La translocation, le passage d'une macromolécule à travers un pore inséré dans une membrane, est impliquée dans de nombreux processus biologiques. On peut citer comme exemple le transport d'ARN ou de protéines entre les composants de la cellule, et l'infection d'une cellule par le passage d'un ADN viral à travers la membrane cellulaire. Aujourd'hui la translocation est aussi la base d'applications technologiques, comme le fait d'utiliser les pores en tant que détecteurs pour le séquençage rapide de molécules ou en tant que filtre moléculaire. La compréhension du processus de translocation est importante à la fois d'un point de vue fondamental et pour la fabrication de nouveaux dispositifs de translocation à usage spécifique. Dans cette thèse, nous réalisons des expériences et des simulations informatiques pour étudier certains des effets les plus importants mis en jeu lors de la translocation.Nous utilisons des simulations informatiques avec un modèle à ``gros grain'' pour étudier qualitativement l'influence d'une interaction attractive entre les parois du pore et un polymère en train de transloquer. Nous montrons que la position de l'interaction influence la fréquence d'entrée et le temps de résidence du polymère dans le pore. La fréquence d'entrée est plus grande lorsque l'entrée du pore est attractive. Le comportement du temps de résidence avec la longueur du polymère est qualitativement et quantitativement affecté par la position de l'interaction dans le pore. Cependant, quelle que soit la position de l'interaction, nous observons que le temps de translocation augmente linéairement avec la longueur du polymère lorsque le polymère est plus long que le pore. Cette observation est qualitativement en accord avec des données expérimentales publiées.Lorsque la translocation est lente, la corrélation entre les mouvements des monomères confinés dans le pore peut jouer un rôle important. Cet effet n'a pas été pris en compte jusqu'à présent. Nous développons un nouveau modèle pour la translocation de polymères, inspiré par le processus d'exclusion asymétrique (ASEP process), qui permet d'étudier spécifiquement cet effet. Nous montrons que les mouvements corrélés des monomères confinés dans le pore génèrent un comportement du temps de résidence avec la longueur du polymère qui est qualitativement similaire à ce qui est habituellement interprété comme la présence d'une barrière d'énergie libre dans le processus de translocation, même lorsqu'une telle barrière n'existe pas. Notre modèle réduit fortement le temps de simulation comparé aux simulations de dynamique moléculaire traditionnelles (quelques secondes contre quelques mois pour un système similaire). Cette accélération provient de l'idéalisation des portions du polymère à l'extérieur du pore. Une telle idéalisation est également présente dans les modèles largement utlisés de type Fokker-Planck, mais dans notre cas le comportement de la partie de la chaîne confinée dans le pore est mieux modélisé.Enfin nous réalisons des expériences pour tester l'existence d'un flot électro-osmotique (EOF) à travers le nanopore d'alpha-hémolysine de staphylococcus aureus. Malgré de nombreux travaux ces dernières années, la question de l'EOF à travers l'un des nanopores biologiques les plus utlisés fait toujours débat. Nous montrons qu'un EOF existe à travers l'alpha-hémolysine et qu'il contrôle la fréquence d'entrée et le temps de résidence de molécules neutres (beta-cyclodextrines) dans le nanopore. La force de l'EOF dépend du type de cation en solution. En particulier nous montrons que l'EOF est plus fort en présence de LiCl que de KCl. / Translocation, the passage of a macromolecule through a pore inserted in a membrane, is involved in many biological processes. Examples include the transport of RNA or proteins between cell components, and the infection of a cell by the passage of a viral DNA through the cell membrane. Today translocation is also the basis of technological applications, such as using pores as sensors for fast molecule sequencing or molecular sieves. The comprehension of the translocation process is important both from a fundamental point of view and for the design of new translocation setups for specific uses.In this thesis both experiments and computer simulations are used to investigate some of the most important effects at work during translocation.Coarse-grained computer simulations are used to study qualitatively the influence of an attractive interaction between the pore walls and a translocating polymer. The location of the interaction is shown to influence both the entry frequency and residence time of the polymer in the pore. The entry frequency is greater when the pore entry is attractive. The behaviour of the residence time with the polymer length is qualitatively and quantitatively affected by the location of the interaction within the pore. Nevertheless, regardless of the location of the interaction, a linear increase of the residence time with polymer length occurs when the polymer becomes longer than the pore. This observation is in qualitative agreement with published experimental data.In the case of slow translocation the correlation between the movements of the monomers confined in the pore may be important. This effect has not been considered previously. A new model of polymer translocation, inspired by the asymmetric exclusion process (ASEP), is developped which enables to specifically investigate this effect. The correlated movements of the monomers confined in the pore are shown to give rise to a behaviour of the residence time with polymer length which is qualitatively similar to what is usually interpreted as the presence of a free-energy barrier in the translocation process, even when such barrier is absent. Our model greatly reduces the simulation time compared to traditional molecular dynamics simulations (several seconds versus several months for similar systems). This speed up comes from the idealization of the portions of the polymer outside the pore. Such idealization is also present in the widely used Fokker-Planck models, but in our case the behaviour of the portion of the chain confined in the pore is better modelled.Finally, experiments are performed to probe the existence of an electro-osmotic flow (EOF) through the nanopore of alpha-hemolysin, from staphylococcus aureus. Despite numerous works during past years, the question of EOF through one of the most commonly used biological nanopores is still under debate. An EOF is shown to exist through alpha-hemolysin and to control the entry frequency and residence time of neutral molecules (beta-cyclodextrins) in the nanopore. The strength of the EOF depends on the type of cations in solution. In particular EOF is shown to be stronger in LiCl solution than in KCl solution.

