Studies on the {\it in vivo} function of a phosphatidylinositol transfer protein from {\it Saccharomyces cerevisiae}

Phosphatidylinositol transfer proteins (PI-TP's) catalyze the transfer of phosphatidylinositol and phosphatidylcholine between membranes in vitro. However the in vivo function of these proteins is unknown. In this thesis we have used a combined biochemical and genetic approach to determine the importance of PI-TP in vivo. An oligonucleotide based on the amino terminal sequence of the PI-TP from Saccharomyces cerevisiae, was used to screen a yeast genomic library for the gene encoding PI-TP (PIT1 gene). Yeast strains transformed with the positive clones showed overproduction of transfer activities and transfer protein in the 100,000 x g supernatants. The 5$\sp\prime$ terminus of the PIT1 gene correlates with the predicted codons for residues 3-30 of the determined protein sequence. Tetrad analysis of a heterozygous diploid (PIT1/pit1::LEU2) revealed that the PIT1 gene is essential for cell growth. Non-viable spores could be rescued by transformation of the above diploid prior to sporulation, with a plasmid borne copy of the wild type gene. Sequencing of the entire PIT1 gene has revealed that the PIT1 gene is identical to the SEC14 gene. The sec14 ts mutant which exhibits conditional defects at the Golgi stage of protein secretion, is also temperature sensitive for PI-TP activity in vitro. These findings represent the first instance in which a physiological function has been assigned to any phospholipid transfer protein. ^

Zoning and the built environment: A public health analysis of zoning in Austin, Texas

The built environment is part of the physical environment made by people and for people. Because the built environment is such a ubiquitous component of the environment, it acts as an important pathway in determining health outcomes. Zoning, a type of urban planning policy, is one of the most important mechanisms connecting the built environment to public health. This policy analysis research paper explores how zoning regulations in Austin, Texas promote or prohibit the development of a healthy built environment. A systematic literature review was obtained from Active Living Research, which contained literature published about the relationships between the built environment, physical activity, and health. The results of these studies identified the following four components of the built environment that were associated to health: access to recreational facilities, sprawl and residential density, land use mix, and sidewalks and their walkability. A hierarchy analysis was then performed to demonstrate the association between these aspects of the built environment and health outcomes such as obesity, cardiovascular disease, and general health. Once these associations had been established, the components of the built environment were adapted into the evaluation criteria used to conduct a public health analysis of Austin's zoning ordinance. A total of eighty-eight regulations were identified to be related to these components and their varying associations to human health. Eight regulations were projected to have a negative association to health, three would have both a positive and negative association simultaneously, and nine were indeterminable with the information obtained through the literature review. The remaining sixty-eight regulations were projected to be associated in a beneficial manner to human health. Therefore, it was concluded that Austin's zoning ordinance would have an overwhelmingly positive impact on the public's health based on identified associations between the built environment and health outcomes.^

Analysis of the E1-ubiquitin activating enzyme, Uba1, in cell death and tissue growth in Drosophila

Ubiquitination is an essential process involved in basic biological processes such as the cell cycle and cell death. Ubiquitination is initiated by ubiquitin-activating enzymes (E1), which activate and transfer ubiquitin to ubiquitin-conjugating enzymes (E2). Subsequently, ubiquitin is transferred to target proteins via ubiquitin ligases (E3). Defects in ubiquitin conjugation have been implicated in several forms of malignancy, the pathogenesis of several genetic diseases, immune surveillance/viral pathogenesis, and the pathology of muscle wasting. However, the consequences of partial or complete loss of ubiquitin conjugation in multi-cellular organisms are not well understood. Here, we report the characterization of nba1, the sole E1 in Drosophila. We have determined that weak and strong nba1 alleluias behave genetically different and sometimes in opposing phenotypes. For example, weak uba1 alleluias protect cells from cell death whereas cells containing strong loss-of-function alleluias are highly apoptotic. These opposing phenotypes are due to differing sensitivities of cell death pathway components to ubiquitination level alterations. In addition, strong uba1 alleluias induce cell cycle arrest due to defects in the protein degradation of Cyclins. Surprisingly, clones of strong uba1 mutant alleluias stimulate neighboring wild-type tissue to undergo cell division in a non-autonomous manner resulting in severe overgrowth phenotypes in the mosaic fly. I have determined that the observed overgrowth phenotypes were due to a failure to downregulate the Notch signaling pathway in nba1 mutant cells. Aberrant Notch signaling results in the secretion of a local cytokine and activation of JAK/STAT pathway in neighboring cells. In addition, we elucidated a model describing the regulation of the caspase Dronc in surviving cells. Binding of Dronc by its inhibitor Diap1 is necessary but not sufficient to inhibit Dronc function. Ubiquitin conjugation and Uba1 function is necessary for the negative regulation of Dronc. ^

