The present thesis investigates the hypothesis according to which the arithmetic retrieval deficits observed in children with math disabilities (MD) would be due to an inhibition deficit. In the first chapter, two experiments showed that children with MD (with or without reading disabilities), compared to normally-achieving children, do not present impairments in three inhibition functions (filtering, suppression, and blocking). The second chapter focused on interference in arithmetic tasks. Experiment 3 revealed that, in a multiplication verification task, children with MD were not more sensitive to interference than control children (of the same age or of the same math skills). In contrast, Experiment 4 showed that children with poor math skills were more sensitive to multiplication-related interference (i.e., the negative effect of multiplications on additions) than children with good math skills. Nevertheless, the third chapter established that the arithmetic retrieval deficits of children with MD (as well as the results of Experiment 4) can be accounted for without the recourse to inhibition. Indeed, Experiment 5 demonstrated that children with MD have poor memory representations of difficult single-digit multiplications (i.e., weak and incorrect problem-answer associations), which is sufficient to account for their retrieval deficits. Finally, in the last chapter, we considered the possibility that sensitivity to interference is involved in MD but during the development of arithmetic facts representations (not during their retrieval) and could lead to the poor representations observed in children with MD. In experiment 6, we found that counting to solve an addition might interfere with the memorization of the addition's addends (hence, with the development of problem-answer associations) but there was no evidence that children with MD are more sensitive to this interference than control children. Altogether, these data provide converging evidence against the inhibition-deficit hypothesis and suggest that poor arithmetic representations represent a better candidate as a causal factor of MD. / Le but de cette thèse est d'investiguer l'hypothèse d'un rôle causal des processus d'inhibition dans les Troubles d'Apprentissage de l'Arithmétique (TAA). Selon cette hypothèse, les difficultés de récupération des faits arithmétiques (p.ex., les tables de multiplications) observés dans les TAA seraient dues à des déficits d'inhibition. Le premier chapitre présente deux expériences dans lesquelles nous avons testé trois fonctions d'inhibition (filtrage, suppression et blocage) chez des enfants avec TAA global (avec et sans troubles de lecture associés) et avec troubles spécifiques de récupération de faits arithmétiques (Expériences 1 et 2). Les résultats de ces études n'ont pas mis en évidence de trouble d'inhibition chez ces enfants. Le second chapitre s'est focalisé sur les effets d'interférence classiquement observés dans des tâches arithmétiques. Dans l'expérience 3, nous avons mis en évidence un effet de confusion associative (p.ex., 3 x 7 est plus difficile à rejeter que 3 x 7 = 26) significatif chez des enfants avec troubles de récupération de faits arithmétiques. Néanmoins, cet effet était équivalent à celui observé chez des enfants contrôle. En revanche, dans l'expérience 4, nous avons observé que l'effet interférent de la multiplication sur l'addition était plus important chez des enfants avec faibles capacités de récupération de faits arithmétiques que des enfants avec fortes capacités de récupération. Toutefois, dans le troisième chapitre, nous démontrons que ces derniers résultats, et plus globalement, les troubles de récupération de faits arithmétiques, peuvent être interprétés à la lumière de modèles théoriques qui n'incluent pas de processus d'inhibition. En effet, l'expérience 5 révèle que chez des enfants avec de tels troubles de récupération (en comparaison d'enfants contrôle), les multiplications difficiles (p.ex., 6 x 7) étaient plus faiblement associées à leurs réponses correctes et associées à de réponses incorrectes, ce qui, selon certains modèles théoriques, était suffisant pour rendre compte de déficits de récupération. Ces résultats vont à l'encontre de l'hypothèse de déficits d'inhibition comme facteur causal des troubles de récupération arithmétique dans les TAA. Cependant, dans le dernier chapitre, nous considérons la possibilité que la sensibilité à l'interférence jouait un rôle important dans le développement des représentations arithmétiques (et non dans leur récupération). Dans l'expérience 6, notre hypothèse était que compter pour résoudre une addition pouvait interférer avec la mémorisation des opérandes de cette addition et donc, avec la formation des associations entre problème et réponse correcte. Si les résultats obtenus allaient dans ce sens, nous n'avons trouvé aucune évidence indiquant que cet effet interférent était plus important chez des enfants avec troubles de récupération de faits arithmétiques que chez des enfants contrôle. En conclusion, il semble que de pauvres représentations des faits arithmétiques, plutôt que des déficits d'inhibition, soient à l'origine des troubles de récupération de ces faits dans les TAA.

