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The retinoblastoma family of phosphoproteins consisting of the retinoblastoma protein (pRB) and the two structurally related proteins p130 and p107 play an important role in the negative regulation of cell cycle progression. Hypophosphorylated pocket proteins interact with the different members of the E2F family and repress the transcription of E2F-dependent genes and consequently suppress cell cycle progression through the G0/G1 transition and the restriction point in G1. Mitogenic stimulation results in sequential activation of cyclin/CDK complexes in mid to late G1, leading to subsequent hyperphosphorylation at multiple Ser/Thr sites of pocket proteins triggering dissociation of pocket protein/E2F complexes. This disruption leads to de-repression of many E2F dependent genes whose products are essential for cell cycle progression. The traditional view has been that pocket proteins continue to be hyperphosphorylated through the S and G2 phases and following cyclin/CDK inactivation during mitotic exit become dephosphorylated by action of PP1. However, our lab observed that upon treatment of asynchronously growing cells with the CDK inhibitor Flavopiridol or CHX, pocket proteins, are rapidly dephosphorylated correlating with the inactivation of G1/CDKs and down regulation of D-type cyclins, respectively. Pocket protein dephosphorylation was prevented by pre-treating these cells with phosphtase inhibitors at a concentration selective for PP2A, implicating PP2A or PP2A-like serine/threonine phosphatase in this iii process. The involvement of PP2A on pocket protein dephosphorylation was further strengthened by the observation that SV40 small t antigen (ST) delays/prevents p107 dephosphorylation. Moreover, a physical association between PP2A/C and p130/p107 was observed throughout the cell cycle that was not affected by CHX treatment, strongly suggesting that CHX-induced dephosphorylation is not the result of increased pocket protein targeting by PP2A, but rather that a dynamic equilibrium between CDKs and PP2A is shifted to dephosphorylation when CDK activity is compromised. This dynamic equilibrium operates throughout the cell cycle. PP2A is a trimeric enzyme complex consisting of a catalytic C, a structural A and substrate specific B subunit. There are four families of regulatory B subunits designated B, B’, B’’ and B’’’, each with several members encoded by genes with multiple splice variants that mediate substrate specificity and subcellular localization. It has been reported recently that in excess of 200 functional distinct PP2A holoenzymes can assemble with distinct specificities. Therefore, to gain insight into the mechanisms that regulate the steady state phosphorylation of pocket proteins throughout the cell cycle, it was essential to identify the specific holoenzyme complexes involved. To this end, it was identified that a PP2A trimeric holoenzyme containing B55α specifically targets and dephosphorylates p107/p130 both in vitro and in mammalian cells. B55α associates directly with the spacer of p107 and this interaction seems to be indirectly enhanced by the C-terminus of p107. The decreased association of p107 with PP2A/C of the B55α/PP2A holoenzyme complex upon treatment with ST further confirmed the role of B55α in mediating p107-PP2A/C interaction. Our data also revealed an interaction between B55α and p130, but not pRb, which appears to prefer a PR70, suggesting selectivity in the interaction of pocket proteins with distinct PP2A holoenzymes. In accordance with this, recombinant purified B55α dephosphorylates p107 in vitro. Limited ectopic expression of B55α but not other subunits, result in ST sensitive dephosphorylation of p107 and p130 in cells. Further shRNA mediated knockdown of B55α results in hyperphosphorylation of p107 and p130. This suggests that the cellular levels of B55α are critical in modulating the phosphorylation status of p107/p130 rather than just catalyzing the dephosphorylation of these proteins when the activity of CDKs is compromised. Since ST disrupts the B55α/PP2A holoenzyme complex by binding to the PP2A-A-C dimer and leads to hyperphosphorylation of pocket proteins it is conceivable that ST mediates its effects on cell proliferation at least in part, via inactivation of the PP2A holoenzymes that activates pocket proteins. Given the sensitivity of p107 phosphorylation to the cellular levels of B55α, future analyses should ascertain if deregulation of B55α leads to hyperphosphorylation of pocket proteins and abnormal cell cycle progression. / Molecular Biology and Genetics

