Protein kinase C signalling involved in prothoracicotropic hormone‐stimulated prothoracic glands in the silkworm, Bombyx mori

In the present study, the participation of protein kinase C (PKC) signalling in prothoracicotropic hormone (PTTH)‐stimulated ecdysteroidogenesis in Bombyx prothoracic glands (PGs) is demonstrated and characterized. PTTH stimulated phosphorylation of a 37‐kDa protein in Bombyx PGs both in vitro and in vivo, as recognized by a PKC substrate antibody. Treatment with either A23187 or thapsigargin also stimulated this 37‐kDa protein phosphorylation. PTTH‐stimulated phosphorylation of the 37‐kDa protein was markedly attenuated in the absence of Ca2+. The phospholipase C (PLC) inhibitor, U73122, greatly inhibited PTTH‐stimulated phosphorylation of this protein, indicating the involvement of Ca2+ and PLC. A mitogen‐activated protein kinase/extracellular signal‐regulated kinase (ERK) kinase (MEK) inhibitor (U0126), a phosphoinositide 3‐kinase (PI3K) inhibitor (LY294002) and a chemical activator of adenosine 5′‐monophosphate‐activated protein kinase (AMPK) (5‐aminoimidazole‐4‐carboxamide‐1‐β‐d‐ribofuranoside) did not affect PTTH‐stimulated phosphorylation of the 37‐kDa protein, implying that ERK and PI3K/AMPK are not the upstream signalling pathways for PKC‐dependent protein phosphorylation. The mitochondrial oxidative phosphorylation inhibitors (the uncoupler carbonyl cyanide p‐trifluoromethoxyphenylhydrazone and diphenylene iodonium) inhibited PTTH‐stimulated phosphorylation of the 37‐kDa protein, indicating its redox regulation. Treatment with PKC inhibitors (either calphostin C, chelerythrine C or rottlerin) reduced PTTH‐stimulated phosphorylation of the 37‐kDa protein. PTTH‐stimulated ecdysteroidogenesis was also inhibited by treatment with rottlerin, thus further confirming participation of PKC‐dependent phosphorylation in PTTH signalling. From these results, we demonstrated that redox‐regulated PTTH‐stimulated PKC signalling is involved in ecdysteroid secretion in Bombyx PGs. The participation of PKC in Bombyx PTTH signaling was demonstrated. PTTH stimulated PKC‐dependent phosphorylation of a 37‐kDa protein. Inhibition of PKC signaling partly decreased PTTH‐stimulated ecdysone secretion.