Calorie availability pivotally affects metabolic rate, reproductive fitness, growth, and survival in fish (Tocher, 2003). Calories in the form of fatty acids are the most significant source of ATP for many species of fish. Accordingly, fish manipulate storage and mobilization of fatty acids as part of their natural history resulting in a variety of outcomes. Striped bass increase intracellular lipid droplets 13-fold in red muscle during cold acclimation (Egginton and Sidell, 1989) and salmon increase serum cholesterol without eating during spawning migrations (Farrell and Munt, 1983).

The expression of fat metabolism genes is regulated by free long chain fatty acids and their metabolic by-products, however the definitive mechanism by which they do so remains elusive (Duplus et al., 2000). Characterizing the molecular signaling behind these changes in fat metabolism has traditionally been approached by looking at candidate proteins, (e.g. fatty acid binding proteins (FABPs) (Londraville and Sidell, 1995)) organelle function (peroxisomes (Crockett and Sidell, 1993)), and enzymatic indicators of fatty acid flux (Sidell et al., 1995). However, these are specific indicators, not an integrated global mechanism. One family of transcription factors, the peroxisome proliferated-activated receptors (PPARs), has the potential to attach a mechanism to the metabolic changes detailed above.

Acting as lipid sensors, PPARs are a nuclear hormone receptor superfamily of transcriptional regulators that are responsive to polyunsaturated fatty acids (PUFAs) (Issemann and Green, 1990), and their metabolic derivatives that act to orchestrate metabolism (Evans et al., 2004). Various PPAR isoforms have been linked to regulation of lipid metabolism from their associated pathologies in mammals (Bilsen et al., 2002). PPAR? is a critical mediator in the rodent liver, responding to perturbation of lipid metabolism but also directly and indirectly directing genes involved in hepatocyte growth (Corton et al., 2000). PPAR’s from fish were first identified in Atlantic salmon (Ruyter et al., 1997) and subsequently cloned (Andersen et al., 2000). PPAR’s have also been identified in zebrafish (Escriva et al., 1997) with several homologous mammalian isoforms (PPAR?, PPAR?, and PPAR?) being conclusively identified by immunohistochemistry (Ibabe et al., 2002). I hypothesized that high-calorie diets in zebrafish will up-regulate activity of PPARs. I tested this hypothesis using proteomics to identify which proteins change with diet. I did not find an up-regulation of proteins under the control of PPARs but I did identify several proteins associated with stress.

tudying the effects of a single protein in isolation is limited in explanatory power because of the ways proteins interact in vivo. Proteomics quantitatively estimates alterations in protein expression within a tissue at a moment in time (Abbott, 1999). Its' power is that it focuses on the part of the genome that is expressed, (including the unexpected), thus more effectively estimating differences in expression among different groups or experimental treatments. Proteomics’ utility is in sifting through thousands of proteins to find likely targets for further study. My approach was a hypothesis-generating step, to find these targets that could then be rigorously characterized with immunoblotting, nucleic acid hybridization, cloning, and sequence-functional expression in future studies.

CONTENTS

LIST OF TABLES
LIST OF FIGURES
CHAPTER

I. INTRODUCTION
II. MATERIALS AND METHODS
III. RESULTS
IV. DISCUSSION

REFERENCES

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