Cellular Creatine/Phosphocreatine Homeostasis

Florida State University, Tallahassee FL

Investigators

Abstract

The basic laws of thermodynamics apply to organisms as they do to the physical world. As a consequence, all living cells must metabolically breakdown large organic molecules such as carbohydrates and in the process trap some of the energy released in the form of a compound known as adenosine triphosphate, ATP for short. ATP functions as the cell's energy currency. Its chemical breakdown is used to drive a variety of energetically unfavorable processes such as biosynthesis, cellular movement like contraction, and pumping substances uphill across biological membranes. There are instances where demand for ATP exceeds supply as might take place at the onset of burst muscle contraction. In many cells a compound known as phosphocreatine is present which, through the action of an enzyme called creatine kinase, is capable of replenishing ATP as quickly as it is used (at least in the short term). This energy buffering role of phosphocreatine is critical in the functioning of a broad spectrum of cells. The precursor for phosphocreatine is creatine. In vertebrates, a fraction of the creatine required is obtained from the diet while the rest is synthesized in the liver and pancreas. Creatine is then carried in the blood to cells where it is transported by a special membrane transport protein (creatine transporter) into the cell and then converted to phosphocreatine. Genetic defects in creatine biosynthesis and transport produce severe pathological effects in humans including profound mental retardation. Many diverse invertebrate groups accumulate large quantities of creatine/phosphocreatine in their cells. This is particularly true of certain marine species such as sea urchins which are broad-cast fertilizers. These animals literally shed their eggs and sperm into to the sea water; this strategy necessitates production of massive amounts of sperm which have very high levels of creatine/phosphocreatine. Thus, there is a seasonally high demand for creatine yet it is not clear at all how these animals obtain and transport this vital substance to cells where it is needed. The proposed research effort seeks to trace the evolution of creatine biosynthetic and membrane transport capacities by investigating these processes in selected groups of invertebrates. Initial efforts will focus on 1) using sensitive isotopic labeling techniques to determine whether the two key enzymes of creatine biosynthesis are present in these animals; and 2) using modern molecular genetic approaches to localize the tissue expression of these enzymes, and to determine whether there is increased presence of these proteins at peak reproductive activity (when demand for creatine is high). A second facet of this research will center on determining the nature of creatine transport into cells where it accumulates as high concentrations of creatine/phosphocreatine. Once again, molecular genetic methods will be used to express the transport proteins in a cell system where they can be studied and characterized most efficiently. The overall results will yield mechanistic information about creatine biosynthesis and transport as well as provide a picture of the evolution of these processes from the lower invertebrates to more advanced animals.