One of the major themes throughout the works of Nathaniel Hawthorne portrays an image of men and women who are physically and morally isolated from the world. So extensive was Hawthorne's interest in this problem that it emerges as a distinct concern in his philosophy. This thesis will discuss the process, as put forth by Hawthorne, by which an individual is drawn apart from humanity and then will describe the only solution the author saw for the isolated individual. Only male isolates can be brought back to the world, and only love for a pure woman can accomplish this act. Basic definitions and explanations of isolation of the heart or of the intellect will be given and supported by specific examples; the means of redemption will also be explained. Orientation to the life of Hawthorne will be provided, since the entire concept of isolation and redemption seemingly evolved from the author's personal experience and environment. Characters other than those involved.in this concept of isolation and salvation will be approached only as they serve to more clearly delineate and clarify this particular aspect of Hawthorne's philosophy. / Master of Arts

A dynamic pressure measuring system composed of long connecting tube, transducer, D-C amplifier and galvanometer oscillograph were used in a wind study conducted by the Department of Agricultural Engineering, Virginia Polytechnic Institute. The instrumentation was used to measure and record dynamic pressures due to wind velocity. Data recorded in that study indicated velocity head pressure oscillations on some surfaces of the test structure in excess of 100 Hz. This investigation was undertaken to see if the recorded oscillations could have been generated in the measuring system or if they were truly windpressure variations. Components of the pressure measuring system were modeled by transfer functions. From these transfer functions, a system operational transfer function was determined and used to define system frequency response. The frequency analysis indicated the system was severely limited in response by the oscillating air column constrained within the interconnecting tube. The usable frequency range of the system (+ 10% tolerable amplitude error allowable) was found to go from zero to 34.2 Hz. By eliminating the air column this frequency range could have been extended to 99.4 Hz. Phase shifts in these frequency ranges were found to be negligible. Transducers were mounted rigidly to exterior walls of the test structure. The measured fundamental natural frequencies of these walls (transducer mountings) were found to be well within the usable frequency range of the instrumentation; and, therefore, a source of vibration pickup. To improve the measuring system reliability, recommendations were made to eliminate the air column and stiffen the transducer mountings. / Master of Science

Le cycle cellulaire est hautement régulé par la phosphorylation réversible de plusieurs effecteurs. La kinase dépendante des cyclines Cdk1 déclenche la mitose en induisant le bris de l’enveloppe nucléaire, la condensation des chromosomes et la formation du fuseau mitotique. Chez les animaux métazoaires, ces évènements sont contrés par la protéine phosphatase PP2A-B55, qui déphosphoryle plusieurs substrats de Cdk1. La kinase Greatwall (Gwl) est activée par le complexe cycline B-Cdk1 en début de mitose et induit ensuite l’inhibition de PP2A-B55 via Endos/Arpp19. Toutefois, les mécanismes moléculaires qui régulent Gwl sont encore peu connus. Nous avons montré que Gwl a une activité s’opposant à PP2A-B55, qui collabore avec la kinase Polo pour assurer l’attachement du centrosome au noyau et la progression du cycle cellulaire dans le syncytium de l’embryon de la drosophile. Ensuite, nous avons trouvé dans des cellules de drosophile que Gwl est localisée au noyau pendant l’interphase, mais qu’elle se relocalise au cytoplasme dès la prophase, avant le bris de l’enveloppe nucléaire. Nous avons montré que cette translocation de Gwl est cruciale pour sa fonction et qu’elle dépend de la phosphorylation de plusieurs résidus de la région centrale de Gwl par les kinases Polo et Cdk1. Cette région centrale contient également deux séquences de localisation nucléaire (respectivement NLS1 et NLS2). De plus, nos résultats suggèrent que la phosphorylation de Gwl par la kinase Polo promeut sa liaison avec la protéine 14-3-3ε, ce qui favorise la rétention cytoplasmique de Gwl. Le rôle de Cdk1 dans cette translocation reste quant à lui inconnu. De plus, nous avons montré que le complexe cycline B-Cdk1 entre dans le noyau avant que Gwl ne soit transportée dans le cytoplasme. Cdk1 