Reproduction de : Thèse d' exercice : Médecine : Nancy 1 : 2003. / Thèse : 2003NAN11046. Titre provenant de l'écran-titre.

Le lymphome des cellules du manteau (LCM) est un lymphome d’une rare agressivité causée par la translocation chromosomique t(11; 14)(q13; q32) juxtaposant le locus de la cycline D1 (CCND1) sur le chr 11 avec le locus de la chaîne lourde de l'immunoglobuline (IgH) sur le chr 14. En conséquence, une cycline D1 proto-oncogène devient active alors qu’elle n’est pas exprimée dans les cellules-B normales. L’hypothèse initiale semble indiquer une influence directe du fort enhancer IgH sur le promoteur du gène CCND1 afin de surexprimer sa transcription. Quoi qu’il en soit, le locus CCND1 peut être éloigné jusqu'à 200kb du point de cassure du chromosome. Nous avons montré que le locus 11q13 relocalise depuis la périphérie du noyau jusque au centre actif de transcription et au nucléole (Allinne et al., 2014). Ce phénomène qui mène à l’activation du locus entier, suggère un mécanisme epigénétique de régulation des gènes dans les LCM plutôt que simplement un simple effet enhancer-promoteur. Plusieurs nouveaux traitements contre le MCL ont été proposés, y compris les inhibiteurs d’histone deacetylase (HDACis) qui impliquent des mécanismes épigénétiques. Dans les lignées cellulaires LMC, les HDACis sont décrites comme aillant des effets antiprolifératifs, et paradoxalement, le niveau d’expression protéique de la cycline D1 dans la cellule diminue. Jusqu'à présent, il n'y a pas une compréhension claire de ce phénomène, de même que les mécanismes d’action des HDACis reste inconnus. Pour ces raisons, une étude du « paysage épigénétique » sur les loci 11q13 et 14q32 devrait fortement améliorer notre connaissance sur les mécanismes de surexpression de la cycline D1 dans les LMC. L’objectif de ce travail est d'étudier la structure de la chromatine dans le locus réarrangé (11; 14)(q13; q32) dans des cellules LMC par rapport au locus 11q13 et 14q32 dans les lymphocytes humains normaux. Nous avons ensuite étudié l'effet de différentes HDACis sur le locus réarrangé (11; 14)(q13; q32) à plusieurs niveaux: l'acétylation des histones / la méthylation de la chromatine ainsi que sa conformation et l'expression des gènes. Nous avons montré que t(11:14)(q13; q32) conduit à la surexpression de CCND1 avec un groupe de gènes couvrant plus de 15 Mb autour du point de translocation. Les mêmes gènes, sensibles à la dérégulation par la translocation t(11; 14, réagissent au traitement HDACi en augmentant leur expression. Nos résultats indiquent que bien que HDACi stimule la désagrégation de l'hétérochromatine sur l'ensemble du génome, les promoteurs de gènes restent à l'abri de ces effets. / Mantle cell lymphoma (MCL) is a rare aggressive lymphoma caused by the chromosome translocation t(11;14)(q13;q32) juxtaposing the cyclin D1 (CCND1) locus on chr 11 with the immunoglobulin heavy chain (IgH) locus on chr 14. As a result, a proto-oncogene cyclin D1 which is not expressed in normal B-cells, becomes active. The initial hypothesis favored direct influence of the strong IgH enhancer on CCND1 gene promoter to upregulate its transcription. However, the CCND1 locus may be as far as 200 kb from the chromosome breakpoint. We have shown that 11q13 locus relocalizes from the nucleus periphery towards the transcriptionally active center and nucleolus (Allinne et al., 2014). This may lead to activation of the entire locus, suggesting an epigenetic mechanism of gene regulation in MCL, rather than just simple enhancer-promoter effect.Several new treatments are proposed for MCL, including histone deacetylase inhibitors (HDACis) with epigenetic mechanism of action. In MCL cell lines, HDACis were shown to have antiproliferative effects, and paradoxically, they decreased the cyclin D1 protein level in the cells. Until now, there is no clear understanding of this phenomenon, nor of HDACis mechanism of action. Therefore, a study of «epigenetic landscape» in 11q13 and 14q32 loci should significantly advance our knowledge about the mechanisms of cyclin D1 upregulation in MCL.The purpose of the present work was to study chromatin structure in the rearranged (11;14)(q13;q32) locus in MCL cells as compared to the 11q13 and 14q32 loci in normal human lymphocytes. We then studied the effect of different HDACis on the rearranged (11;14)(q13;q32) locus at several levels: histone acetylation / methylation, chromatin conformation and gene expression.We have shown that t(11:14)(q13;q32) translocation leads to overexpression of CCND1 along with a group of genes spanning over 15 Mb around the translocation point. The same genes, sensitive to deregulation by t(11;14) translocation, react to the HDACi treatment by increasing their expression. Importantly, while HDACi stimulates genome-wide disaggregation of heterochromatin, genes’ promoters stay shielded from its effect.

