Rescue of light responses in the Drosophila "null" phospholipase C mutant, norpAP24, by the diacylglycerol kinase mutant, rdgA, and by metabolic inhibition.
Light responses in Drosophila are reportedly abolished in severe mutants of the phospholipase C (PLC) gene, norpA. However, on establishing the whole-cell recording configuration in photoreceptors of the supposedly null allele, norpAP24, we detected a small ( approximately 15 pA) inward current that represented spontaneous light channel activity. The current decayed during approximately 20 min, after which tiny residual responses (<2 pA) were elicited by intense flashes. Both spontaneous currents and light responses appeared to be mediated by residual PLC activity, because they were enhanced by impairing diacylglycerol (DAG) kinase function by mutation (rdgA) or by restricting ATP but were reduced or abolished by a mutation of the PLC-specific Gq alpha subunit. It was reported recently that metabolic inhibition activated the light-sensitive transient receptor potential and transient receptor potential-like channels, even in norpAP24, leading to the conclusion that this action was independent of PLC (Agam, K., von Campenhausen, M., Levy, S., Ben-Ami, H. C., Cook, B., Kirschfeld, K., and Minke, B. (2000) J. Neurosci. 20, 5748-5755). However, we found that channel activation by metabolic inhibitors in norpAP24 was strictly dependent on the residual PLC activity underlying the spontaneous current, because the inhibitors failed to activate any channels after the spontaneous current had decayed. By contrast, polyunsaturated fatty acids invariably activated the channels independently of PLC. The results strongly support the obligatory requirement for PLC and DAG in Drosophila phototransduction, suggest that activation by metabolic inhibition is primarily because of the failure of diacylglycerol kinase, and are consistent with the proposal that polyunsaturated fatty acids, which are potential DAG metabolites, act directly on the channels.
Hardie
Roger C
RC
Cambridge University Department of Anatomy, Cambridge CB2 3DY, United Kingdom. rch14@hermes.cam.ac.uk
Martin
Fernando
F
Chyb
Sylwester
S
Raghu
Padinjat
P
D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000008:administration & dosage; D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D019852:Diacylglycerol Kinase / Q000235:genetics* / Q000378:metabolism; D004075:Diglycerides / Q000378:metabolism; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000201:enzymology* / Q000235:genetics / Q000528:radiation effects; D004553:Electric Conductivity; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D019204:GTP-Binding Proteins / Q000502:physiology; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000502:physiology; D019476:Insect Proteins / Q000187:drug effects / Q000502:physiology; D008027:Light; D009154:Mutation; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054799:Phospholipase C beta; D010786:Photoreceptor Cells / Q000502:physiology; D050051:Transient Receptor Potential Channels; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D014785:Vision, Ocular / Q000502:physiology
D015220:Calcium Channels; D004075:Diglycerides; D029721:Drosophila Proteins; D004791:Enzyme Inhibitors; D005231:Fatty Acids, Unsaturated; D019476:Insect Proteins; D050051:Transient Receptor Potential Channels; C063169:trp protein, Drosophila; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D000255:Adenosine Triphosphate; D019852:Diacylglycerol Kinase; D010738:Type C Phospholipases; C468371:NorpA protein, Drosophila; D054799:Phospholipase C beta; D019204:GTP-Binding Proteins; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D019297:2,4-Dinitrophenol