title: NorpA and itpr mutants reveal roles for phospholipase C and inositol (1,4,5)- trisphosphate receptor in Drosophila melanogaster renal function.
abstract: Mutants of norpA, encoding phospholipase C beta (PLC beta), and itpr, encoding inositol (1,4,5)-trisphosphate receptor (IP(3)R), both attenuate response to diuretic peptides of Drosophila melanogaster renal (Malpighian) tubules. Intact tubules from norpA mutants severely reduced diuresis stimulated by the principal cell- and stellate cell-specific neuropeptides, CAP(2b) and Drosophila leucokinin (Drosokinin), respectively, suggesting a role for PLC beta in both these cell types. Measurement of IP(3) production in wild-type tubules and in Drosokinin-receptor-transfected S2 cells stimulated with CAP(2b) and Drosokinin, respectively, confirmed that both neuropeptides elevate IP(3) levels. In itpr hypomorphs, basal IP(3) levels are lower, although CAP(2b)-stimulated IP(3) levels are not significantly reduced compared with wild type. However, CAP(2b)-stimulated fluid transport is significantly reduced in itpr alleles. Rescue of the itpr(90B.0) allele with wild-type itpr restores CAP(2b)-stimulated fluid transport levels to wild type. Drosokinin-stimulated fluid transport is also reduced in homozygous and heteroallelic itpr mutants. Measurements of cytosolic calcium levels in intact tubules of wild-type and itpr mutants using targeted expression of the calcium reporter, aequorin, show that mutations in itpr attenuated both CAP(2b)- and Drosokinin-stimulated calcium responses. The reductions in calcium signals are associated with corresponding reductions in fluid transport rates. Thus, we describe a role for norpA and itpr in renal epithelia and show that both CAP(2b) and Drosokinin are PLC beta-dependent, IP(3)-mobilising neuropeptides in Drosophila. IP(3)R contributes to the calcium signalling cascades initiated by these peptides in both principal and stellate cells.
authors.affiliation: Institute of Biomedical and Life Sciences, Division of Molecular Genetics, University of Glasgow, Glasgow G11 6NU, UK.
authors.forename: Valerie P
authors.initials: VP
authors.lastname: Pollock
authors.forename: Jonathan C
authors.initials: JC
authors.lastname: Radford
authors.forename: Susan
authors.initials: S
authors.lastname: Pyne
authors.forename: Gaiti
authors.initials: G
authors.lastname: Hasan
authors.forename: Julian A T
authors.initials: JA
authors.lastname: Dow
authors.forename: Shireen-A
authors.initials: SA
authors.lastname: Davies
mesh_terms: D000331:Aequorin; D000483:Alleles; D000818:Animals; D015220:Calcium Channels / Q000235:genetics / Q000502:physiology*; D020013:Calcium Signaling / Q000502:physiology; D017931:DNA Primers; D004231:Diuresis / Q000502:physiology; D029721:Drosophila Proteins / Q000378:metabolism; D004331:Drosophila melanogaster / Q000502:physiology*; D020869:Gene Expression Profiling; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism / Q000502:physiology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007527:Isoenzymes / Q000235:genetics / Q000502:physiology*; D008317:Malpighian Tubules / Q000378:metabolism / Q000502:physiology*; D009154:Mutation; D009479:Neuropeptides / Q000378:metabolism; D054799:Phospholipase C beta; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000502:physiology*; D010738:Type C Phospholipases / Q000235:genetics / Q000502:physiology*
chemical_list: D015220:Calcium Channels; D017931:DNA Primers; D029721:Drosophila Proteins; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007527:Isoenzymes; C468143:Lk protein, Drosophila; D009479:Neuropeptides; D018160:Receptors, Cytoplasmic and Nuclear; D000331:Aequorin; D015544:Inositol 1,4,5-Trisphosphate; D010738:Type C Phospholipases; D054799:Phospholipase C beta