Thesis (MSc)--Stellenbosch University, 1982. / ENGLISH ABSTRACT: No abstract available / AFRIKAANSE OPSOMMING: Geen opsomming beskikbaar

No description available.

No description available.

Acute lymphoblastic leukaemia is the most common childhood cancer with cure rates of approximately 80%. This success can be attributed to the introduction of risk stratification for patients and employment of intensified treatment regimes for patients with high risk disease. However, the identification of prognostically important leukaemia subtypes, unfortunately, is an labour-intensive process. In addition, despite the success in treating childhood ALL, specific subgroups of patients nevertheless still have poor survival rates. This is particularly true for leukaemias characterised by chromosomal translocations involving the MLL oncogene on chromosome 11q23. By using a novel bioinformatics approach, GeneRave, a set of 12 classifier genes (PTPRK, FOS, ENG, Lgal-S1, TCFL5, LRMP, CTGF, IGJ, MX1, PENK, CD3D and HBG1) was selected from a publicly available U95 Affymetrix microarray dataset. Real time PCR carried out on a blinded cohort of 58 primary ALL samples yielded an accuracy of 86%. The absence of PENK gene expression in the majority of ALL samples tested appears to have decreased the overall accuracy. Nevertheless, the results indicate that this method of classification can be easily and quickly performed and therefore may be useful as an adjunct to routinely used methodology in the diagnostic classification of childhood ALL. Parellal screening of a 34,000 chemical small molecules library identified 30 ???hits??? that exhibited specificity toward leukaemia, and many of which shared structural similarity. The cytotoxic effect of these compounds was further investigated in a panel of 19 cell lines that included 3 MLL-translocated (MV411, THP-1 and PER-485), 7 non-MLL-translocated leukaemias (REH, Jurkat, K562, HL60, Hal-01, UOB-B, NB4), 2 immortalized normal blood cell lines, 4 non-leukaemic tumour cell lines (Calu6, MCF7, BE(2)-C, and HeLa) and 3 normal cell lines (HSF, MCF10a and MRC5). In particular, two compounds were identified, SM6 and SM7, that were highly effective at killing MLL-translocated cell lines in the low micromolar range while having little or no effect on the other cell lines. Treatment of PER-485 cells with SM6 and SM7 showed mark down-regulation of the MLL chimaeric fusion protein together with its down-stream targets. One other more broadly acting anti-leukaemia compounds were also identified.

