FUNCTIONAL SIGNIFICANCE OF INTRACELLULAR REGULATION OF GLUTAMINE TRANSPORTERS IN NORMAL AND MALIGNANT CELLS

Glutamine, the most abundant amino acid in plasma and cerebrospinal fluid, is important for many metabolic processes, e.g. biosynthesis of amino acids, purines and pyrimidines. Glutamine is a major N- and C-donor involved in inter-organ and inter-cellular shuttling of metabolites. It is a signaling molecule as transport into cells has direct effects on cell function. We have molecularly characterized a family of amino acid transporters with high affinity for glutamine. Transport of glutamine by SN1 and SN 2 is electroneutral and bidirectional due to coupling to Na+ in symport and H+ in antiport. In contrast, SAT1 and SAT2, that are unable to bind H+, carry unidirectional transport accumulating glutamine inside the cells. SN1 is regulated through several me chanisms: through proton action directly on the SN1 protein, at the protein synthesis level, and by stabilization of mRNA via the pH-response element. The level of SN1 protein is increased in glioblastoma multiforme, and SN1-mediated glutamine transport i nto anaplastic cells may be a limiting factor for growth and metastatic ability. We therefore want to study regulation of SN1 and SN2 in normal cells and in tumors. We want to explore intracellular modifying proteins/enzymes acting on SN1 or SN2 and poten tial linkage to oncogenes. Regulation by phosphorylation and the target residues will be characterized, before site-directed mutagenesis and expression in heterologous systems is performed to map the effects on expression levels, membrane trafficking and functional activity. Pathways of controlled protein degradation will be studied in normal, neoplastic and apoptotic cells. Additional regulatory mechanisms may be revealed in SN1 and SN2-deficient mice. The project aims to give insight into cell growth an d development of neoplasia. The data may explain functional changes in the tumor surroundings, such as altered synaptic transmission in the vicinity of CNS tumors leading to development of novel therapeutic schemes.