Human sulfotransferases and their role in chemical metabolism

Sulfonation is an important reaction in the metabolism of numerous xenobiotics, drugs, and endogenous compounds. A supergene family of enzymes called sulfotransferases (SULTs) catalyze this reaction. In most cases, the addition of a sulfonate moiety to a compound increases its water solubility and decreases its biological activity. However, many of these enzymes are also capable of bioactivating procarcinogens to reactive electrophiles. In humans three SULT families, SULT1, SULT2, and SULT4, have been identified that contain at least thirteen distinct members. SULTs have a wide tissue distribution and act as a major detoxification enzyme system in adult and the developing human fetus. Nine crystal structures of human cytosolic SULTs have now been determined, and together with site-directed mutagenesis experiments and molecular modeling, we are now beginning to understand the factors that govern distinct but overlapping substrate specificities. These studies have also provided insight into the enzyme kinetics and inhibition characteristics of these enzymes. The regulation of human SULTs remains as one of the least explored areas of research in the field, though there have been some recent advances on the molecular transcription mechanism controlling the individual SULT promoters. Interindividual variation in sulfonation capacity may be important in determining an individual's response to xenobiotics, and recent studies have begun to suggest roles for SULT polymorphism in disease susceptibility. This review aims to provide a summary of our present understanding of the function of human cytosolic sulfotransferases.

Molecular dissection of the Munc18c/syntaxin4 interaction: Implications for regulation of membrane trafficking

Sec1p/Munc18 (SM) proteins are believed to play an integral role in vesicle transport through their interaction with SNAREs. Different SM proteins have been shown to interact with SNAREs via different mechanisms, leading to the conclusion that their function has diverged. To further explore this notion, in this study, we have examined the molecular interactions between Munc18c and its cognate SNAREs as these molecules are ubiquitously expressed in mammals and likely regulate a universal plasma membrane trafficking step. Thus, Munc18c binds to monomeric syntaxin4 and the N-terminal 29 amino acids of syntaxin4 are necessary for this interaction. We identified key residues in Munc18c and syntaxin4 that determine the N-terminal interaction and that are consistent with the N-terminal binding mode of yeast proteins Sly1p and Sed5p. In addition, Munc18c binds to the syntaxin4/SNAP23/VAMP2 SNARE complex. Pre-assembly of the syntaxin4/Munc18c dimer accelerates the formation of SNARE complex compared to assembly with syntaxin4 alone. These data suggest that Munc18c interacts with its cognate SNAREs in a manner that resembles the yeast proteins Sly1p and Sed5p rather than the mammalian neuronal proteins Munc18a and syntaxin1a. The Munc18c-SNARE interactions described here imply that Munc18c could play a positive regulatory role in SNARE assembly.

Oscillatory regulation of hes1: Discrete stochastic delay modelling and simulation

Discrete stochastic simulations are a powerful tool for understanding the dynamics of chemical kinetics when there are small-to-moderate numbers of certain molecular species. In this paper we introduce delays into the stochastic simulation algorithm, thus mimicking delays associated with transcription and translation. We then show that this process may well explain more faithfully than continuous deterministic models the observed sustained oscillations in expression levels of hes1 mRNA and Hes1 protein.

Accelerated leap methods for simulating discrete stochastic chemical kinetics

Biologists are increasingly conscious of the critical role that noise plays in cellular functions such as genetic regulation, often in connection with fluctuations in small numbers of key regulatory molecules. This has inspired the development of models that capture this fundamentally discrete and stochastic nature of cellular biology - most notably the Gillespie stochastic simulation algorithm (SSA). The SSA simulates a temporally homogeneous, discrete-state, continuous-time Markov process, and of course the corresponding probabilities and numbers of each molecular species must all remain positive. While accurately serving this purpose, the SSA can be computationally inefficient due to very small time stepping so faster approximations such as the Poisson and Binomial τ-leap methods have been suggested. This work places these leap methods in the context of numerical methods for the solution of stochastic differential equations (SDEs) driven by Poisson noise. This allows analogues of Euler-Maruyuma, Milstein and even higher order methods to be developed through the Itô-Taylor expansions as well as similar derivative-free Runge-Kutta approaches. Numerical results demonstrate that these novel methods compare favourably with existing techniques for simulating biochemical reactions by more accurately capturing crucial properties such as the mean and variance than existing methods.

