CHARACTERIZATION OF SERINE KINASE ACTIVITY ASSOCIATED WITH MOLONEY MURINE SARCOMA VIRUS V-MOS GENE PRODUCTS

Various Moloney murine sarcoma virus (Mo-MuSV) isolates contain a cellular sequence, termed mos, which is responsible for the transforming ability of Mo-MuSV. A serine kinase activity has been found to be associated with mos gene products of several isolates of Mo-MuSV. A mutant of Mo-MuSV strain 124 (designated MuSV ts110) is temperature-sensitive (ts) for transformation and encodes two proteins, P85('gag-mos) (an 85,000 M(,r) protein encoded by the gag and mos genes) and P58('gag), at the permissive temperature (28(DEGREES)C). At the nonpermissive temperature (39(DEGREES)C), only P58('gag) is found in MuSV ts110-infected NRK cells (6m2 cells). Both P85('gag-mos) and P58('gag) were phosphorylated when anti-gag immune complexes containing these proteins were incubated at 22(DEGREES)C with (lamda)-('32)P -ATP and MnCl(,2). The kinase detected in anti-gag complexes from 6m2 cells at permissive temperature was associated with P85('gag-mos) since immune complexes from 39(DEGREES)C 6m2 cells, which lack P85('gag-mos), produced no phosphorylated P58('gag) molecules. In addition, an anti-mos complex (anti-mos 37-55 complexes) allowed in vitro phosphorylation of P85('gag-mos) in the absence of P58('gag). No kinase activity was detectable with other gag gene products (e.g., Mo-MuSV-124 P62('gag)), suggesting that the P85('gag-mos) kinase activity was present within the mos portion of the protein. The P85('gag-mos) kinase activity was very thermolabile upon shifting 6m2 cells from permissive to nonpermissive temperatures (t(, 1/2) for inactivation = 5 min). In contrast, a spontaneous revertant of MuSV ts110 encodes a larger gag-mos protein (termed P100('gag-mos)) which contained a kinase activity stable to 39(DEGREES)C. Using the optimal conditions developed for the P85('gag-mos) kinase, Mo-MuSV-encoded p37('mos) was also found to be associated with a serine kinase activity. Phosphorylation of p37('mos) and a 43 Kd protein (super-phosphorylated p37('mos)) occurred in anti-mos(37-55) complexes from Mo-MuSV-124 acutely-infected NIH 3T3 cells, but neither in mos 37-55 peptide-blocked anti-mos(37-55) complexes nor in immune complexes from uninfected NIH 3T3 cells. Antibodies directed against the C-terminus of v-mos were found to inhibit the in vitro phosphorylation of p37('mos), suggesting that the extreme C-terminal sequence of v-mos may be important for an intrinsic kinase activity. This inhibitory action by antibodies to the C-terminus of p37('mos), when considered with all the other data reported here, provides convincing evidence that the v-mos gene encodes a serine protein kinase activity. ^

STRUCTURAL ANALYSIS OF THE TRANSCRIPTION PRODUCTS OF A TEMPERATURE-SENSITIVE PHENOTYPIC MUTANT OF MOLONEY SARCOMA VIRUS--MUSV TS110

Cells infected with a temperature sensitive phenotypic mutant of Moloney sarcoma virus (MuSVts110) exhibit a transformed phenotype at 33('(DEGREES)) and synthesize two virus specific proteins, p85('gag-mos), a gag-mos fusion protein and p58('gag), a truncated gag precursor protein (the gag gene codes for viral structural proteins and mos is the MuSV transforming gene). At 39('(DEGREES)) only p58('gag) is synthesized and the morphology of the cells is similar to uninfected NRK parental cells. Two MuSVts110 specific RNAs are made in MuSVts110-infected cells, one of 4.0 kb in length, the other of 3.5 kb. Previous work indicated that each of these RNAs arose by a single central deletion of parental MuSV genetic material, and that p58('gag) was made by the 4.0 kb RNA and p85('gag-mos) from the 3.5 kb RNA. The objective of my dissertation research was to map precisely the deletion boundaries of both of the MuSVts110 RNAs, and to determine the proper reading frame across both deletion borders. This work succeeded in arriving at the following conclusions: (a) Using S-1 nuclease analysis and primer extension sequencing, it was found that the 4.0 kb MuSVts110 RNA arose by a 1488 base deletion of 5.2 kb parental MuSV genomic RNA. This deletion resulted in an out of frame fusion of the gag and mos genes that resulted in the formation of a "stop" codon which causes termination of translation just beyond the c-terminus of the gag region. Thus, this RNA can only be translated into the truncated gag protein p58('gag). (b) S-1 analysis of RNA from cells cultivated at different temperatures demonstrated that the 4.0 kb RNA was synthesized at all temperatures but that synthesis of the 3.5 kb RNA was temperature sensitive. These observations supported the data derived from blot hybridization experiments the interpretation of which argued for the existence of a single provirus in MuSVts110 infected cells, and hence only a single primary transcript (the 4.0 kb RNA). (c) Analyses similar to those described in (a) above showed that the 3.5 kb RNA was derived from the 4.0 kb MuSVts110 RNA by a further deletion of 431 bases, fusing the gag and mos genes into a continuous reading frame capable of directing synthesis of the p85('gag-mos) protein. These sequence data and the presence of only one MuSVts110-specific provirus, indicate that a splice mechanism is employed to generate the 3.5 kb RNA since the gag and mos genes are observed to be fused in frame in this RNA. . . . (Author's abstract exceeds stipulated maximum length. Discontinued here with permission of author.) UMI ^

Collective Voluntary Agreements and the Production of Less Polluting Products

Recently, some industries have collectively agreed not to produce models that do not meet an energy efficiency (and hence an environmental) standard. This paper presents a simple model that can be used to examine a voluntary collective agreement to limit or completely eliminate the low efficiency model of a given product (e.g., a low efficiency washing machine). We show that, when there is competition between firms, a collective agreement to limit or even eliminate production of the polluting model can actually increase profits for all firms in the industry. This suggests that a collective agreement of this type might actually be beneficial to firms, while at the same time improving environmental quality. However, the implicit enforcement that comes from the public nature of the commitment is necessary to ensure this outcome. This suggests that, by promoting such agreements, policymakers may be able to achieve substantial environmental gains with relatively little inducement. The impact on social welfare will then depend on whether these gains are sufficiently large to offset consumer losses from reductions in product variety and the associated price increases.

