Natural Killer (NK) cells are highly mobile, specialized sub-populations of lymphocytic cells that survey their host to identify and eliminate infected or tumor cells. They are one of the key players in innate immunity and do not need prior activation through antigen recognition to deliver cytotoxic packages and release messenger chemicals to recruit immune cells. Cytohesin associated scaffolding protein (CASP) is a lymphocyte specific adaptor protein that forms complexes with vesicles and sorting proteins including SNX27 and Cytohesin-1. In this study I show that by using stably integrated shRNA, CASP has a direct role in the secretion of messenger cytokines including IFN-γ, and impedes NK cell motility and ability to kill tumor cells. I also show that CASP is post-translationally modified by ubiquitination and cleavage by granzyme B. CASP is an essential and multi-faceted protein, which has a very diverse role in NK cell specific immune functions.

nÃo hà / This study analyzes the performance of the Course of Professional Self-Sufficiency(CASP) on 2009, in its role of preparing differentiated professionals for the job market in terms of inclusion and raising income. Statistics analysis combined with multiple regression and Binary Dependent Variable models are developed to determine the effectiveness of this program and verify: i) participate to the program implies in an increase, though modest, in labor income; ii) against the expectations of the traditional Mincerian analysis, non-white people have higher probability to be employed; iii) independent of race, male people have a tendency to get out of an unemployment situation. These results are in according to the current changes of labor market and with the level of qualification of labor demanded in the job market of CearÃ. / Este trabalho analisa o desempenho do Curso de Auto-SuficiÃncia Profissional (CASP) no ano de 2009, no seu papel de preparar profissionais diferenciados para o mercado de trabalho nos quesitos inserÃÃo no trabalho e aumento de renda. AnÃlises de estatÃsticas descritivas combinadas a modelos de regressÃo mÃltipla e de variÃvel dependente binÃria permitiram avaliar a eficÃcia deste programa e constatar que: i) participar do programa implica em um incremento, embora modesto, na renda do trabalho; ii) ao contrÃrio do esperado em uma anÃlise minceriana tradicional, os indivÃduos nÃo-brancos possuem uma maior probabilidade em conseguir emprego e iii) independente da raÃa, os indivÃduos do sexo masculino tendem a sair mais rÃpido de uma situaÃÃo de desemprego. Tais constataÃÃes condizem com as mudanÃas atuais do mercado de trabalho e com o grau de qualificaÃÃo da mÃo-de-obra exigido no mercado de trabalho cearense.

Predicting protein structure using its primary sequence has always been a challenging topic in biochemistry. Although it seems as simple as finding the minimal energy conformation, it has been quite difficult to provide an accurate yet reliable solution for the problem. On the one hand, the lack of understanding of the hydrophobic effect as well as the relationship between different stabilizing forces, such as hydrophobic interaction, hydrogen bonding and electronic static interaction prevent the scientist from developing potential functions to estimate free energy. On the other hand, structure databases are limited with redundant structures, which represent a noncontinuous, sparsely-sampled conformational space, and preventing the development of a method suitable for high-resolution, high-accuracy structure prediction that can be applied for functional annotation of an unknown protein sequence. Thus, in this study, we use molecular dynamics simulation as a tool to sample conformational space. Structures were generated with physically realistic conformations that represented the properties of ensembles of native structures. First, we focused our study on the relationship among different factors that stabilize protein structure. Using a wellcharacterized mutation system of the B-hairpin, a fundamental building block of protein, we were able to identify the effect of terminal ion-pairs (salt-bridges) on the stability of the beta-hairpin, and its relationship with hydrophobic interactions and hydrogen bonds. In the same study, we also correlated our theoretical simulations qualitatively with experimental results. Such analysis provides us a better understanding of beta-hairpin stability and helps us to improve the protein engineering method to design more stable hairpins. Second, with large-scale simulations of different representative protein folds, we were able to conduct a fine-grained analysis by sampling the continuous conformational space to characterize the relationship among backbone conformation, side-chain conformation and side-chain packing. Such information is valuable for improving high-resolution structure prediction. Last, with this information, we developed a new prediction algorithm using packing information derived from the conserved relative packing groups. Based on its performance in CASP7, we were able to draw the conclusion that our simulated dataset as well as our packing-oriented prediction method are useful for template based structure prediction.

Background: Magnetic resonance imaging (MRI) techniques are rarely used in the context of abdominal sepsis and in sepsis research. This study investigates the impact of MRI for monitoring septic peritonitis in an animal model (colon ascendens stent-induced peritonitis, CASP). The CASP model closely mimics that of human disease and is highly standardized. The most frequently employed readout parameter in mouse CASP studies is prolonged or decreased rate of survival. Monitoring the progression of peritonitis via MRI could provide a helpful tool in the evaluation of severity. The use of alternative readout systems could very well reduce the number of research animals. Perspectively, clinical improvement after certain treatment could be classified. Methods: This study describes for the first time MRI findings following the induction of septic peritonitis in mice using the CASP model. Two sublethal groups of mice with septic peritonitis were investigated. Each had received one of two differing stent diameters in order to control the leakage of feces into the abdominal cavity. Each mouse served as its own control. Imaging and analyses were performed blinded. Gut diameters, stomach volume, abdominal organ wall diameters, and volume of the adrenal glands were measured. Serum corticosterone levels were detected using ELISA. Serum IL-6, TNF-α, IL-1β, and IL-10 levels were screened by cytometric bead array. Statistical analysis was performed using the Mann-Whitney U test for nonparametric probes and the Kruskal-Wallis and t tests. Results: Using a 7-tesla MRI scanner 24 and 48 h after induction of septic peritonitis, interenteric fluid, organ swelling of spleen and adrenal glands, as well as dilatation of the stomach were compared to nonseptic conditions. Swelling of adrenal glands resulted in an increased serum corticosterone level. In addition, the wall of the intestine bowel was thickened. Based upon these findings, an MRI score (MRI sepsis score, MSS) for abdominal sepsis in mice was established. Reduced stent sizes led to reduced severity of the abdominal sepsis, which could be reproduced in the MSS, which is described here for the first time. Conclusions: Intraabdominal variations during septic peritonitis are detectable by MRI techniques. MRI methods should become a more important tool for the evaluation of abdominal peritonitis. MSS could provide an interesting tool for the evaluation of therapeutic strategies.