Branched tubular organs, such as the lung and vascular system fulfill the respiratory needs of most animals. Optimal tissue function relies on the uniform sizes and shapes of the constituting branches in each organ. The Drosophila tracheal airways provide a recognized genetic model system for identification and characterization of tube size regulators. We found that the programmed secretion and assembly of the apical extracellular matrix (ECM) is required for the expansion of the trachea and salivary glands (SG) tubes. We have characterized Vermiform (Verm) and Serpentine (Serp), two chitin-binding proteins with predicted polysaccharide deacetylase domains (ChLDs). Verm and Serp mutants show overelongated tubes, suggesting that luminal ECM modification restricts tracheal tube elongation. The luminal deposition of ChLDs, but not other secreted components, depends on paracellular septate junction integrity (SJs) in the tracheal epithelium. Deletion of the deacetylase domain renders Serp-GFP intracellular, arguing that the deacetylase domain harbors uncharacterized secretion signals. To explore this possibility we transferred the deacetylase domain from Serp to Gasp, another tracheal luminal protein, which requires the Emp24 adaptor for ER exit. The Gasp-Deac-GFP chimera was normally secreted in emp24 mutants indicating that the deacetylase domain contains potential ER-exit signals. To identify such signals we characterized conserved sequence motifs in the Serp deacetylase domain. Mutations of the N-glycosylation sites gradually reduced Serp-GFP luminal deposition suggesting that increased glycosylation enhances apical Serp secretion. By contrast, substitutions in three conserved amino acid stretches completely blocked the ER-exit of Serp-GFP. The mutated proteins were N-glycosylated suggesting that the motifs may be involved in a subsequent protein-folding step or facilitate ER exit through interactions with unidentified specific adaptors. / At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Epithelial cells called enterocytes line the lumen of the small intestine and are responsible for nutrient processing and barrier maintenance. Enterocytes have highly ordered actin arrays, or brush borders, on their apical surfaces. The brush border is composed of microvilli, membrane based protrusions of parallel actin bundles. Within microvilli, myosin-1a laterally links the actin cytoskeleton to the overlying membrane, and contributes to membrane tension regulation and vesicle shedding. These physiological functions require proper localization of this motor, a process that depends on the membrane binding tail homology 1 (TH1) domain. The goal of this thesis is to provide mechanistic details as to how myosin-1a targets to microvillar membrane and how its cellular dynamics are controlled. We find that in vitro and in cells myosin-1a interacts electrostatically with phosphatidylserine through basic residues in two independent bona fide membrane binding motifs. Because membrane binding controls myosin-1a targeting and previously published solution kinetic studies show the motor/actin interaction is short lived, we hypothesized that TH1 is the master regulator of dynamics for this molecule. We used live cell single molecule total internal reflection fluorescence microscopy in combination with single particle tracking and mean squared displacement analysis to measure membrane bound lateral mobility for myosin-1a and TH1. Many myosin-1a molecules display long-lived low mobility dynamics. Similar events are absent from TH1 analysis, indicating the motor domain makes an unexpected contribution to limiting mobility of myosin-1a at the membrane/cytoskeletal interface. Structure/function analysis confirmed this result and revealed the neck region is also important to controlling myosin-1a dynamics. In the context of full length myosin-1a, the neck region also plays a role in regulating localization, perhaps through a conformational change that involves calmodulin/calcium interactions. This is the first study to examine live cell dynamics for any class I myosin at single molecule resolution. The results presented within this thesis provide novel insight as to how myosin-1a cellular targeting and dynamics are controlled, and how biochemical and biophysical properties of myosin-1a manifest in cells to help this molecule carry out physiological roles in the brush border.