Creating Useful Products From Connecticut's 2000 LIDAR Data Set JHR 08-314 Project 07-2

The State of Connecticut owns a LIght Detection and Ranging (LIDAR) data set that was collected in 2000 as part of the State’s periodic aerial reconnaissance missions. Although collected eight years ago, these data are just now becoming ready to be made available to the public. These data constitute a massive “point cloud”, being a long list of east-north-up triplets in the State Plane Coordinate System Zone 0600 (SPCS83 0600), orthometric heights (NAVD 88) in US Survey feet. Unfortunately, point clouds have no structure or organization, and consequently they are not as useful as Triangulated Irregular Networks (TINs), digital elevation models (DEMs), contour maps, slope and aspect layers, curvature layers, among others. The goal of this project was to provide the computational infrastructure to create a first cut of these products and to serve them to the public via the World Wide Web. The products are available at http://clear.uconn.edu/data/ct_lidar/index.htm.

Instrumental variable method for the causal relationship between soluble receptor for advanced glycation end-products (sRAGE) and risk of pancreatic cancer

This thesis project is motivated by the potential problem of using observational data to draw inferences about a causal relationship in observational epidemiology research when controlled randomization is not applicable. Instrumental variable (IV) method is one of the statistical tools to overcome this problem. Mendelian randomization study uses genetic variants as IVs in genetic association study. In this thesis, the IV method, as well as standard logistic and linear regression models, is used to investigate the causal association between risk of pancreatic cancer and the circulating levels of soluble receptor for advanced glycation end-products (sRAGE). Higher levels of serum sRAGE were found to be associated with a lower risk of pancreatic cancer in a previous observational study (255 cases and 485 controls). However, such a novel association may be biased by unknown confounding factors. In a case-control study, we aimed to use the IV approach to confirm or refute this observation in a subset of study subjects for whom the genotyping data were available (178 cases and 177 controls). Two-stage IV method using generalized method of moments-structural mean models (GMM-SMM) was conducted and the relative risk (RR) was calculated. In the first stage analysis, we found that the single nucleotide polymorphism (SNP) rs2070600 of the receptor for advanced glycation end-products (AGER) gene meets all three general assumptions for a genetic IV in examining the causal association between sRAGE and risk of pancreatic cancer. The variant allele of SNP rs2070600 of the AGER gene was associated with lower levels of sRAGE, and it was neither associated with risk of pancreatic cancer, nor with the confounding factors. It was a potential strong IV (F statistic = 29.2). However, in the second stage analysis, the GMM-SMM model failed to converge due to non- concaveness probably because of the small sample size. Therefore, the IV analysis could not support the causality of the association between serum sRAGE levels and risk of pancreatic cancer. Nevertheless, these analyses suggest that rs2070600 was a potentially good genetic IV for testing the causality between the risk of pancreatic cancer and sRAGE levels. A larger sample size is required to conduct a credible IV analysis.^

Effect of Certified Organic Products on Soybean Aphid

The soybean aphid (Aphis glycines), native to China, has become the most economically damaging insect in soybeans in northeast Iowa. Soybean aphid may have up to 18 generations per year, beginning with overwintering eggs on the alternate host buckthorn. In spring, winged aphids migrate from buckthorn to nearby emerged soybeans. Generations advance in these fields, and then another winged migration occurs in summer spreading from these fields to others. A third migration occurs in fall with aphids moving back to buckthorn. Depending on the season, soybean proximity to buckthorn, and soybean aphid migration patterns, populations of aphids tend to peak in soybeans anywhere from late July to early September. With higher aphid populations, the production of honeydew (the excrement of the aphid) and the resulting black fungus that grows on it (sooty mold) may become apparent. Aphid feeding may cause stunted plants, reduced pods and seeds, and may also transmit viruses that could cause mottling and distortion of leaves, reduced seed set, and discolored seeds.

Tecnologías de conservación por métodos combinados en pimiento, chaucha y berenjena

La tecnología de los métodos combinados permite reducir la intensidad del tratamiento térmico y mantener las propiedades organolépticas en el producto final, mediante una combinación de obstáculos que aseguran la estabilidad y seguridad microbiana. El objetivo de este trabajo fue aplicar dicha tecnología en la conservación de tres hortalizas: pimiento, chaucha y berenjena y analizar la calidad y vida útil de los productos obtenidos. La elaboración de las conservas se efectuó del siguiente modo: las hortalizas fueron cortadas y posteriormente escaldadas en solución de ácidos cítrico, láctico y acético. Terminada esta etapa se envasaron con la solución de relleno, constituida por los ácidos de la solución de escaldado con la adición de ácido ascórbico. Los análisis que se realizaron tanto en el material fresco como en los productos procesados fueron: humedad, pH, hidratos de carbono, lípidos, proteína, vitamina C, °Brix, actividad enzimática y acidez total. En los productos ya procesados se realizó el control microbiológico. Los resultados indican que la aplicación de los métodos combinados permite conservar los productos elaborados a temperatura ambiente y se mantiene su seguridad microbiológica.