Klonierung und funktionelle Charakterisierung des Extraneuronalen Monoamin-Transporters (EMT) des Hausschweins

Together with the other members of the non-neuronal monoamine-transporter family, EMT is responsible for the re-uptake of catecholamines and monoamines after their release into the synaptic cleft and from the circulation. EMT in fact transports other organic cations and its expression at excretion organs designates it to a further role: that of an Organic Cation-Transporter, which allows excretion of xenobiotics and physiological metabolites.From the first cloning of EMT in 1998 until now a lot has been discovered concerning its substrates and localization. This stays in contrast to how it is regulated, which is an important clue to clarify its physiological and pathophysiological role. In this area, facts are missing.For that there was a need to provide the basics for an interspecific promoter comparison to identify conserved areas which are strong indices to be relevant for regulation. To allow this comparison to be a powerful tool for identifying “clue-regions”, EMT of a further species had to be cloned.In the course of my work I succeeded in cloning the open reading frame (ORF) of the porcine EMT for the first time. Starting from the cDNA an EMTp (p: porcine)- expressing cell line has been created that allowed me to characterize the functional protein. In doing so an extensive similarity to the human EMT turned out which is consistent with that on nucleic acid basis. Beyond this it has been shown that acetylcholine is an EMT substrate. This indicates that EMT is possibly involved in the “non-neuronal acetylcholine system”.The comparison of the recorded substance spectrum between transporter and control cell line showed after incubation with DMEM/FCS a clear and reproducible signal at m/z 254. As up to now there is no known EMT substrate having such mass, it is maybe a new EMT substrate