These cells show a delay in migration. These observations lead to our hypothesis that Slug controls an early epithelial cell differentiation stage during mammary gland physiological growth.

Les synapses se forment selon plusieurs étapes comprenant la synaptogenèse, la maturation et la plasticité synaptique. Les molécules d’adhésion et les récepteurs ionotropiques du glutamate ont des rôles clés dans ces processus. Lors de ma thèse, je me suis intéressée à la dynamique des interactions impliquant deux protéines membranaires, la N-cadhérine et le récepteur NMDA. La N-cadhérine joue un rôle important dans l’induction de la croissance axonale mais les mécanismes moléculaires sous-jacents sont peu connus. Par des approches de vidéo-microscopie et de pinces optiques, j’ai démontré que la transmission directe des forces générées par le flux rétrograde d’actine aux adhésions N-cadhérines, via les caténines, induit l’avancée du cône de croissance. Les récepteurs NMDA synaptiques ont un rôle crucial dans la maturation et la plasticité synaptique, néanmoins, les mécanismes moléculaires régulant la distribution des récepteurs NMDA synaptiques sont peu connus. En combinant le développement de peptides compétiteurs divalents et des approches d’imagerie haute résolution, nous avons étudié la dynamique de surface des récepteurs NMDA endogènes. Mes résultats montrent que l’interaction dynamique entre les protéines d’échafaudage à domaine PDZ et les récepteurs NMDA contenant la sous-unité NR2A régule leur rétention synaptique et leur distribution de surface. Le déplacement des récepteurs NMDA contenant la sous-unité NR2A en dehors des synapses est compensé par une insertion synaptique de récepteurs contenant la sous-unité NR2B, indiquant que l’ancrage synaptique des différents sous-types de récepteurs NMDA est différentiellement régulé. De plus, cette redistribution des récepteurs NMDA affecte la maturation et la plasticité synaptique. L’ensemble de ces résultats révèle une régulation rapide et spécifique des récepteurs NMDA synaptiques par les protéines à domaine PDZ suggérant un rôle de l’organisation de la densité postsynaptique dans la stabilisation synaptique des récepteurs et les processus adaptatifs. / The formation of synapses follows different steps including synaptogenesis, maturation and plasticity. Adhesion molecules and ionotropic receptors play key roles in these processes. During my thesis, I have been interested in the dynamics of the interactions mediated by two membrane proteins, N-cadherin and the NMDA receptor N-cadherin plays important roles in axon outgrowth, but the molecular mechanisms underlying this effect are mostly unknown. Using live imaging and optical trapping, I demonstrated that the direct transmission of actin-based traction forces to N-cadherin adhesions, through catenin partners, drives growth cone advance. Synaptic NMDA receptors (NMDARs) play key roles during synaptic refinement and plasticity, however the molecular mechanisms that govern the distribution of the synaptic surface NMDARs are largely unknown. We investigated the dynamics of endogenous NMDARs using high-resolution single particle imaging and a newly-developed biomimetic divalent competing ligand. My results show that the dynamic interaction between PDZ domain-containing scaffold proteins and NR2A-NMDARs regulates their synaptic retention and surface distribution. Interestingly, a rapid displacement of NR2A-NMDARs out of synapses is paralleled by a compensatory increase in NR2B-NMDARs, providing functional evidence that the sites of synaptic anchoring of native surface NR2-NMDARs are different. Furthermore, such redistribution of surface NR2-NMDARs strongly impairs synaptic maturation and plasticity. Together, these data reveal a rapid and specific regulation of surface NR2-NMDARs by PDZ domain-containing scaffolds in synapses, supporting a role of the postsynaptic density architecture in regulating specific NR2-NMDAR retention and synaptic adaptation.

Epithelial tissues are composed of a sheet of adherent cells and are present in all metazoans. Their broad function is to compartmentalize tissues and enable the regulated exchange of nutrients and waste between the internal and external environments. To accomplish this function, cells require a specific organization: an apico-basal polarity that provides directionality and intercellular adhesion mediated by adherens junctions that hold cells together. How the epithelia architecture is initiated and maintained remains to be fully elucidated. Adherens junctions and the polarity proteins are functionally linked, as a loss of the main component of AJs: E-cadherin leads to a loss of apico-basal polarity, while disturbing apico-basal polarity results in a re-localization of E-Cadherin. Therefore is challenging to study either pathway in isolation.During my thesis I explored the role of Crumbs, a polarity protein, in the regulation of E-Cadherin in both AJ maturation and maintenance. During maturation of AJs in Drosophila embryo, I demonstrated for the first time by using quantitative high-resolution microscopy PALM that Crumbs regulates E-Cadherin clusters size and their homogenous distribution along the junction. In conclusion, my thesis work provides the first dissection of polarity proteins in E-Cadherin regulation apart from polarity pathways. / Epithelial tissues are composed of a sheet of adherent cells and are present in all metazoans. Their broad function is to compartmentalize tissues and enable the regulated exchange of nutrients and waste between the internal and external environments. To accomplish this function, cells require a specific organization: an apico-basal polarity that provides directionality and intercellular adhesion mediated by adherens junctions that hold cells together. How the epithelia architecture is initiated and maintained remains to be fully elucidated. Adherens junctions and the polarity proteins are functionally linked, as a loss of the main component of AJs: E-cadherin leads to a loss of apico-basal polarity, while disturbing apico-basal polarity results in a re-localization of E-Cadherin. Therefore is challenging to study either pathway in isolation.During my thesis I explored the role of Crumbs, a polarity protein, in the regulation of E-Cadherin in both AJ maturation and maintenance. During maturation of AJs in Drosophila embryo, I demonstrated for the first time by using quantitative high-resolution microscopy PALM that Crumbs regulates E-Cadherin clusters size and their homogenous distribution along the junction. In conclusion, my thesis work provides the first dissection of polarity proteins in E-Cadherin regulation apart from polarity pathways.

No description available.

Melanoma remains an aggressive form of skin cancer with limited treatment options. Novel methods to treat primary tumors and prevent metastatic disease can lead to improved survival for those diagnosed with melanoma. Through this work, we have evaluated the antineoplastic effects of Rhodiola crenulata (R. crenulata) root extracts on B16-F10 melanoma both in vitro and in vivo. In this study, we observed that R. crenulata treatment resulted in an increased cell death as well as a reduced cell growth, proliferation and migration in vitro. Additionally, we observed that R. crenulata decreased the expression of integrin β1 and vimentin, and increased expression of E-cadherin upon in vitro treatment. Further, we observed in a topical R. crenulata based cream therapy, a more radial growth pattern of tumors as well as a reduced mitotic activity and increased tumor necrosis. Markedly, we observed that mice supplemented with R. crenulata orally in their drinking water also displayed reduced establishment of metastatic foci in a disseminated model of melanoma. Collectively, these findings reveal that R. crenulata exhibits striking anti-tumorigenic and anti-metastatic properties, and that this extract may increase survival and harbor potential novel adjuvant therapy for the treatment of melanoma.