Outre leur polarité apico-basale, certaines cellules épithéliales développent une seconde polarité, appelée Polarité Planaire Cellulaire (PCP). L'axe de la PCP est orienté perpendiculairement à l'axe de polarité apico-basale et régit l'orientation uniforme de certaines structures, comme les poils ou cils, non seulement à l'échelle de la cellule mais également au sein du tissu. L'épithélium cochléaire est l'un des meilleurs modèles d'étude de PCP chez les mammifères. En effet, les cellules neuro-épitheliales qui le composent, soutenues par des cellules de soutien, présentent à leur apex, des touffes ciliaires dont l'orientation est parfaitement coordonnée par la voie de la polarité planaire. Les deux premiers gènes impliqués dans la PCP chez les mammifères, Vangl2 et Scrib1, ont été identifiés sur la base du phénotype de la cochlée chez les mutants. L'analyse de la localisation de Vangl2 dans l'organe de Corti a également révélé une localisation asymétrique proximo-distale et transitoire de la protéine, perpendiculaire à l'axe apico-basal classique. Cette asymétrie apparaît à la jonction entre deux types cellulaires : une cellule sensorielle ciliée et une cellule de soutien. J'ai pu montrer au cours de mes travaux de thèse que cette asymétrie était majoritairement due à une accumulation de Vangl2 du côté distal des cellules de soutien, et que dans une moindre mesure, Vangl2 pouvait ségréger du côté distal des cellules ciliées. Cette localisation subcellulaire très précise et limitée dans l'espace semble être indépendante de l'expression du gène Scrib1 dans les cellules ciliées. La délétion du gène Scrib1 dans les cellules ciliées m'a toutefois permis de mettre en évidence que ce gène avait un rôle autonome dans la régulation de la PCP, et que les cellules de soutien de l'organe de Corti pouvaient jouer un rôle prépondérant dans le contrôle de la PCP. Mes travaux ont également permis de mettre en évidence que GIPC1 avait un rôle dans la régulation de la PCP et le maintien de l'intégrité des touffes ciliaires des cellules sensorielles, et que le complexe GIPC1/Myosine VI pouvait réguler l'établissement de l'asymétrie de Vangl2 dans l'organe de Corti. / Several epithelia exhibit a second polarity perpendicular to the apico-basal axis, called planar polarity and that governs the orientation of structures such as stereocilia and hear. Our laboratory studies planar polarity, using mammalian cochlear sensory epithelium and we focus our studies on Vangl2, that we identified as the first mammalian planar polarity gene. Vangl2 encodes a four-transmembrane protein that contains a PDZ binding domain in its C-terminus tail. Vangl2 is asymmetrically located at the junction between mechanosensory hair cells and supporting cells, and this asymmetry appears important for planar cell polarity. I have shown in my thesis, using STED microscopy, that Vangl2 asymmetry is mainly due to an accumulation of Vangl2 to the distal side of supporting cells. I sought to dissect the molecular role of Vangl2 by analysing its trafficking within the cochlear epithelium. Deletion analysis shows that the last 12 amino acids, unlike its N-terminus tail are essential for Vangl2 endoplasmic reticulum sorting, its plasma membrane targeting and its function. Conditional mutant mice analysis show that Scrib1, which we have previously shown, interacts with Vangl2 through the PDZ binding domain of its C-terminal tail, is not the protein mediating this asymmetry. My work also highlight that GIPC1 had a role in the regulation of PCP and maintaining the integrity of hair bundles of sensory cells, and that the complex GIPC1/Myosin VI could regulate Vangl2 asymmetry in the organ of Corti.

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Drosophila Fat is a large atypical cadherin molecule. Genetic assays show that Fat has multiple function, however, the mechanism of Fat function is poorly understood. Hence, I undertook a biochemical approach to determine the mechanistic function of Fat. Previous data indicated that Fat might be processed; I further confirmed the precursor-product relationships between these proteins. I then looked at sub cellular localization of Fat. My preliminary data suggests that the smaller 110 kDa forms of Ft goes to the nucleus. To characterize the interaction between Fat and Atro, only known binding partner of Fat, I conducted pull-down assays that indicate Fat has multiple binding sites for Atro. However, the interaction is weak, and different experimental conditions will be needed to characterize the interaction. The only known downstream target of both Fat and Atro in PCP is four-jointed. I provided evidence that fjlacZ1.2kb is regulated by the Ecdysone receptor.

