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Originally published as Genetics Published Articles Ahead of Print on February 3, 2005.
Genetics, Vol. 169, 1541-1552, March 2005, Copyright © 2005
doi:10.1534/genetics.104.035139
Transient Receptor Potential-Like Channels Are Essential for Calcium Signaling and Fluid Transport in a Drosophila Epithelium
Matthew R. MacPherson*,1,
Valerie P. Pollock*,1,
Laura Kean*,
Tony D. Southall*,
Maria E. Giannakou*,
Kate E. Broderick*,
Julian A. T. Dow*,
Roger C. Hardie
and
Shireen A. Davies*,2
* IBLS Division of Molecular Genetics, University of Glasgow, Glasgow G11 6NU, United Kingdom
Department of Anatomy, University of Cambridge, Cambridge CB2 3DY, United Kingdom
2 Corresponding author: Division of Molecular Genetics, Institute of Biomedical and Life Sciences, Anderson Complex, University of Glasgow, 56 Dumbarton Rd., Glasgow G11 6NU, United Kingdom.
E-mail: s.a.davies{at}bio.gla.ac.uk
Calcium signaling is an important mediator of neuropeptide-stimulated fluid transport by Drosophila Malpighian (renal) tubules. We demonstrate the first epithelial role, in vivo, for members of the TRP family of calcium channels. RT-PCR revealed expression of trp, trpl, and trp
in tubules. Use of antipeptide polyclonal antibodies for TRP, TRPL, and TRP showed expression of all three channels in type 1 (principal) cells in the tubule main segment. Neuropeptide (CAP2b)-stimulated fluid transport rates were significantly reduced in tubules from the trpl302 mutant and the trpl;trp double mutant, trpl302;trp343. However, a trp null, trp343, had no impact on stimulated fluid transport. Measurement of cytosolic calcium concentrations ([Ca2+]i) in tubule principal cells using an aequorin transgene in trp and trpl mutants showed a reduction in calcium responses in trpl302. Western blotting of tubule preparations from trp and trpl mutants revealed a correlation between TRPL levels and CAP2b-stimulated fluid transport and calcium signaling. Rescue of trpl302 with a trpl transgene under heat-shock control resulted in a stimulated fluid transport phenotype that was indistinguishable from wild-type tubules. Furthermore, restoration of normal stimulated rates of fluid transport by rescue of trpl302 was not compromised by introduction of the trp null, trp343. Thus, in an epithelial context, TRPL is sufficient for wild-type responses. Finally, a scaffolding component of the TRPL/TRP-signaling complex, INAD, is not expressed in tubules, suggesting that inaD is not essential for TRPL/TRP function in Drosophila tubules.
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