Mathematical modelling and optimal multivariable control of chemical processes

This work reports the developnent of a mathenatical model and distributed, multi variable computer-control for a pilot plant double-effect climbing-film evaporator. A distributed-parameter model of the plant has been developed and the time-domain model transformed into the Laplace domain. The model has been further transformed into an integral domain conforming to an algebraic ring of polynomials, to eliminate the transcendental terms which arise in the Laplace domain due to the distributed nature of the plant model. This has made possible the application of linear control theories to a set of linear-partial differential equations. The models obtained have well tracked the experimental results of the plant. A distributed-computer network has been interfaced with the plant to implement digital controllers in a hierarchical structure. A modern rnultivariable Wiener-Hopf controller has been applled to the plant model. The application has revealed a limitation condition that the plant matrix should be positive-definite along the infinite frequency axis. A new multi variable control theory has emerged fram this study, which avoids the above limitation. The controller has the structure of the modern Wiener-Hopf controller, but with a unique feature enabling a designer to specify the closed-loop poles in advance and to shape the sensitivity matrix as required. In this way, the method treats directly the interaction problems found in the chemical processes with good tracking and regulation performances. Though the ability of the analytical design methods to determine once and for all whether a given set of specifications can be met is one of its chief advantages over the conventional trial-and-error design procedures. However, one disadvantage that offsets to some degree the enormous advantages is the relatively complicated algebra that must be employed in working out all but the simplest problem. Mathematical algorithms and computer software have been developed to treat some of the mathematical operations defined over the integral domain, such as matrix fraction description, spectral factorization, the Bezout identity, and the general manipulation of polynomial matrices. Hence, the design problems of Wiener-Hopf type of controllers and other similar algebraic design methods can be easily solved.

Innovation in the pharmaceutical industry:a study of the effects of regulation on the U.K. pharmaceutical industry

A history of government drug regulation and the relationship between the pharmaceutical companies in the U.K. and the licensing authority is outlined. Phases of regulatory stringency are identified with the formation of the Committees on Safety of Drugs and Medicines viewed as watersheds. A study of the impact of government regulation on industrial R&D activities focuses on the effects on the rate and direction of new product innovation. A literature review examines the decline in new chemical entity innovation. Regulations are cited as a major but not singular cause of the decline. Previous research attempting to determine the causes of such a decline on an empirical basis is given and the methodological problems associated with such research are identified. The U.K. owned sector of the British pharmaceutical industry is selected for a study employing a bottom-up approach allowing disaggregation of data. A historical background to the industry is provided, with each company analysed or a case study basis. Variations between companies regarding the policies adopted for R&D are emphasised. The process of drug innovation is described in order to determine possible indicators of the rate and direction of inventive and innovative activity. All possible indicators are considered and their suitability assessed. R&D expenditure data for the period 1960-1983 is subsequently presented as an input indicator. Intermediate output indicators are treated in a similar way and patent data are identified as a readily-available and useful source. The advantages and disadvantages of using such data are considered. Using interview material, patenting policies for most of the U.K. companies are described providing a background for a patent-based study. Sources of patent data are examined with an emphasis on computerised systems. A number of searches using a variety of sources are presented. Patent family size is examined as a possible indicator of an invention's relative importance. The patenting activity of the companies over the period 1960-1983 is given and the variation between companies is noted. The relationship between patent data and other indicators used is analysed using statistical methods resulting in an apparent lack of correlation. An alternative approach taking into account variations in company policy and phases in research activity indicates a stronger relationship between patenting activity, R&D Expenditure and NCE output over the period. The relationship is not apparent at an aggregated company level. Some evidence is presented for a relationship between phases of regulatory stringency, inventive and innovative activity but the importance of other factors is emphasised.

H2S regulation of nitric oxide metabolism

Nitric oxide (NO) and hydrogen sulfide (H2S) are two major gaseous signaling molecules that regulate diverse physiological functions. Recent publications indicate the regulatory role of H2S on NO metabolism. In this chapter, we discuss the latest findings on H2S-NO interactions through formation of novel chemical derivatives and experimental approaches to study these adducts. This chapter also addresses potential H2S interference on various NO detection techniques, along with precautions for analyzing biological samples from various sources. This information will facilitate critical evaluation and clearer insight into H2S regulation of NO signaling and its influence on various physiological functions.

Sex steroid feedback regulation of pituitary gonadotropins during early secondary oocyte growth in coho salmon (Oncorhynchus kisutch) – a potential target of endocrine disruption.