pourrait donc activer Gwl et phosphoryler ses substrats nucléaires, à l’abri de PP2A-B55 qui est largement cytoplasmique. Gwl est ensuite exclue du noyau et relocalisée dans le cytoplasme afin d’induire l’inhibition de PP2A-B55. Cela permet de synchroniser les événements de phosphorylation se produisant dans le noyau et dans le cytoplasme. Fait intéressant, un mécanisme de régulation de la localisation de Gwl similaire à cela a été découvert chez l’humain et chez la levure, suggérant que ce mécanisme est conservé entre différentes espèces. / Reversible phosphorylation of proteins, triggered by cyclically activated kinases and phosphatases, is a key mechanism to control cell cycle progression. CyclinB-Cdk1 is a crucial kinase phosphorylating a large number of substrates to trigger mitotic entry. However, in metazoans, it is counteracted mainly by a Protein Phosphatase 2A carrying the B55 regulatory subunit (PP2A-B55). On the other hand, the Greatwall (Gwl) kinase is activated by CyclinB-Cdk1 upon mitotic entry and subsequently induces the inhibition of PP2A-B55 by Endos/Arpp19, thus promoting mitotic entry and maintenance. Nonetheless, the regulatory mechanisms of Gwl are less clear. We demonstrated that in Drosophila syncytial embryos, PP2A-B55 is negatively regulated by Gwl, but collaborates with Polo kinase to ensure both nucleus attachment of centrosome and faithful cell cycle progression. Later, we discovered that in Drosophila, the subcellular localization of Gwl changes dramatically throughout the cell cycle. Gwl is nuclear in interphase but suddenly becomes mostly cytoplasmic in prophase before nuclear envelope breakdown. Such translocation is important for Gwl’s function and requires the phosphorylation of Gwl by both Polo kinase and Cdk1 in the region containing two Nuclear Localization Signals (NLSs). Phosphorylation of Gwl by Polo likely promotes its association with14-3-3ε thereby promoting Gwl cytoplasmic retention, whereas Cdk1’s role in this translocation remains elusive. Moreover, I found that most cyclin B is imported into the nucleus before Gwl translocates to the cytoplasm. Therefore, Cdk1 can activate Gwl and phosphorylate its nuclear substrates without the perturbation of PP2A-B55 which is largely cytoplasmic. Subsequently, Gwl translocates into cytoplasm to mediate the inhibition of PP2A-B55 so that the phosphorylation events can be synchronized between the nucleus and the cytoplasm. Interestingly, similar spatial regulation of Gwl was also uncovered in mammal cells and in yeast, implying a conserved regulatory mechanism across species.

There is a strong body of psychological research implicating the mass media in the aetiology of adolescent girls' negative body image and eating behaviours. The present thesis aims to extend this research by examining potential factors – namely, media type, body perfect internalisation and materialism – that make girls more vulnerable to the negative impact of the mass media. An initial meta-analysis (Chapter 3) collated the findings of existing research examining the impact of ‘body perfect' media on adolescents' body image; examining gender, age and media type as moderators of this effect. Chapter 4 examined the relative roles of both media type and media model identification (a key dimension of body perfect internalisation), within the mass media and body image relationship. Using both survey and experimental methods (N = 199), it was found that adolescent girls' habitual tendency to identify with media models, was a more potent vulnerability factor within the mass media and body image relationship, than media type. Due to the limitations associated with existing measures of body perfect internalisation, a new measure of body perfect internalisation was developed in Chapter 5 (N =373), which was subsequently utilised in the final experiments of the thesis. Chapter 6 demonstrated that acute music video exposure had a more potent negative impact on girls' body image than still media images (N = 142); an effect that was fully mediated by wishful character identification and also moderated by body perfect internalisation. Chapter 7 consists of two studies that demonstrate the important role which materialism plays within the mass media, body image and eating behaviour relationship. In Study 1, structural equation modelling identified a direct pathway between materialism and restrained eating that was independent of body image (N = 199). This finding was further replicated in an exposure experiment, which demonstrated that brief exposure to materialistic media causes acute diet-like behaviours in adolescent girls (N = 180).

