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The human c-myc gene is a cellular proto-oncogene composed of three exons and two introns. Transcription of c-myc is controlled by two promoters, Pl and P2. The activity of these promoters is regulated by many factors, such as cellular transcription factors E2F, YYl, and HSV-1 immediate-early proteins, ICPO, ICP4. Many regulatory elements located both upstream of and between P 1 and P2 have been identified, and some of these are required for optimum expression of c-myc. In this thesis research, a region downstream from P2 in the c-myc exon 1 was identified by its response to transactivation by HSV-1 immediate-early proteins, ICPO and ICP4. The purpose of this research was to examine this region for regulatory sites that respond to HSV-1 infection. I hypothesized that after HSV-1 infection, ICPO and ICP4 activate c-myc expression, in part, through regulatory sequences present in exon 1. To test for this hypothesis, reporter plasmids containing (I) the c-myc promoter (from - 101 bp relative to Pl) and exon 1 coupled to the bacterial CAT gene were constructed. (ii) The c-myc exon sequences used were either intact (wild-type) or they were constructed with various deletions. The activities of these plasmids were examined in transient expression assays. To analyze protein binding, electrophoretic mobility shift assay (EMSA) and completion EMSAs were carried out. The results from these experiments lead to the following conclusions: (i) ICP4 and ICPO serve as activators, whereas ICP27 inhibits c-myc gene expression. (ii) The region from +332 to +513 within the c-myc exon 1 contains an important element required for transactivation of the c-myc gene by HSV-1 proteins. (iii) Cellular proteins, including factor YYl, bind to the region from +332 to +513 in the c-myc exon 1. Although the exact mechanism by which HSV-1 immediate-early proteins regulate cmyc gene expression is still not clear, it gives rise to a possibility that this regulation is caused by turning on or activation of the cellular regulatory proteins by ICP4 and ICPO. The cellular proteins in turn activate the c-myc gene expression by interacting with the ciselement downstream from P2.

A regulatory element involved in the transcriptional activation of the major capsid protein (VP5) of herpes simplex virus type 1 was identified and characterized in this research project. Gel mobility shift assay with nuclear extracts from both uninfected and HSV-1 infected HeLa cells identified two major protein-DNA complexes involving the VP5 promoter. No viral specific complex found. DNase I and orthophenanthroline-cu+ footprint analyses in the same laboratory revealed that the two complexes involve a single binding site, GGCCATCTTGAA, located between -64 and -75 bp relative to the VP5 cap site. To determine the function of this leaky-late binding site (LBS) in VP5 gene activation, mutated VP5 promoters with deletion and insertion around LBS were constructed and linked to a reporter gene, bacterial chloramphenicol acetyltransferase gene. The effect of mutations were tested in transient expression assay. Deletion of LBS resulted in seven to eight-fold reduction in the level of transactivation of the chloramphenicol acetyltransferase gene by superinfection with HSV-1 or by cotransfection of HSV immediate-early genes. These results indicated LBS is involved in the maximal transactivation of the VPS gene. A search of published gene sequences found the homologs of LBS exist in a number of HSV-1 By promoters, and other viral promoters, as well as cellar promoters. Some of these homologs have found involved in the transcription regulation.