Thesis (Ph.D.)--University of Washington, 2016-06

Physico-chemical assessment of sediments of Tamarindo estuary and its tributaries, Guanacaste, Costa Rica

Sediments of Tamarindo estuary and its tributaries were evaluated in order to determine the effect of anthropogenic activities of the surrounding communities. The evaluation relied on texture, total phosphorus, organic matter, and heavy metal (Pb, Cu, Ni and Cr) analyses. The study spanned four sampling campaigns between October 2007 and April 2008, including four climatic seasons: rainy season, rainy-dry transition, dry season and dryrainy transition. Five sampling points were selected at Tamarindo estuary and three additional sites were set at Matapalo, Lomas and Lajas rivers. Ontario Ministry of the Environment regulation was used to evaluate sediment quality. It was determined that theanthropogenic activities of the surrounding communities exert a negative impact over the sediment quality; the pollution effect caused by such activities was rated as lower effect.

REGULATION OF SHAPE DYNAMICS AND ACTIN POLYMERIZATION DURING COLLECTIVE CELL MIGRATION

This thesis aims to understand how cells coordinate their motion during collective migration. As previously shown, the motion of individually migrating cells is governed by wave-like cell shape dynamics. The mechanisms that regulate these dynamic behaviors in response to extracellular environment remain largely unclear. I applied shape dynamics analysis to Dictyostelium cells migrating in pairs and in multicellular streams and found that wave-like membrane protrusions are highly coupled between touching cells. I further characterized cell motion by using principle component analysis (PCA) to decompose complex cell shape changes into a serial shape change modes, from which I found that streaming cells exhibit localized anterior protrusion, termed front narrowing, to facilitate cell-cell coupling. I next explored cytoskeleton-based mechanisms of cell-cell coupling by measuring the dynamics of actin polymerization. Actin polymerization waves observed in individual cells were significantly suppressed in multicellular streams. Streaming cells exclusively produced F-actin at cell-cell contact regions, especially at cell fronts. I demonstrated that such restricted actin polymerization is associated with cell-cell coupling, as reducing actin polymerization with Latrunculin A leads to the assembly of F-actin at the side of streams, the decrease of front narrowing, and the decoupling of protrusion waves. My studies also suggest that collective migration is guided by cell-surface interactions. I examined the aggregation of Dictyostelim cells under distinct conditions and found that both chemical compositions of surfaces and surface-adhesion defects in cells result in altered collective migration patterns. I also investigated the shape dynamics of cells suspended on PEG-coated surfaces, which showed that coupling of protrusion waves disappears on touching suspended cells. These observations indicate that collective migration requires a balance between cell-cell and cell-surface adhesions. I hypothesized such a balance is reached via the regulation of cytoskeleton. Indeed, I found cells actively regulate cytoskeleton to retain optimal cell-surface adhesions on varying surfaces, and cells lacking the link between actin and surfaces (talin A) could not retain the optimal adhesions. On the other hand, suspended cells exhibited enhanced actin filament assembly on the periphery of cell groups instead of in cell-cell contact regions, which facilitates their aggregation in a clumping fashion.

Sexual selection in a tropical fruit fly : role of a plant derived chemical in mate choice

The specific mechanisms by which selective pressures affect individuals are often difficult to resolve. In tephritid fruit flies, males respond strongly and positively to certain plant derived chemicals. Sexual selection by female choice has been hypothesized as the mechanism driving this behaviour in certain species, as females preferentially mate with males that have fed on these chemicals. This hypothesis is, to date, based on studies of only very few species and its generality is largely untested. We tested the hypothesis on different spatial scales (small cage and seminatural field-cage) using the monophagous fruit fly, Bactrocera cacuminata. This species is known to respond to methyl eugenol (ME), a chemical found in many plant species and one upon which previous studies have focused. Contrary to expectation, no obvious female choice was apparent in selecting ME-fed males over unfed males as measured by the number of matings achieved over time, copulation duration, or time of copulation initiation. However, the number of matings achieved by ME-fed males was significantly greater than unfed males 16 and 32 days after exposure to ME in small cages (but not in a field-cage). This delayed advantage suggests that ME may not influence the pheromone system of B. cacuminata but may have other consequences, acting on some other fitness consequence (e.g., enhancement of physiology or survival) of male exposure to these chemicals. We discuss the ecological and evolutionary implications of our findings to explore alternate hypotheses to explain the patterns of response of dacine fruit flies to specific plant-derived chemicals.