Similarly, Pdfr5304 mutant flies showed a reduction in the absolute levels of sex pheromone expression relative to w1118 controls ( Figure 4B).

Conclusions, however, could not be drawn regarding the absence of temporal changes in the level of expression in Pdfr5304, since w1118 failed to show the predicted subjective day/night differences when sampled at circadian time (CT) 4–6 and 16–18. The decrease in sex pheromone levels in Pdf01 and Pdfr5304 were part of an overall reduction in the total amount of CHCs, encompassing all chemical classes of hydrocarbon compounds ( Table S6). Thus, Pdf and Pdfr are necessary for regulating the expression of sex pheromones and CHCs in general. Next, we asked whether rescuing Pdf would restore sex pheromone expression. We found that the Pdfresc transgene rescued the expression of 5-T this website and 7-P of Pdf01 mutant males to near

wild-type levels, but it failed to recover 7-T expression or subjective day/night differences ( Figure 4A and Table S6). The rescue of sex pheromone expression, although www.selleckchem.com/products/icotinib.html only partial, further demonstrates the requirement for PDF in regulating oenocyte physiology. The gene encoding the PDF receptor, Pdfr, is expressed by the oenocytes ( Figure S3), an indication that the oenocytes are primed to respond to direct stimulation by PDF. To determine whether the oenocytes can respond directly to the PDF peptide, we used the Gal4/UAS system to specifically target the expression of a cell membrane-tethered form of PDF (tPDF; Choi et al., 2009) to the oenocytes. The oenocyte Gal4 driver oe-Gal4 ( Billeter et al., 2009) was used to drive tPDF expression. Flies expressing tPDF in the oenocytes were compared to those expressing a scrambled PDF peptide (tPDF-scr), a peptide containing the disarranged component amino acids present in PDF. Flies heterozygous for Sitaxentan oe-Gal4, UAS-tPDF, and UAS-tPDF-scr transgenes served as negative controls. The misexpression of tPDF in the oenocytes resulted in a significant increase in the levels of 7-T and 7-P relative to tPDF-scr

and heterozygous controls at all time points sampled on DD6 (Figure 5A). Opposite to the decreased expression found with the Pdf01 and Pdfr5304 mutations, tPDF induced an overall increase in the total amount of CHCs, affecting all chemical classes of cuticular compounds analyzed, except CVA ( Table S7). Thus, the oenocytes have the ability to directly respond to the PDF peptide. Moreover, the relationship between the gain-of-function expression of tPDF and the loss-of-function Pdf01 and Pdfr5304 mutants with regard to sex pheromone expression suggests that a common regulatory pathway may be alternately activated and repressed by these genetic manipulations. The PDF ligand is thought to signal exclusively through PDFR to affect the circadian rhythm in locomotor activity.