The herpes simplex virus 1 (HSVl) VP5 gene codes for the major viral capsid protein. Understanding of the mechanism of how the VP5 gene is regulated in host cells will help us to understand the molecular action of the HSV 1 life cycle and its interplay with the host cell gene expression machinery (transcription and translation). This may ultimately provide scientific bases for both better prevention and cure of HSV 1 caused diseases. Previous work from Dr. Robert L. Millette' s laboratory has indicated that a 164 base pair region of the VP5 promoter gene could activate the transcription of an attached reporter gene (bacteria CAT gene). Furthermore, a 12 bp (GGCCATCTTGAA) cis-acting element situated within the 164 bp promoter region was required for the promoter activity. To understand the function of this cis-element in the regulation of VP5 transcription and to identify the trans-acting factors interacting with this element, gel mobility shift assays were first carried out using the fragment containing the 12 bp site as the probe. A cellular factor, YY 1, was found to bind to this site in a sequence specific manner. Based on the oligonucleotide competition assays, partial protease digestions, and antibody supershift assays, it became clear that two cellular factors bound to the VP5 promoter. These were related, if not identical, to the previously identified Yin-Yang- 1 factor (YY 1), and transcription factor the SPl. Site-directed mutagenesis studies indicated that these two factors bind to distinct sites on the 164 bp fragment. Point mutations studies on the 12 bp YYl binding site demonstrated that seven of the 12 bp were required for YY 1-DNA complex formation and the first four bp in the 12 bp were very important for VP5 gene regulation. Also, it was found that YY 1 performs both positive and negative regulator function in VP5 gene regulation. In conclusion, two cellular transcription factors, YY 1 and SPl, play a major role in VP5 gene expression.

This dissertation has two topics. The rst one is about rotating a supporting hyperplane on the convex hull of a nite point set to arrive at one of its facets. We present three procedures for these rotations in multiple dimensions. The rst two procedures rotate a supporting hyperplane for the polytope starting at a lower dimensional face until the support set is a facet. These two procedures keep current points in the support set and accumulate new points after the rotations. The rst procedure uses only algebraic operations. The second procedure uses LP. In the third procedure we rotate a hyperplane on a facet of the polytope to a dierent adjacent facet. Similarly to the rst procedure, this procedure uses only algebraic operations. Some applications to these procedures include data envelopment analysis (DEA) and integer programming. The second topic is in the eld of containment problems for polyhedral sets. We present three procedures to nd a circumscribing simplex that contains a point set in any dimension. The rst two procedures are based on the supporting hyperplane rotation ideas from the rst topic. The third circumscribing simplex procedure uses polar cones and other geometrical properties to nd facets of a circumscribing simplex. One application of the second topic discussed in this dissertation is in hyperspectral unmixing.

Copies of author's previously published articles inserted inside back cover. / Bibliography: leaves 137-179. / x, 179, [26] leaves, [28] leaves of plates : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / The aims of this study are to determine whether latent HSV genomes are a result of viral DNA amplification in the PNS during the establishment phase and to investigate the relationship between HSV DNA copy number and viral transcriptional activity during latent infection of the PNS. In order to map the distribution of viral nucleic acid sequences in latently infected sensory ganglia, experiments are undertaken using a mouse model that makes novel use of the segmental sensory innervation of flank skin. / Thesis (Ph.D.)--University of Adelaide, Dept. of Microbiology and Immunology, 1995?

Momentum vectors of fragment ions produced by the Coulomb explosion of CO2z+ (z = 3 - 6) and CS2z+ (z = 3 - 13) in an intense laser field (~50 fs, 1 x 1015 W/cm2) are determined by the triple coincidence imaging technique. The molecular structure from symmetric and asymmetric explosion channels is reconstructed from the measured momentum vectors using a novel simplex algorithm that can be extended to study larger molecules. Physical parameters such as bend angle and bond lengths are extracted from the data and are qualitatively described using an enhanced ionization model that predicts the laser intensity required for ionization as a function of bond length using classical, over the barrier arguments. As a way of going beyond the classical model, molecular ionization is examined using a quantum-mechanical, wave function modified ADK method. The ADK model is used to calculate the ionization rates of H2, N2, and CO2 as a function of initial vibrational level of the molecules. A strong increase in the ionization rate, with vibrational level, is found for H2, while N2 and CO2 show a lesser increase. The prospects for using ionization rates as a diagnostic for vibrational level population are assessed.