INFANT MORTALITY IN THE MEXICAN AMERICAN COMMUNITY: AN ANALYSIS OF DIFFERENTIALS IN INFANT, NEONATAL, AND POSTNEONATAL MORTALITY BY ETHNICITY AND PARENTAL NATIVITY, 1974-75 SINGLE LIVE BIRTH COHORT, HARRIS COUNTY, TEXAS

The paradoxically low infant mortality rates for Mexican Americans in Texas have been attributed to inaccuracies in vital registration and idiosyncracies in Mexican migration in rural areas along the U.S.-Mexico border. This study examined infant (IMR), neonatal (NMR), and postneonatal (PNMR) mortality rates of Mexican Americans in an urban, non-border setting, using linked birth and death records of the 1974-75 single live birth cohort (N = 68,584) in Harris County, Texas, which includes the city of Houston and is reported to have nearly complete birth and death registration. The use of parental nativity with the traditional Spanish surname criterion made it possible to distinguish infants of Mexican-born immigrants from those of Blacks, Anglos, other Hispanics, and later-generation, more Anglicized Mexican Americans. Mortality rates were analyzed by ethnicity, parental nativity, and cause of death, with respect to birth weight, birth order, maternal age, legitimacy status, and time of first prenatal care.^ While overall IMRs showed Spanish surname rates slightly higher than Anglo rates, infants of Mexican-born immigrants had much lower NMRs than did Anglos, even for moderately low birth weight infants. However, among infants under 1500 grams, presumably unable to be discharged home in the neonatal period, Mexican Americans had the highest NMR. The inconsistency suggested unreported deaths for Mexican American low birth weight infants after hospital discharge. The PNMR of infants of Mexican immigrants was also lower than for Anglos, and the usual mortality differentials were reversed: high-risk categories of high birth order, high maternal age, and late/no prenatal care had the lowest PNMRs. Since these groups' characteristics are congruent with those of low-income migrants, the data suggested the possibility of migration losses. Cause of death analysis suggested that prematurity and birth injuries are greater problems than heretofore recognized among Mexican Americans, and that home births and "shoebox burials" may be unrecorded even in an urban setting.^ Caution is advised in the interpretation of infant mortality rates for a Spanish surname population of Mexican origin, even in an urban, non-border area with reportedly excellent birth and death registration. ^

1906 Polk's Medical Register and Directory of North America (Texas Portion Only)

List of Physicians and Surgeons arranged by location, giving post office address with population and location, the School practiced, date and college of graduation, all the existing and extinct medical colleges in North America, with locations, officers, number of professors, lecturers, demonstrators, etc., the various medical societies, penal, reformatory and charitable state institutions, hospitals, sanitariums, dispensaries, asylums and other medical institutions, boards of health, boards of medical examiners, health officers at principal points, the laws of registration and other laws relating to the profession, medical journals with names of editors, frequency of publication and subscription rates, medical libraries, a therapeutic classification of American health resorts, mineral springs, official list of officers of the medical departments of the U.S. Army, Navy and Marine Hospital Service, the U.S. Indian Service, roster of examining surgeons of the U.S. Pension Department, a descriptive sketch of each state, territory and province, embodying such matters as location, boundary, extent in miles and acres, latitude and longitude, statistics relating to climate, temperature, rate of mortality, etc. Full particulars of all national and inter-state associations and societies relating to medicine and surgery, and an INDEX TO THE PHYSICIANS OF THE UNITED STATES. Arranged alphabetically, with the number of the page and column in which the name appears.

