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First published online January 27, 2004
Journal of Experimental Biology 207, 743-748 (2004)
Published by The Company of Biologists 2004
doi: 10.1242/jeb.00818

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Drosophila CG8422 encodes a functional diuretic hormone receptor

Erik C. Johnson¹, Laura M. Bohn^2,* and Paul H. Taghert^1,

¹ Department of Anatomy and Neurobiology, Washington University School of Medicine, Saint Louis, MO 63110, USA
² Department of Cell Biology, Duke University, Durham, NC 27710, USA

View larger version (73K): [in a new window]   Fig. 1. HEK-293 cells expressing CG8422 display ßarr2-GFP translocation in response to diuretic hormone 44 (DH) neuropeptide. A 10-min time series imaging a single CG8422-expressing cell before and following exposure to 10-6 mol l-1 DH (final concentration). Numbers at the bottom right of each panel refer to minutes after DH exposure. Prior to peptide application, most of the fluorescence is uniformly distributed in the cytoplasm. Within 1 min, and at times thereafter, the fluorescence translocates to the cell membrane. Similar results were obtained from three independent transfections: the majority of GFP-positive cells displayed clear ß-arrestin translocation within minutes of exposure to DH.

View larger version (72K): [in a new window]   Fig. 2. CG8422-expressing cells internalize ßarr2-GFP fluorescence after diuretic hormone 44 (DH)-induced translocation. (A) A typical cell prior to peptide application; note the uniform cytoplasmic distribution of fluorescence. (B) A different cell after peptide exposure; note the prominent disposition of fluorescence in large vesicles throughout the cell. Numbers at the bottom right of each panel refer to minutes after DH exposure.

View larger version (13K): [in a new window]   Fig. 3. Diuretic hormone 44 (DH) activation of CG8422-expressing HEK-293 cells produces a dose-dependent increase in CRE-luciferase activity. Dose-response curve for DH activation of CRE-luc gene expression in cells transiently co-expressing CG8422. The calculated EC50 is 1.47 nmol l-1. Values are means ± S.E.M. and represent the results pooled from three experiments that were performed in triplicate. Values observed with exposures to DH above 5x10-10 mol l-1 were statistically different from values observed with exposure to vehicle alone.

View larger version (19K): [in a new window]   Fig. 4. CG8422 transcripts are consistently detected in adult head RNA. Data mined from microarray results that were described by Lin et al. (2002) using adult head RNA from the genotypes and environmental conditions listed in the text. L:D, 12 h:12 h light:dark; D:D, constant darkness. Values are medians + median average deviation (M.A.D.). Values within bars represent the number of microarrays. CG8422 levels were not significantly different in WT (wild type) L:D versus WT D:D (P=0.28), in per (period mutant) L:D versus per D:D (P=0.25), in WT L:D versus per L:D (P=0.33) or in WT D:D versus per D:D (P=0.06), as indicated by the Mann-Whitney U test.

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