Momentum vectors of fragment ions produced by the Coulomb explosion of CO2z+ (z = 3 - 6) and CS2z+ (z = 3 - 13) in an intense laser field (~50 fs, 1 x 1015 W/cm2) are determined by the triple coincidence imaging technique. The molecular structure from symmetric and asymmetric explosion channels is reconstructed from the measured momentum vectors using a novel simplex algorithm that can be extended to study larger molecules. Physical parameters such as bend angle and bond lengths are extracted from the data and are qualitatively described using an enhanced ionization model that predicts the laser intensity required for ionization as a function of bond length using classical, over the barrier arguments. As a way of going beyond the classical model, molecular ionization is examined using a quantum-mechanical, wave function modified ADK method. The ADK model is used to calculate the ionization rates of H2, N2, and CO2 as a function of initial vibrational level of the molecules. A strong increase in the ionization rate, with vibrational level, is found for H2, while N2 and CO2 show a lesser increase. The prospects for using ionization rates as a diagnostic for vibrational level population are assessed.

This thesis presents an approach for identifying critical model parameters in dynamic air-conditioning systems using limited sensor information. The expansion valve model and the compressor model parameters play a crucial role in the system model's accuracy. In the past, these parameters have been estimated using a mass flow meter; however, this is an expensive devise and at times, impractical. In response to these constraints, a novel method to estimate the unknown parameters of the expansion valve model and the compressor model is developed. A gray box model obtained by augmenting the expansion valve model, the evaporator model, and the compressor model is used. Two numerical search algorithms, nonlinear least squares and Simplex search, are used to estimate the parameters of the expansion valve model and the compressor model. This parameter estimation is done by minimizing the error between the model output and the experimental systems output. Results demonstrate that the nonlinear least squares algorithm was more robust for this estimation problem than the Simplex search algorithm. In this thesis, two types of expansion valves, the Electronic Expansion Valve and the Thermostatic Expansion Valve, are considered. The Electronic Expansion Valve model is a static model due to its dynamics being much faster than the systems dynamics; the Thermostatic expansion valve model, however, is a dynamic one. The parameter estimation algorithm developed is validated on two different experimental systems to confirm the practicality of its approach. Knowing the model parameters accurately can lead to a better model for control and fault detection applications. In addition to parameter estimation, this thesis also provides and validates a simple usable mathematical model for the Thermostatic expansion valve.

In order to search for bioactive compounds, we have studied the chemical constituents from the organic extracts of one soft corals Klyxum simplex and two gorgonians Subergorgia mollis and Briareum excavatum. This study had led to the isolation of twenty-seven natural compounds 1¡V27, including nineteen new eunicellin¡Vtype diterpenoids, simplexins A¡VS (1¡V19) from K. simplex, one new steroid 11£\,15£\-diacetoxy-17£]-pregna-4,20- dien-3-one (20) along with one known compound 17£]-pregna-4,20-dien-3- one (21) from S. mollis, and six new briarane¡Vtype diterpenoids briaexcavatolides Q (22), S¡VV (23-26) and W (27) from B. excavatum. The structure of these compounds were established by the detailed spectroscopic analysis (IR, MS, 1D¡B2D NMR) and by comparison of the physical and spectral data with those of the related known compounds. The absolute configuration of 1 was determined by using a modified Mosher's method. The cytotoxicity of compounds 1¡V6, 9, 13¡V15 against the MCF-7 (human breast adenocarcinoma), Hep2 (human laryngeal carcinoma), Daoy (human medulloblastoma), and Hela (human cervical epitheloid carcinoma) cancer cell lines were determined. Compounds 1, 4 and 14 showed weak inhibition against the growth of MCF-7, Hep2 and Daoy, but did not inhibit the growth of Hela cells. Compound 5 exhibited a weak cytotoxicity against the growth of MCF-7, Hep2, Hela and Daoy cells. In addition, the activity of compounds 1¡V6, 9, 13¡V15 to inhibit the pro-inflammatory iNOS and COX-2 protein expression in LPS-stimulated RAW264.7 macrophage cells was estimated. Compound 5 showed the best anti-inflammatory activity among the all tested compounds.