1902 Polk's Medical Register and Directory of the United States and Canada (Texas portion only)

List of Physicians and Surgeons arranged by states and provinces, giving post office address with population and location, the School practiced, date and college of graduation, all the existing and extinct medical colleges in the United States and Canada, with locations, officers, number of professors, lecturers, demonstrators, etc., the various medical societies, state prisons, hospitals, sanitariums, dispensaries, asylums and other medical institutions, boards of health, boards of medical examiners, a synopsis of the laws of registration and other laws relating to the profession, medical journals with names of editors, frequency of publication and subscription rates, medical libraries, mineral springs, official list of officers of the medical departments of the U.S. Army, Navy and Marine Hospital Service, roster of examining surgeons of the U.S. Pension Department, a descriptive sketch of each state, territory and province, embodying such matters as location, boundary, extent in miles and acres, latitude and longitude, statistics relating to climate, temperature, rate of mortality, number of deaths from consumption, etc. full particulars of all national associations and societies relating to medicine and surgery, and an INDEX TO THE PHYSICIANS OF THE UNITED STATES. Arranged alphabetically, with the number of the page on which the name appears.

Comparison of Tumor Shrinkage and Cumulative Dose Distribution for Lung Cancers

Background: The physical characteristic of protons is that they deliver most of their radiation dose to the target volume and deliver no dose to the normal tissue distal to the tumor. Previously, numerous studies have shown unique advantages of proton therapy over intensity-modulated radiation therapy (IMRT) in conforming dose to the tumor and sparing dose to the surrounding normal tissues and the critical structures in many clinical sites. However, proton therapy is known to be more sensitive to treatment uncertainties such as inter- and intra-fractional variations in patient anatomy. To date, no study has clearly demonstrated the effectiveness of proton therapy compared with the conventional IMRT under the consideration of both respiratory motion and tumor shrinkage in non-small cell lung cancer (NSCLC) patients. Purpose: This thesis investigated two questions for establishing a clinically relevant comparison of the two different modalities (IMRT and proton therapy). The first question was whether or not there are any differences in tumor shrinkage between patients randomized to IMRT versus passively scattered proton therapy (PSPT). Tumor shrinkage is considered a standard measure of radiation therapy response that has been widely used to gauge a short-term progression of radiation therapy. The second question was whether or not there are any differences between the planned dose and 5D dose under the influence of inter- and intra-fractional variations in the patient anatomy for both modalities. Methods: A total of 45 patients (25 IMRT patients and 20 PSPT patients) were used to quantify the tumor shrinkage in terms of the change of the primary gross tumor volume (GTVp). All patients were randomized to receive either IMRT or PSPT for NSCLC. Treatment planning goals were identical for both groups. All patients received 5 to 8 weekly repeated 4-dimensional computed tomography (4DCT) scans during the course of radiation treatments. The original GTVp contours were propagated to T50 of weekly 4DCT images using deformable image registration and their absolute volumes were measured. Statistical analysis was performed to compare the distribution of tumor shrinkage between the two population groups. In order to investigate the difference between the planned dose and the 5D dose with consideration of both breathing motion and anatomical change, we re-calculated new dose distributions at every phase of the breathing cycle for all available weekly 4DCT data sets which resulted 50 to 80 individual dose calculations for each of the 7 patients presented in this thesis. The newly calculated dose distributions were then deformed and accumulated to T50 of the planning 4DCT for comparison with the planned dose distribution. Results: At the end of the treatment, both IMRT and PSPT groups showed mean tumor volume reductions of 23.6% ( 19.2%) and 20.9% ( 17.0 %) respectively. Moreover, the mean difference in tumor shrinkage between two groups is 3% along with the corresponding 95% confidence interval, [-8%, 14%]. The rate of tumor shrinkage was highly correlated with the initial tumor volume size. For the planning dose and 5D dose comparison study, all 7 patients showed a mean difference of 1 % in terms of target coverage for both IMRT and PSPT treatment plans. Conclusions: The results of the tumor shrinkage investigation showed no statistically significant difference in tumor shrinkage between the IMRT and PSPT patients, and the tumor shrinkage between the two modalities is similar based on the 95% confidence interval. From the pilot study of comparing the planned dose with the 5D dose, we found the difference to be only 1%. Overall impression of the two modalities in terms of treatment response as measured by the tumor shrinkage and 5D dose under the influence of anatomical change that were designed under the same protocol (i.e. randomized trial) showed similar result.