The Notch and TGF13 signaling pathways have been shown to play important roles in regulating endothelial-to-mesenchymal transition (EndMT) during cardiac morphogenesis. EndMT is the process by which endocardial cells of the atrioventricular canal and the outflow tract repress endothelial cell phenotype and upregulate mesenchymal cell phenotype. EndMT is initiated by inductive signals emanating from the overlying myocardium and inter-endothelial signals and generate the cells that form the heart valves and atrioventricular septum. The Notch and TGFf3 pathway are thought to act in parallel to modulate endothelial phenotype and promote EndMT. Vascular endothelial (VE) cadherin is a key regulator of cardiac endothelial cell phenotype and must be downregulated during EndMT. Accordingly, VE-cadherin expression remains stabilized in the atrioventricular canal and outflow tract of Notchl-deficient mouse embryos, while activation of the Notch or TGFP pathways results in decreased VE-cadherin expression in endothelial cells. However, the downstream target gene(s) that are involved in regulating endothelial cell phenotype and VE-cadherin expression remain largely unknown. In this thesis the transcriptional repressor Slug is demonstrated to be expressed by the mesenchymal cells and a subset of endocardial cells of the atrioventricular canal and outflowtract during cardiac morphogenesis. Slug is demonstrated to be required for cardiac development through its role in regulating EndMT in the cardiac cushion. Data presented in Chapter 6 further suggests that Slug-deficiency in the mouse is compensated for by a increase in Snail expression after embryonic day (E) 9.5, which restores EndMT in the cardiac cushions. Additionally, the Notch pathway, via CSL, directly binds and regulates expression of the Slug promoter, while a close Slug family member, Snail is regulated by the TGFB pathway in endothelial cells. While Notch does not directly regulate Snail expression, Notch and TGFB act synergistically to regulate Snail expression in endothelial cells. It is further demonstrated that Slug is required for Notch mediated EndMT, binds to and represses the VE-cadherin promoter, and induces a motile phenotype. Collectively the data demonstrate that Notch signaling directly regulates Slug, but not Snail, expression and that the combined expression of Slug and Snail are required for cardiac cushion morphogenesis.

Epithelial-to-Mesenchymal Transformation (EMT) is an important process in development, and occurs during key steps in both valvular and coronary vessel development. This dissertation uses transcriptional profiling strategies to identify novel regulators of EMT during cardiogenesis. An early step in valvulogenesis occurs when endocardial cells overlaying the cushions of the atrioventricular canal (AVC) and outflow tract (OFT) undergo EMT to yield valvular interstitial cells. We developed an unbiased strategy to identify genes important in endocardial EMT using a spatial transcriptional profile. RNA-seq analysis of gene expression between AVC, OFT, and ventricles (VEN) isolated from chick (HH18) and mouse (E11.0) embryos at comparable stages of development was performed. EMT occurs in the AVC and OFT cushions but not the VEN at this time. We identified genes >2-fold enriched in chick and mouse cushions compared to VEN. Gene ontology and Gene Regulatory Network (GRN) analysis of cushion-enriched genes were consistent with cells undergoing EMT. Further analysis accurately identified and validated previously unrecognized novel candidate genes (Meis2, Id1, Hapln1, Foxp2) and the NF-κB pathway as regulators of endocardial cell EMT in vitro. Epicardial EMT is a critical step in coronary vessel formation which is dysregulated in mice lacking TGFβR3. To elucidate the role of TGFβR3 in EMT we developed a strategy to identify genes downstream of TGFβR3 in cultured epicardial cells. Tgfbr3+/+ and Tgfbr3-/- immortalized epicardial cells were incubated with vehicle or ligands known to promote TGFβR3-dependent invasion (TGFβ1, TGFβ2, BMP2) and harvested for RNA-seq analysis. GRN analysis of genes >2-fold differentially expressed between Tgfbr3+/+ and Tgfbr3-/- cells in each ligand incubation group revealed dysregulated NF-ĸB signaling. TGFβ2 or BMP2 incubation stimulated NF-ĸB activity in Tgfbr3+/+ but not Tgfbr3-/- epicardial cells. Inhibiting NF-ĸB signaling reduced TGFβ2- or BMP2-promoted invasion of Tgfbr3+/+ cell, further supporting a role for NF-ĸB signaling in promoting invasion downstream of TGFβR3. The genes and signaling pathways identified through our analysis yield the first comprehensive list of candidate genes whose expression is dependent on TGFβR3 signaling. These transcriptional profiling strategies identified and validated novel regulators of endocardial and epicardial EMT.

