Ectopic expression of a Drosophila InsP(3)R channel mutant has dominant-negative effects in vivo.
The inositol 1,4,5-trisphosphate (InsP(3)) receptor is a tetrameric intracellular calcium channel. It is an integral component of the InsP(3) signaling pathway in multicellular organisms, where it regulates cellular calcium dynamics in many different contexts. In order to understand how the primary structure of the InsP(3)R affects its functional properties, the kinetics of Ca(2+)-release in vitro from single point mutants of the Drosophila InsP(3)R have been determined earlier. Among these, the Ka901 mutant in the putative selectivity-filter of the pore is of particular interest. It is non-functional in the homomeric form whereas it forms functional channels (with altered channel properties) when co-expressed with wild-type channels. Here we show that due to its changed functional properties the Ka901 mutant protein has dominant-negative effects in vivo. Cells expressing Ka901:WT channels exhibit much higher levels of cytosolic Ca(2+) upon stimulation as compared with cells over-expressing just the wild-type DmInsP(3)R, thus supporting our in vitro observations that increased Ca(2+) release is a property of heteromeric Ka901:WT channels. Furthermore, ectopic expression of the Ka901 mutant channel in aminergic cells of Drosophila alters electrophysiological properties of a flight circuit and results in defective flight behavior.
Srikanth
Sonal
S
National Centre for Biological Sciences, Tata Institute of Fundamental Research, Gandhi Krishi Vigyan Kendra Campus, Bangalore 560 065, India.
Banerjee
Santanu
S
Hasan
Gaiti
G
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D015220:Calcium Channels / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D018076:DNA, Complementary / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D005426:Flight, Animal / Q000502:physiology; D015870:Gene Expression; D017344:Genes, Insect; D053496:Inositol 1,4,5-Trisphosphate Receptors; D009474:Neurons / Q000378:metabolism; D017354:Point Mutation; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D015220:Calcium Channels; D018076:DNA, Complementary; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D011976:Receptors, Muscarinic; D011993:Recombinant Fusion Proteins; D002217:Carbachol