Renal Cryoablation: Investigation of Periprocedural Visualization Tools and Treatment Response Quantification

Cryoablation for small renal tumors has demonstrated sufficient clinical efficacy over the past decade as a non-surgical nephron-sparing approach for treating renal masses for patients who are not surgical candidates. Minimally invasive percutaneous cryoablations have been performed with image guidance from CT, ultrasound, and MRI. During the MRI-guided cryoablation procedure, the interventional radiologist visually compares the iceball size on monitoring images with respect to the original tumor on separate planning images. The comparisons made during the monitoring step are time consuming, inefficient and sometimes lack the precision needed for decision making, requiring the radiologist to make further changes later in the procedure. This study sought to mitigate uncertainty in these visual comparisons by quantifying tissue response to cryoablation and providing visualization of the response during the procedure. Based on retrospective analysis of MR-guided cryoablation patient data, registration and segmentation algorithms were investigated and implemented for periprocedural visualization to deliver iceball position/size with respect to planning images registered within 3.3mm with at least 70% overlap and a quantitative logit model was developed to relate perfusion deficit in renal parenchyma visualized in verification images as a result of iceball size visualized in monitoring images. Through retrospective study of 20 patient cases, the relationship between likelihood of perfusion loss in renal parenchyma and distance within iceball was quantified and iteratively fit to a logit curve. Using the parameters from the logit fit, the margin for 95% perfusion loss likelihood was found to be 4.28 mm within the iceball. The observed margin corresponds well with the clinically accepted margin of 3-5mm within the iceball. In order to display the iceball position and perfusion loss likelihood to the radiologist, algorithms were implemented to create a fast segmentation and registration module which executed in under 2 minutes, within the clinically-relevant 3 minute monitoring period. Using 16 patient cases, the average Hausdorff distance was reduced from 10.1mm to 3.21 mm with average DSC increased from 46.6% to 82.6% before and after registration.

Analysis of Agrobacterium tumefaciens VirB6 and VirB9 of the VirB /D4 type IV secretion system

Agrobacterium tumefaciens uses the VirB/D4 type IV secretion system (T4SS) to translocate oncogenic DNA (T-DNA) and protein substrates to plant cells. Independent of VirD4, the eleven VirB proteins are also essential for elaboration of a conjugative pilus termed the T pilus. The focus of this thesis is the characterization and analysis of two VirB proteins, VirB6 and VirB9, with respect to substrate translocation and T pilus biogenesis. Observed stabilizing effects of VirB6 on other VirB subunits and results of protein-protein interaction studies suggest that VirB6 mediates assembly of the secretion machine and T pilus through interactions with VirB7 and VirB9. Topology studies support a model for VirB6 as a polytopic membrane protein with a periplasmic N terminus, a large internal periplasmic loop, five transmembrane segments, and a cytoplasmic C terminus. Topology studies and Transfer DNA immunoprecipitation (TrIP) assays identified several important VirB6 functional domains: (i) the large internal periplasmic loop mediates interaction of VirB6 with the T-DNA, (ii) the membrane spanning region carboxyl-terminal to the large periplasmic loop mediates substrate transfer from VirB6 to VirB8, and (iii) the terminal regions of VirB6 are required for substrate transfer to VirB2 and VirB9. To analyze structure-function relationships of VirB9, the phenotypic consequences of dipeptide insertion mutations were characterized. Substrate discriminating mutations were shown to selectively export the oncogenic T-DNA and VirE2 to plant cells or a mobilizable IncQ plasmid to bacterial cells. Mutations affecting VirB9 interactions with VirB7 and VirB10 were localized to the C- and N- terminal regions respectively. Additionally, “uncoupling” mutations identified in VirB11 and VirB6 that block T pilus assembly, but not substrate transfer to recipient cells, were also identified in VirB9. These results in conjunction with computer analysis establish that VirB9, like VirB6, is also composed of distinct regions or domains that contribute in various ways to secretion channel activity and T pilus assembly. Lastly, in vivo immunofluorescent studies suggest that VirB9 localizes to the outer membrane and may play a role similar to that of secretion/ushers of types II and III secretion systems to facilitate substrate translocation across this final bacterial barrier. ^