By 2001, elephants had been translocated (mainly from Kruger National Park) to 58 small, fenced reserves in South Africa. All but two introductions took place since 1989. We document important aspects of the population dynamics of elephants in these reserves using data collected in a survey conducted in 2001. The mean population size was 45 elephants, with an average density of 0.25 elephants/km2. Populations have a female bias with 0.79 males to females. Populations have 19% adult males, and 31% adult females. On average, almost 50% of the population comprises adult and subadult females, indicating an immanent potential for large population growth. Births were not significantly different froma 1:1 sex ratio. When two extreme populations were removed, mean mortality rate was 0.4% per annum.Population growth rates averaged 8.3%,but five reserves had growth rates above 13%, and the highest annual growth rate was 16.5% per annum. Twenty-seven populations already have densities above 0.2 elephants/km2, and eight reserves have densities above 0.4 elephants/km2. Assuming a 12% per annum growth (feasible given the data presented), over half the reserves will have densities above 0.33 elephants/km2 within five years. These results indicate that the translocation of elephants has been successful, with most populations reproducing at a rate far exceeding expectations. This has serious implications for owners and managers, as some form of population control (contraception, removals, culling etc.) needs to be urgently planned for implementation as soon as possible in most, and probably all small reserves.

No description available.

Acute lymphoblastic leukaemia is the most common childhood cancer with cure rates of approximately 80%. This success can be attributed to the introduction of risk stratification for patients and employment of intensified treatment regimes for patients with high risk disease. However, the identification of prognostically important leukaemia subtypes, unfortunately, is an labour-intensive process. In addition, despite the success in treating childhood ALL, specific subgroups of patients nevertheless still have poor survival rates. This is particularly true for leukaemias characterised by chromosomal translocations involving the MLL oncogene on chromosome 11q23. By using a novel bioinformatics approach, GeneRave, a set of 12 classifier genes (PTPRK, FOS, ENG, Lgal-S1, TCFL5, LRMP, CTGF, IGJ, MX1, PENK, CD3D and HBG1) was selected from a publicly available U95 Affymetrix microarray dataset. Real time PCR carried out on a blinded cohort of 58 primary ALL samples yielded an accuracy of 86%. The absence of PENK gene expression in the majority of ALL samples tested appears to have decreased the overall accuracy. Nevertheless, the results indicate that this method of classification can be easily and quickly performed and therefore may be useful as an adjunct to routinely used methodology in the diagnostic classification of childhood ALL. Parellal screening of a 34,000 chemical small molecules library identified 30 ???hits??? that exhibited specificity toward leukaemia, and many of which shared structural similarity. The cytotoxic effect of these compounds was further investigated in a panel of 19 cell lines that included 3 MLL-translocated (MV411, THP-1 and PER-485), 7 non-MLL-translocated leukaemias (REH, Jurkat, K562, HL60, Hal-01, UOB-B, NB4), 2 immortalized normal blood cell lines, 4 non-leukaemic tumour cell lines (Calu6, MCF7, BE(2)-C, and HeLa) and 3 normal cell lines (HSF, MCF10a and MRC5). In particular, two compounds were identified, SM6 and SM7, that were highly effective at killing MLL-translocated cell lines in the low micromolar range while having little or no effect on the other cell lines. Treatment of PER-485 cells with SM6 and SM7 showed mark down-regulation of the MLL chimaeric fusion protein together with its down-stream targets. One other more broadly acting anti-leukaemia compounds were also identified.