The molecular mechanisms that regulate survival of embryonic neural precursors are still relatively ill-defined. Here, we have asked whether the p53 family member p63 plays any role during this developmental window, focusing upon the embryonic cerebral cortex. We show that genetic knockdown of p63 either in culture or in the embryonic telencephalon causes apoptosis of cortical precursors and newly-born cortical neurons, and that this can be rescued by expression of ΔNp63, but not TAp63 isoforms. This cortical precursor apoptosis is the consequence of deregulated p53 activity, since both basal precursor apoptosis and that induced by loss of p63 are rescued by coincident genetic silencing of p53. Finally, we demonstrate that the third p53 family member, ΔNp73, does not regulate survival of cortical precursor cells, but that it collaborates with ΔNp63 to ensure the survival of newly-born cortical neurons. Thus, the balance of ΔNp63 versus p53 determines the life versus death of embryonic cortical precursors. To assess if p63 plays a similar role in the adult brain, we examined mice haploinsufficient in p63 (p63+/-); we observed that aging, but not young, mice show deficits in short term memory and vertical exploratory behaviour. These results establish a foundation upon which the role of p63 in aging can be further characterized. Given the importance of p73 in aging and in preventing neurodegeneration, we aimed to create a conditional p73 knock out mouse (floxed p73). This mouse will allow for lineage-specific characterization of p73 function and will circumvent the reduced life span displayed by 4 mice lacking p73 (p73-/-). The work presented in this thesis has advanced our knowledge on the role of the p53 family members (p53, p63, and p73) in regulating survival and death in the nervous system. This knowledge will be important to understand neuropathology and to design appropriate therapeutic interventions.

The purposes of this dissertation were to: (1) formulate and develop a critical thinking/critical reading model based on professional opinion and the review of the literature in these areas; (2) create a critical thinking/critical reading appraisal for grades three through six; and (3) validate the critical thinking/critical reading appraisal instrument.The first step in the development of the Worden Critical Thinking/Critical Reading Model was to review the literature and related research in the area of critical thinking, critical reading and language development. A "first draft" of the model with an accompanying explanation of it was submitted to selected faculty members in the areas of elementary education and reading. Once this model was evaluated, further developed and refined it was sent to eighty authorities across the country for a more extensive evaluation. These authorities represented the areas of reading diagnosis, evaluation and testing, cognitive processes, linguistics and language development.Once the model was developed by this author and the sub-skills of critical reading identified, the Worden Critical Thinking/Critical Reading Appraisal was begun. Both the passages and test items were written on reading levels ranging from second reading level to sixth reading level. All these test items with accompanying passages were submitted to a predetermined panel of judges composed of university faculty members and reading specialists. These judges were asked to answer each test question, evaluate the clarity of the compositions and questions, and indicate the skill they felt was being measured.The passages and questions were then formed into the Worden Critical Thinking/Critical Reading Appraisal and administered to a population of students in grades three through six. The results of the first administration were statistically analyzed and a second form of the Worden Critical Thinking/Critical Reading Appraisal was created. This also was administered to a population of third, fourth, fifth and sixth grade students.Data from the validation procedures for both appraisal forms were analyzed to determine the validity (face, content, and. construct; and reliability. An item analysis was also implemented on each roam of the appraisal to determine which items would be retained in a final appraisal form.The Worden Critical Thinking/Critical Reading Appraisal was found to have a test reliability ranging from .80 to .84. The significant finding is that critical thinking and critical reading as defined in this study can be measured using a paper and pencil format. It can not be concluded thus far that this appraisal can serve as a diagnostic instrument to measure specific strengths and weaknesses in the skills of critical reading as identified in the author's model.Based on the findings of this study and subject to its stated limitations, the following conclusions seem warranted:1. A model of critical thinking and critical reading can be designed and developed.2. As a result of this model, certain critical thinking processes and critical reading skills can be identified and defined.3. The general ability to think and read critically can be measured in students with reading levels ranging from three through six.4. Individual sub-skills of critical reading cannot be reliably measured by the Worden Critical Thinking/Critical Reading Appraisal.5. A model of critical reading with separate sub-skills may not be appropriate. These sub--skills, s illustrated in the findings, are difficult, to measure as separate entities.