Protein phosphorylation is a reversible post-translational modification that is critical for the regulation of key cellular processes. It is estimated that two-thirds of all cellular proteins are phosphorylated, with more than 98% of those phosphorylation events occurring on Ser/Thr residues. Protein phosphorylation is mediated by protein kinases and reversed via dephosphorylation by protein phosphatases. Two protein phosphatases, phosphatase 1 (PP1) and Protein phosphatase 2A (PP2A), are thought to account for more than 90% of the total phosphatase activity in eukaryotic cells. PP2A is a highly conserved assortment of heterotrimeric holoenzymes responsible for the dephosphorylation of many regulated phosphoproteins. Substrate recognition and the integration of regulatory cues are mediated by B regulatory subunits that are complexed to the catalytic subunit (C) by a scaffold protein (A). Substrate recruitment of PP2A/B55α, the most abundant PP2A holoenzyme, was thought to be mediated by charge-charge interactions between the surface of B55α and its substrates. Challenging this view, we recently discovered a conserved SLiM (Short Linear Motif) [RK]-V-x-x-[VI]-R in a range of proteins, including substrates such as the retinoblastoma-related protein p107 and TAU (Fowle et al. eLife 2021;10:e63181). Here we report the identification of this SLiM in FAM122A, an inhibitor of B55α/PP2A, and analysis of the associated proteomic datasets that aided in identifying FAM122A, which can assist in the further identification of potential substrates and cellular pathways regulated by this phosphatase. The newly identified conserved SLiM is necessary for FAM122A binding to B55α in vitro and in cells. Computational structure prediction with AlphaFold2 predicts an interaction consistent with the mutational data and supports a mechanism whereby FAM122A uses the ‘SLiM’ in the form of a short α-helix to dock to the B55α top groove. In this model, FAM122A spatially constrains substrate access by occluding the catalytic subunit with a second α-helix immediately adjacent to helix-1. Consistently, FAM122A functions as a competitive inhibitor as it prevents the binding of substrates in in vitro competition assays and the dephosphorylation of CDK substrates by B55α/PP2A in cell lysates. Ablation of FAM122A in human cell lines reduces the rate of proliferation, the progression through cell cycle transitions, and abrogates G1/S and intra-S phase cell cycle checkpoints. FAM122A-KO in HEK293 cells results in the attenuation of CHK1 and CHK2 activation in response to replication stress. Overall, these data strongly suggest that FAM122A is a ‘SLiM’-dependent, substrate-competitive inhibitor of B55α/PP2A that suppresses multiple functions of B55α in the DNA damage response and in timely progression through the cell cycle interphase. In agreement with these findings, ectopic expression of B55α results in the downregulation of 14-3-3σ signaling mediated by ATM and ATR as determined by pathway analysis of phosphoproteomic datasets and a reduction of ATM signaling within the total proteome. Altogether, this work has significantly expanded our understanding of the PP2A/B55 SLiM, resulting from the characterization of FAM122A, a high-affinity substrate inhibitor, and enables future interrogation of novel substrates and signaling networks regulated by PP2A/B55α. / Biomedical Sciences

The subject of this thesis is the description of quantum black holes as a way to probe quantum gravity. Scenarios of a lower Planck scale make quantum gravity experimentally approachable, therefore a theoretical framework is needed in order to be able to probe quantum gravitational effects. We present a field theoretical formalism for the treatment of quantum black holes and their interactions with particles of the Standard Model. We examine the properties and assumptions governing quantum black holes and develop a methodology to examine their behavior using quantum field theory language. We apply our formalism in several different cases and calculate the cross sections for the production of quantum black holes. We use our results to gain phenomenological insights to quantum gravity, such as the derivation of bounds for the Planck mass from Standard Model processes. The distinction between a continuous and a discrete mass spectrum, for a quantum black hole, is discussed and the relevant cross sections presented. Finally, we use quantum black holes as a gateway to supersymmetry and calculate the branching ratios for the decay of quantum black holes into supersymmetric particles.