<p>Interstrand cross-linking (ICL) damage to DNA is cytotoxic as it blocks replication and transcription. This cytotoxicity is exploited in anti-cancer therapies, but increased ICL repair limits the efficacy of these chemotherapies. SNM1A (sensitive to nitrogen mustard 1A), of the beta-CASP family of nucleases, has been shown to participate in the initiation of one of the ICL repair processes. Biochemical studies of SNM1A have been limited due to insolubility and instability of SNM1A in bacteria and insect cell lines and toxicity in human cell lines. Work reported in this thesis describes a novel and efficient method of generating active protein from inclusion body expression of the beta-CASP domain of SNM1A. This refolded beta-CASP domain shows 5’ exonuclease activity on single stranded and double stranded DNA in vitro. Nevertheless, this domain alone is unable to complement <em>pso2</em> null ICL repair defects in<em> S. cerevisiae</em> after exposure to ICL agents. These functional studies of the beta-CASP domain of SNM1A will be helpful in directing future research on its role in ICL repair. Additionally, this will aid future structural and inhibitor studies of this essential interstrand cross-link repair protein, SNM1A.</p> / Master of Science (MSc)

Interstrand cross-links (ICLs) are a type of DNA damage that prevents strand separation required for basic cellular processes. ICL-based anti-cancer therapies exploit the cytotoxic consequences of replication and transcription inhibition, however, they are limited by the ability of the cell to repair DNA crosslinks. The challenge of ICL repair involves coordinating multiple DNA repair pathways to remove damage occurring on both strands of DNA. Participation of factors that are both exclusive and essential to crosslink repair suggests a pathway requirement to process unique structures and/or intermediates arising only in ICL repair. SNM1A is a nuclease required for survival of human cells in response to ICL exposure, but the specific function and role of SNM1A remain unclear. Here we show that, in addition to known 5’-3’exonuclease activity, SNM1A possesses single-strand specific endonuclease activity. Furthermore, SNM1A exhibits translesion nuclease activity on crosslinks which deform the helical backbone, but not non-distorting stable ICLs. We report the identification and characterization of nine small molecules inhibitors of SNM1A, isolated from an in vitro high-throughput screen of nearly 4,000 bioactive compounds. Finally, we demonstrate that inhibitors of SNM1A potentiate the cytotoxicity of ICL-inducing agent cisplatin in HeLa cells. The work in this thesis expands the possible roles of SNM1A in ICL repair and lays the groundwork for SNM1A inhibition in ICL sensitization efforts. / Thesis / Doctor of Philosophy (PhD)

Adhesion between the plasma membrane and the cell wall and the existence of the continuum of these two compartments is needed for signal transmission, e.g. under pathogen attack, during cell expansion and cell wall growth, or in response to environmental conditions. This adhesion is, in addition to the turgor, provided by physical connection of both compartments. One of the best-known examples of physical connection is found in the root system, in the Caspary strip region, where it is required to maintain apoplastic barriers of the root system, even under adverse conditions and consequent plasmolysis. There is little information about the physical interconnection and the participating macromolecules but there are candidates, which could participate in this interaction. The diploma thesis deals with arabinogalactan (AGP) proteins with fasciclin-like domain (namely FLA9 and FLA2). These proteins may play a role in the adhesion of plasma membrane and cell wall and may be involved in the growth regulation and development of the root system. Both genes are are relatively strongly expressed in the roots, especially in the elongation zone and in the cortex, including the endodermis (according to chip data). Also, to a smaller extent, the thesis deals with integrin-like At14A protein. The most interesting...

I analyzed the 2004 and 2006 Health and Retirement Study data to test structural equation models of the quality of life (QOL) construct. The participants (N = 1352) were non-institutionalized individuals aged 42 and older (M = 65.70, SD = 10.88), with an average education of 12.73 years (SD = 2.96) and of varied ethnicities. The results indicated that physical functioning, affective experience, life satisfaction and social support could serve as indicators for a second order QOL factor. Furthermore, the developed QOL model explained 96% of the variance of the CASP-19 (Control, Autonomy, Self-realization and Pleasure), a QOL measure that reflects fulfillment of psychological needs. The results also indicated that Depression and Life Satisfaction are related through reciprocal causation and that Physical Functioning is more likely to cause a change in Depression than the reverse. The results suggest that QOL is a complex, multidimensional concept that should be studied at different levels of analysis.

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The goal of this thesis is test and comparation of the offline tools for prediction of protein structure and creation of metaprediktor, which allows the user to select the appropriate tool, according to given parameters. Testing tool is based on a dataset of proteins, which is based on the SCOP database and it is trying to be as balanced as possible to include proteins from different families and thus could best evaluate individual tools. The results of this thesis are requirements of metaprediktor and also which data and settings can be allowed and processed and how it will be implemented.