University, Diss., 2003--Kassel. / Erscheinungsjahr an der Haupttitelstelle: 2002.

Universiẗat, Diss., 2006--Kassel.

No description available.

No description available.

During multicellular development, cells must make fate decisions that reproducibly generate the correct cell type proportions. It is remarkable that in certain developmental scenarios, seemingly identical cells in a homogenous environment can achieve this. It is thought that this is possible because cell populations exhibit reproducible cell-cell variation in gene expression. How these differences are generated has been intensely studied over the past decade, with transcriptional bursting emerging as an important factor for driving variability between cells. Furthermore, it is thought that chromatin structure around gene promoters is a key regulator of transcriptional bursting. However, key questions remain. What factors regulate chromatin structure at the molecular level? Is the activity of chromatin regulators governed by random processes or entrained by one of many hidden factors such as cell cycle positioning, cell volume, metabolism? Are the proportions of cells exhibiting different bursting patterns regulated to ensure normal cell fate choice and proportioning? To address these questions, we have investigated whether different regulators of chromatin structure affect the pre-stalk/pre-spore fate decision in the social amoebae D. discoideum. We have identified that set1, a methyl-transferase responsible for generating methylation on histone 3 at position lysine 4 (H3K4me), plays a key role in controlling the balance of cell types in multicellular development as in its absence cells become autonomously primed towards a pre-stalk fate. Single cell RNA-sequencing has revealed that genes normally regulated by this modification represent a specific class of hyper-variable genes. We find that this variability is generated by specific set1 dependent repression at these loci, as upon deletion of this enzyme we see an active recruitment of more cells to an expressing state. Our data suggest that set1 activity itself is controlled by the external source of the cell cycle. This cell cycle dependent regulation robustly ensures the correct proportions of cells within the population contain levels of set1 activity that prime 25% of cells towards the pre-stalk lineage and the other 75% to the pre-spore fate. As such we believe our study reveals a novel mechanism linking specific regulation of transcriptional bursting through the activity of set1 to cell fate propensity.

Les amibes se nourrissent principalement de bactéries. Certaines bactéries pouvant résister à la digestion dans la voie phagocytique amibienne s’y retrouvent enrobées et sécrétées dans l’environnement par la suite. Cet enrobage augmente la résistance bactérienne à différents stress. Les bactéries enrobées pourraient favoriser la propagation de maladies infectieuses, mais aucune donnée n’est disponible à ce sujet. Les bactéries pathogènes Escherichia coli O157:H7, Salmonella typhimurium et Pseudomonas aeruginosa sont capables de résister à la digestion amibienne, mais leur capacité à être enrobée est inconnue. L’étude de la résistance de ces bactéries face à l’amibe Dictyostelium discoideum et l’analyse de la viabilité de cette dernière en présence des bactéries ont été réalisées pour mieux cibler les souches bactériennes potentiellement enrobables. Des essais d’enrobage ont été tentés, mais aucune bactérie enrobée n’a été observée en microscopie électronique. D’autres analyses seront requises pour comprendre la propagation des maladies infectieuses via les bactéries enrobées. / Amoebae mainly feed bacteria. Many bacteria are resistant to the digestion in the phagocytic pathway of amoebae. There they can be packaged and then secreted into the environment. Packaging increases the resistance of bacteria to different stresses. Packaged bacteria could improve the spread of infectious diseases, but no data is available regarding that so far. The pathogenic bacteria Escherichia coli O157:H7, Salmonella typhimurium and Pseudomonas aeruginosa are able to resist to digestion by amoeba digestion, but their ability to be packaged is unknown. The study of the bacteria resistance to the amoeba Dictyostelium discoideum and the analysis of the viaibility of the latter in the presence of bacteria were done to better target the bacterial isolates that can be packaged. Assays of packaging of bacteria were done, but no packaged bacteria were observed by electron microscopy. Other analyzes are required to understand the spread of infectious diseases by packaged bacteria.

L'amibe Dictyostelium discoideum sécrète des corps multilamellaires (CMLs) à partir de sa voie endocytique lorsqu'elle est cultivée en présence de bactéries. L'objectif de la présente étude consistait à comprendre le rôle des CMLs dans la physiologie de D. discoideum en caractérisant l'antigène de l'anticorps H36 présent sur les CMLs et celui de l'anticorps H72 qui se retrouve dans la voie endocytique. L'identification des antigènes a été réalisée par une immunoprécipitation suivie par une analyse en spectrométrie de masse. L'identité de l'antigène H36 a été déterminée et a été confirmée par des analyses en Western blot pour être la protease cprG aussi nommée CP7. Dans le cas de l'anticorps H72, l'identification n'a pas été fructueuse. Cette étude démontre que les CMLs sécrétés contiennent donc la protease cprG. Il s'agit de la deuxième description de l'association d'une protease avec des structures membranaires sécrétées par l'amibe D. discoideum.

Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, 1982 / Vita. / Includes bibliographical references. / by Charles Joseph Reynold Verret. / Ph. D. / Ph. D. Massachusetts Institute of Technology, Department of Chemistry

No description available.

PakB is a p21-activated kinase that phosphorylates and activates class I myosins in the social amoeba Dictyostelium discoideum. PakB co-localizes with myosin-I to actin-rich regions of the cell, including macropinocytic cups and the leading edge of migrating cells. Here we show that the cellular localization of PakB depends on the N-terminal region which contains an actin filament binding module and two proline-rich motifs that interact with the SH3 domain of actin-binding protein 1 (dAbp1). dAbp1 co-localized with PakB to actin-rich sites, but in PakBˉ (PakB null) cells dAbp1 adopted a diffuse cytosolic distribution. Overexpression of dAbp1 in PakBˉ cells produced SH3 domain-dependent defects in early development, cell polarization and chemotaxis. We conclude that PakB plays a critical role in regulating the cellular functions of the dAbp1 SH3 domain. PakBˉ cells exhibited a disrupted cortical actin layer and were extremely sensitive to external stresses induced by compression and electroporation. PakBˉ cells showed severe chemotaxis defects when forced to migrate under agarose. The defects were rescued by expression of full-length PakB but not an N-terminal fragment of PakB. The results suggest that loss of PakB kinase activity is responsible for the cortical defects. Immunoblot analysis showed that phosphorylation of MyoD at the TEDS site was significantly reduced in PakBˉ cells. We propose that activation of myosin-I motor activity by PakB plays a critical role in stabilizing the cortical actin cytoskeleton. D. discoideum myosin-II heavy chain kinase A (MHCK-A) is a member of the alpha-kinase family. Competition experiments with Mant-ADP showed that MHCK-A bound ATP with Ki values of 18 and 160 µM in the presence and absence of Mg2+, respectively. ADP and AMP bound 3-fold and 9-fold more weakly than ATP, respectively. The results show that Mg2+ and the nucleotide phosphoryl groups substantially contribute to binding. Mutations of residues in the Pi-pocket and N/D-loop reduced the binding affinity for MgATP, showing that both regulatory sites are coupled to the active site. Phosphorylation of SPOT peptide arrays with MHCK-A revealed a consensus sequence of T-φ/K-φ/K-K/R and showed also phosphorylation of non-Thr-containing peptides. / Thesis (Ph.D, Biochemistry) -- Queen's University, 2013-08-21 14:02:30.015