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内容記述 |
Receptor-activated Ca2+ influx that occurs as a second phase of phosphatidyl inositol-dependent response, has been recently recognized for its physiological significance. Characterization of mammalian homologues of Drosophila TRP proteins is an important clue to understand molecular mechanisms underlying receptor-activated Ca2+ influx in vertebrate cells. Especially, TRP homologues have been proposed to encode store-operated channels (SOC) which is activated through Ca2+ release from the intracellular Ca2+ store and subsequent depletion of the Ca2+ store. However, the hypothesis is still controversial. In order to establish the correlation of cloned TRP homologues and native channels, it is necessary to investigate the functional properties of TRP homologues using the excellent expression system. He has here isolated cDNAs that encode novel TRP homologues, TRP4, TRP5, TRP6, and TRP7 from the mouse brain. All of these homologues were found to be relatively highly expressed in the brain, but their expression was detected also in various organs other than the brain, with various intensities characteristic of each subtype. Among them, first, he characterized the function of the brain-predominant homologue, TRP5. The recombinant expression of the TRP5 cDNA in human embryonic kidney cells potentiated an extracellular Ca2+-dependent increase of [Ca2+]i evoked by ATP, but not by an inhibitor of ER Ca2+-ATPases, thapsigargin. Whole-cell mode of patch-clamp recordings from TRP5-expressing cells demonstrated that ATP application induced a large inward current in the presence of extracellular Ca2+, which reversed at a positive potential. The TRP5 activity was abolished by omission of intracellular Ca2+ and by treatment with a calmodulin antagonist. High concentration of extracellular Ca2+ activated TRP5 without ATP stimulation. On the other hand, omission of extracellular Ca2+ abolished the activity even in the presence of ATP. These results suggest that TRP5 directs the formation of a highly Ca2+-permeable ion channel which can be activated through receptor-operative pathways other than depletion of Ca2+ from Ca2+ stores, and that the activity of this channel is highly dependent on Ca2+ concentrations inside and outside of cells. Second, the TRP7 function was characterized using the same recombinant expression system. The results show that TRP7 directs the formation of a Ca2+-permeable ion channel with significant basal activity. The activity of TRP7 did not depend on extracellular Ca2+ but was enhanced by receptor-stimulation even with the low concentration of ATP which could not induce Ca2+ release from the intracellular Ca2+ store. The obtained functional characters of the two novel TRP homologues together suggest that mammalian TRP homologues are members of a large family of channels responsible for receptor-activated Ca2+ influx including not only SOC but also other diverse subtypes of channels in the family. |
要旨・審査要旨 / Abstract, Screening Result (277.1 kB)