Pendant le bris de l'enveloppe nucléaire, la kinase dépendante des cyclines liée à la cycline B (CDK1-cycline B) et d'autres kinases phosphorylent des protéines nucléaires conduisant au désassemblage des complexes de protéines de l'enveloppe nucléaire. Les protéines nucléaires phosphorylées sont ensuite déphosphorylées par un groupe de phosphatases en sortie mitotique. La protéine phosphatase 2A en complexe avec la sous-unité régulatrice B55 (PP2A-B55) est connue pour être la principale phosphatase à déphosphoryler les protéines critiques à la fin de la mitose. Cependant, les substrats nucléaires déphosphorylés par PP2A-B55 à la sortie mitotique sont peu connus. En utilisant des cellules de drosophile en culture, nous avons démontré que PP2A-B55 est nécessaire pour le recrutement de protéines de l'envelope nucléaire telles que BAF, la protéine de lamina nucléaire Lamin B et la nucléoporine Nup107. De plus, nous avons trouvé que les œufs de femelles des drosophiles hétérozygotes pour une mutation dans les gènes codant pour la Lamine B et Tws (B55 chez la drosophile) n’éclosent pas. Ces œufs présentent divers défauts au stade de la méiose et des divisions nucléaires de l’embryon syncytial. De plus, des tests in vitro et d'autres analyses biochimiques indiquent que PP2A-Tws se lie et déphosphoryle BAF. J'ai d'autres résultats qui suggèrent un rôle de la protéine Ankle2 dans la régulation du recrutement de BAF pour réassembler les noyaux à la sortie mitotique. Mes résultats suggèrent également que Ankle2 en complexe avec PP2A est responsable de la bonne progression mitotique. Mes résultats mettent en évidence l'utilité de la drosophile comme système modèle dans l'étude de différents aspects du cycle cellulaire. Ils démontrent également / During nuclear envelope breakdown, the cyclin-dependent kinase 1 bound to Cyclin B (CDK1-Cyclin B) and other kinases phosphorylate a number of nuclear proteins leading to the disassembly of nuclear envelope protein complexes. Phosphorylated nuclear proteins are then dephosphorylated by a group of phosphatases at mitotic exit. The protein phosphatase 2A in complex with the regulatory subunit B55 (PP2A-B55) is known to be the major phosphatase to dephosphorylate critical proteins at the end of mitosis. However, little was known about the nuclear substrates dephosphorylated by PP2A-B55 at mitotic exit. Using Drosophila cells in culture, we demonstrated that PP2A-B55 is required for the recruitment of nuclear envelope proteins such as BAF, the nuclear lamina protein Lamin B, and the nucleoporin Nup107. Also, eggs from Drosophila females heterozygous for a mutation in genes coding for Lamin B and Tws (B55 in Drosophila), didn’t hatch. These eggs showed various defects during the nuclear division stage and meiosis. Moreover, in vitro assays and other biochemical analyses indicate that PP2A-B55 binds and dephosphorylates BAF. I have other results that suggest a role of the protein Ankle2 in regulating BAF recruitment to reassembling nuclei at mitotic exit. My results also suggest that Ankle2 in complex with PP2A is responsible for the proper mitotic progression. Our results highlight the importance of Drosophila in investigating different aspects of the cell cycle. It also demonstrates a role of PP2A in the nuclear envelope reformation at the end of mitosis.

This study is concerned with the role of sound in the presentation and representation of bodies in performance art that incorporates digital technologies. It consists of a written thesis accompanied by a portfolio with documentation of original artwork. Since the 1960s, performance artists have explored the use of sensor technologies to register signals generated by the body and synthesize or control sound. However, both practical and theoretical approaches to biosignal sonification in this field have almost entirely focused on musical (formalist) perspectives, technological innovation, or heightening the performer's and spectator's awareness of their body's physiology. Little attention has been paid to the usually conspicuous interaction between body and technological equipment and the role of the generated sound in the context of cultural critical debates regarding the performing body. The present study responds to this observation in two ways: Firstly, the written part of the study examines existing biosignal performance practices. It seeks to demonstrate that artists' decisions on the design of sensor technology and sound synthesis or manipulation methods are often complicit in the representation of normative body types and behaviour. Drawing from a concept of the sonified body as a transgressive or ‘freak' body, three critical perspectives on biosignal sonification in digital performance are proposed: A reading of body sonification methods from a gender-critical perspective, an inquiry in the context of Mikhail Bakhtin's concepts of the grotesque and the classical body, and a conceptualization of the sonified body as a posthuman prosthetisized body. This part of the study serves as a framework for its second objective: the development of practical performance strategies to address and challenge cultural conventions concerning ‘the' body's form and role in society. This aspect of the thesis is developed in conjunction with, and further explored in, the artwork documented in the portfolio. The practical part of the study consists of three digital performance works. ELECTRODE (2011) involves an anal electrode that registers the activity of my sphincter muscle and uses this data to synthesize sound. For this work, I modified a commercially available muscle tension sensor device designed for people with faecal incontinence problems. Feedback (2010) encompasses components of a commercially available fetal Doppler sensor intended to listen to the heartbeat of unborn babies. SUIT (2009-2010) encompasses several performances that feature a PVC overall equipped with a loudspeaker, sensor interface and Doppler and humidity sensors.