Drosophila Fat is a large atypical cadherin molecule. Genetic assays show that Fat has multiple function, however, the mechanism of Fat function is poorly understood. Hence, I undertook a biochemical approach to determine the mechanistic function of Fat. Previous data indicated that Fat might be processed; I further confirmed the precursor-product relationships between these proteins. I then looked at sub cellular localization of Fat. My preliminary data suggests that the smaller 110 kDa forms of Ft goes to the nucleus. To characterize the interaction between Fat and Atro, only known binding partner of Fat, I conducted pull-down assays that indicate Fat has multiple binding sites for Atro. However, the interaction is weak, and different experimental conditions will be needed to characterize the interaction. The only known downstream target of both Fat and Atro in PCP is four-jointed. I provided evidence that fjlacZ1.2kb is regulated by the Ecdysone receptor.

No description available.

The gene PRICKLE1 is important for human brain function, as mutations in PRCKLE1 are associated with progressive myoclonus epilepsy (PME). Mutations in prickle orthologs could cause seizures in flies, zebrafish and mice, suggesting a conserved role of Prickle protein in seizure from fruit flies to humans. The underlying molecular mechanism how PRICKLE1 mutation causes PME is still unknown. Prickle1 is part of the planar cell polarity (PCP) pathway, which regulates cell polarity within plane of cell sheets. In Drosophila, prickle is recruited to one side of the cell by another PCP protein, Van Gogh. This asymmetric protein localization of Van Gogh/prickle establishes cell polarity. In zebrafish and Xenopus, loss of Prickle or Van Gogh like (Vangl) genes causes PCP phenotypes, which seemingly supports the Prickle/Vangl protein interaction and the role of Prickle in PCP pathway. The function of Prickle in mammals has not been analyzed. It is possible that mammalian Prickle also interacts with Vangl to mediate PCP signaling based on the conserved role of prickle from Drosophila to Xenopus. If Prickle1 interacts with Vangl and regulate PCP pathway, the PME we observed in humans might be associated with loss of neuronal polarity and impaired neuron activity. Therefore, to understand whether Prickle1 mediates Vangl signaling in mammals could be a step toward revealing the etiology of PME in human patients. Therefore, I analyzed the function of Prickle1 in three developmental processes, the limb development, the palate development, and the caudal migration of facial branchimotor neurons (FBMs), in which the function of PCP pathway, especially Vangl2, has been described. Supporting the interaction between Prickle1 and Vangl, mutations in either Prickle1 or Vangl2 leads to shorter limbs. However, Prickle1 and Vangl2 only have limited overlap in mRNA expression in the digit tips. This raises the question as to how impaired Prickle1/Vangl2 protein interaction in these cells in the digit tips cause defective growth of the whole limb. It also suggests alternate function of Prickle1 other than mediating Vangl2 function. This interaction between Prickle1 and Vangl2 is further challenged by the limited function of Vangl but the essential role of Prickle1 in palate development, which suggests that the function of Prickle1 is independent of Vangl2. In the caudal migration of FBMs, Prickle1 mutation impairs this migration process dose-dependently. This is different from Vangl2 mutation, which completely blocks the caudal migration and partially impairs the lateral migration of FBMs. More importantly, Prickle1 is expressed by the neurons, while Vangl2 functions in the surrounding cells, which again raises the question as to whether and how the two proteins could interact if they are not expressed in the same cell. These results together question the model that Prickle1 is the intracellular partner of Vangl2, but support Prickle1 function might be independent of Vangl. Actually, it is possible that Prickle1 is part of gene expression regulation machinery: Prickle1 mutation affects Wnt5a expression in the limb and Shh expression in the palate. Although this regulation mechanism is still unknown, it suggests that defective gene expression might be related to PME caused by PRICKLE1 mutation.

