Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
SHR Neuro Krebs Kardio Lipid Stoffw Microb
Derler, I; Hofbauer, M; Kahr, H; Fritsch, R; Muik, M; Kepplinger, K; Hack, ME; Moritz, S; Schindl, R; Groschner, K; Romanin, C.
Dynamic but not constitutive association of calmodulin with rat TRPV6 channels enables fine tuning of Ca2+-dependent inactivation.
J Physiol. 2006; 577(Pt 1): 31-44.
Doi: 10.1113/jphysiol.2006.118661
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- Co-Autor*innen der Med Uni Graz
- Groschner Klaus
- Schindl Rainer
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- Abstract:
- The Ca(2+)-selective TRPV6 as well as the L-type Ca(2+) channel are regulated by the Ca(2+)-binding protein calmodulin (CaM). Here, we investigated the interaction of CaM with rat (r)TRPV6 in response to alterations of intracellular Ca(2+), employing Ca(2+)-imaging and patch-clamp techniques. Additionally, confocal Förster resonance energy transfer (FRET) microscopy on living cells was utilized as a key method to visualize in vivo protein-protein interactions essential for CaM regulation of rTRPV6 activity. The effects of overexpressed CaM or its Ca(2+)-insensitive mutant (CaM(MUT)) was probed on various rTRPV6 mutants and fragments in an attempt to elucidate the molecular mechanism of Ca(2+)/CaM-dependent regulation and to pinpoint the physiologically relevant rTRPV6-CaM interaction site. A significant reduction of rTRPV6 activity, as well as an increase in current inactivation, were observed when CaM was overexpressed in addition to endogenous CaM. The Ca(2+)-insensitive CaM(MUT), however, failed to affect rTRPV6-derived currents. Accordingly, live cell confocal FRET microscopy revealed a robust interaction for CaM but not CaM(MUT) with rTRPV6, suggesting a strict Ca(2+) dependence for their association. Indeed, interaction of rTRPV6 or its C terminus with CaM increased with rising intracellular Ca(2+) levels, as observed by dynamic FRET measurements. An rTRPV6Delta(695-727) mutant with the very C-terminal end deleted, yielded Ca(2+) currents with a markedly reduced inactivation in accordance with a lack of CaM interaction as substantiated by FRET microscopy. These results, in contrast with those for CaM-dependent L-type Ca(2+) channel inactivation, demonstrate a dynamic association of CaM with the very C-terminal end of rTRPV6 (aa 695-727), and this enables acceleration of the rate of rTRPV6 current inactivation with increasing intracellular CaM concentrations.
- Find related publications in this database (using NLM MeSH Indexing)
- Animals -
- Calcium - metabolism
- Calcium Signaling - physiology
- Calmodulin - metabolism
- Cell Line -
- Humans -
- Ion Channel Gating - physiology
- Kidney - physiology
- Kinetics -
- Membrane Potentials - physiology
- Rats -
- TRPV Cation Channels - physiology