The molecular mechanisms that regulate survival of embryonic neural precursors are still relatively ill-defined. Here, we have asked whether the p53 family member p63 plays any role during this developmental window, focusing upon the embryonic cerebral cortex. We show that genetic knockdown of p63 either in culture or in the embryonic telencephalon causes apoptosis of cortical precursors and newly-born cortical neurons, and that this can be rescued by expression of ΔNp63, but not TAp63 isoforms. This cortical precursor apoptosis is the consequence of deregulated p53 activity, since both basal precursor apoptosis and that induced by loss of p63 are rescued by coincident genetic silencing of p53. Finally, we demonstrate that the third p53 family member, ΔNp73, does not regulate survival of cortical precursor cells, but that it collaborates with ΔNp63 to ensure the survival of newly-born cortical neurons. Thus, the balance of ΔNp63 versus p53 determines the life versus death of embryonic cortical precursors. To assess if p63 plays a similar role in the adult brain, we examined mice haploinsufficient in p63 (p63+/-); we observed that aging, but not young, mice show deficits in short term memory and vertical exploratory behaviour. These results establish a foundation upon which the role of p63 in aging can be further characterized. Given the importance of p73 in aging and in preventing neurodegeneration, we aimed to create a conditional p73 knock out mouse (floxed p73). This mouse will allow for lineage-specific characterization of p73 function and will circumvent the reduced life span displayed by 4 mice lacking p73 (p73-/-). The work presented in this thesis has advanced our knowledge on the role of the p53 family members (p53, p63, and p73) in regulating survival and death in the nervous system. This knowledge will be important to understand neuropathology and to design appropriate therapeutic interventions.

A dense array of parallel actin-based protrusions called microvilli extend from the apical surface of intestinal epithelial cells (IECs), collectively called the brush border. In addition to serving as the sole site of nutrient absorption, this domain also contains host defense proteins. As the brush border is located at the interface between intestinal contents and tissue it must prevent translocation of pathogens and toxins. However, little is known about its role in host defense. Using a combination of cell biological and microscopy techniques, we asked how the brush border responds to microbial infection. Microvilli release vesicles laden with the host defense enzyme intestinal alkaline phosphatase (IAP) into the intestinal lumen. Here, we find that IAP on these lumenal vesicles (LVs) is biochemically active, able to detoxify a bacterial toxin and limit inflammation. Independent of IAP activity, LVs inhibit attachment of enteropathogenic E. coli (EPEC) to IECs. LVs also limit growth of bacteria, while the presence of microbes stimulates increased LV shedding. Thus, LVs represent a multi-faceted form of host intestinal defense. When EPEC do reach the host cell surface, the brush border is dramatically remodeled, resulting in microvillar effacement and formation of actin-rich pedestals beneath the bacteria. Though many molecular aspects of attachment have been characterized, contributions of the brush border to the attachment process have not been investigated. We find that while brush border integrity is critical for limiting bacterial attachment, EPEC can utilize this domain to recruit actin bundles to sites of attachment. Using live cell microscopy, we show that EPEC stimulates flow of the brush border across the cell surface to sites of attachment, as well as directed elongation of microvilli towards bacterial cells. Microvillar actin also appears in nascent pedestals, and pedestal formation is inhibited in cells overexpressing an actin bundling protein. This work suggests a novel mechanism wherein EPEC-stimulated pedestal formation does not occur exclusively by de novo formation of a branched actin network, but also progresses through repurposing of existing microvillar parallel actin bundles.

In many organisms, a cytokinetic ring directs daughter cell separation following mitosis. While conserved molecular participants in this process have been defined, the signaling events controlling cytokinetic ring function remain obscure. Using a genetically-tractable fission yeast, Schizosaccharomyces pombe, I have investigated mechanisms involved in such signaling, with a particular interest in kinase and phosphatase networks. Through identification of a new subunit of the S. pombe chromosomal passenger complex, I have found that Aurora B kinase influences cytokinesis by mediating Cdc14-family phosphatase accumulation at the cytokinetic ring. In addition, I have discovered that Sid2, a kinase of the S. pombe septation initiation network, phosphorylates cytokinetic formin Cdc12 to reverse formin multimerization and allow cytokinetic ring maintenance. My studies also indicate that cytokinesis impacts cell cycle-dependent polarized growth, and that phosphosignaling at the cytokinetic ring ensures robust growth following cell division. These studies advance our understanding of molecular cues regulating cytokinesis, and broaden knowledge concerning the consequences of this control.