In this study, the phytoremediation techniques are used to treat the soil contaminated by pentachlorophenol(PCP).First, four plants species were selected,including Allium tuberosum, Vigna radiata (L.) Wilczek, Pennisetum alopecuroides, and Medicago sativa to compare their treatment efficiencies for PCP in soil.The experimental results showed that the species of Allium tuberosum presented the highest degradation rate 76% after 35-day test run with the initial concentration of 20mg/kg in soil. In the second stage,the species of Allium tuberosum was thus selected to run the tests of feasibility of using phytoremediayion to treat the soils contaminated byPCP.During the e xperiment,the pot tests inside a greenhouse were run for 330 days.The result indicated that the species of Allium tuberosum contributed to the increase of microorganism and dehydrogenase activity in the soil. Bisides,we also found that adding with nutrients could help Allium tuberosum to depress the PCP stress.The test with vegetation of Allium tuberosum and addition of nutrients showed that the PCP degradation rate was measured equal to 98.4% with the concentration of PCP degraded from 42mgkg-1 to 0.68mgkg-1 after 330days. Finally, molecule biotechnology of PCR-DGGE was applied to the test of observing the microbiota in the soils.According to the test results,we found that the diversity of microorganisms could be raised through planting the species of Allium tuberosum. The microbiota in the soils with PCP pollutant have more varieties than the microbiota in soils without vegetation, which was infered that the addition of PCP might stimulate the vitality of microbes in the soils. Moreover, comparing the microbiota on rhizosphere of the plant species and in the bulk soils, it was found that the actitivies of root exudates might be able to increase the varieties of rhizospheric microorganisms.

Pentachlorophenol (PCP) is a chloride-containing aromatic compound which is mostly used for preserving wood and leather, but still one can easily detect this compound present in the waste water generated by various industries such as petrifaction, oil-refining, and etc. PCP, due to its chemical property of being stable and highly toxic, would cause severe and irreparable environmental pollution once exposed to open air. This study is intended to explore the feasibility of dealing the problem of PCP with biodegradation. The examination results showed that, except for absorption, the suspension of contaminated soil (aerobic incubation), nonetheless, could effectively degrade PCP during a period of 90 days without the aid of any extra carbon source. (0.62 mg/L/day). The degradation rate was further greatly improved by adding sodium acetate, molasses, and sludge cake (sodium acetate added: 4.15 mg/L/day; molasses added: 1.05 mg/L/day; sludge cake added:0.83 mg /L/day). None of four experimental groups of aerobic sludge, anaerobic sludge, contaminated soil (anaerobic incubation), and Fe3+reaction could degrade PCP after 135 days, 174 days, 250 days, and 124 days, respectively, regardless of whether any sources of carbon were added or not. A bacterium which used PCP as the sole carbon source was isolated from the contaminated soil. After 16s rDNA sequence analysis, it had 98% degree of similarity to Pseudomonas mendocina and was designated as Pseudomonas mendocina NSYSU. The PCP (40 mg/L) degradation rate of Pseudomonas mendocina NSYSU was 9.33 mg/L/day, and the degradation rate would slow down as PCP concentration increased. At a PCP concentration of 320 mg/L, PCP degradation was completely inhibited, although an active population of Pseudomonas mendocina NSYSU was still present in these cultures. The study also indicated that the addition of various carbon sources such as sodium acetate and glucose did not facilitate the degradation of PCP with the degradation rate of 8.11 mg/L/day for sodium acetate, and that of 7.55 mg/L/day for glucose. Analysis from examining several environmental factors showed that the optimal condition for PCP degradation is that of 30¢J, pH6, and in the presence of oxygen. The end products of PCP degradation were detected by GC-MS. After 6 days of incubation, PCP was gradually disappeared and the metabolic intermediate product, acetic acid was detected. The chloride ion concentration also increased by 21.8 mg/L, which is approximately equal to the original total chloride content in PCP (66% of chloride content). In conclusion, PCP could be effectively and completely degraded by Pseudomonas mendocina NSYSU.

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