S6 phosphorylation results from prothoracicotropic hormone stimulation of insect prothoracic glands: a role for S6 kinase.
The insect prothoracic glands are the source of steroidal molting hormone precursors and the glands are stimulated by a brain neuropeptide, prothoracicotropic hormone (PTTH). Previous work from this laboratory revealed that PTTH acts via a cascade including Ca2+/calmodulin activation of adenylate cyclase, protein kinase A, and the subsequent phosphorylation of a 34 kDa protein (p34) hypothesized, but not proven, to be the S6 protein of the 40S ribosomal subunit. The immunosuppressive macrolide, rapamycin, is a potent inhibitor of cell proliferation, a signal transduction blocker, and also prevents ribosomal S6 phosphorylation in mammalian systems. We demonstrate here that rapamycin inhibited PTTH-stimulated ecdysteroidogenesis in vitro by the prothoracic glands of the tobacco hornworm, Manduca sexta, with half-maximal inhibition at a concentration of about 5 nM. At concentrations above 5 nM, there was a 75% inhibition of ecdysteroid biosynthesis. Similar results were observed with the calcium ionophore (A23187), a known stimulator of ecdysteroidogenesis. Most importantly, the inhibition of ecdysteroid biosynthesis was accompanied by the specific inhibition of the phosphorylation of p34, indicating that p34 indeed is ribosomal protein S6. In vivo assays revealed that injection of rapamycin into day 6 fifth instar larvae resulted in a decreased hemolymph ecdysteroid titer and a dose-dependent delay in molting and metamorphosis. When S6 kinase (S6K) activity was examined using rapamycin-treated prothoracic glands as the enzyme source and a synthetic peptide (S6-21) or a 40S ribosomal subunit fraction from Manduca tissues as substrate, the date revealed that rapamycin inhibited S6K activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Song
Q
Q
Department of Biology, University of North Carolina, Chapel Hill 27599-3280.
Gilbert
L I
LI
D000818:Animals; D004440:Ecdysone / Q000096:biosynthesis; D007301:Insect Hormones / Q000502:physiology*; D007814:Larva; D008675:Metamorphosis, Biological / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D009036:Moths / Q000196:embryology* / Q000235:genetics; D009479:Neuropeptides / Q000502:physiology; D010766:Phosphorylation; D011090:Polyenes / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011679:Pupa; D038601:Ribosomal Protein S6; D019893:Ribosomal Protein S6 Kinases; D012269:Ribosomal Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus; D013909:Thorax / Q000196:embryology
D007301:Insect Hormones; D009479:Neuropeptides; D011090:Polyenes; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; D004440:Ecdysone; C009089:prothoracicotropic hormone; D017346:Protein Serine-Threonine Kinases; D019893:Ribosomal Protein S6 Kinases; D020123:Sirolimus