Essential role of phosphatidylinositol 3‐kinase in insulin‐induced activation and phosphorylation of the cGMP‐inhibited cAMP phosphodiesterase in rat adipocytes studies using the selective inhibitor wortmannin

Abstract

Incubation of rat adipocytes with wortmannin, a potent and selective phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor, completely blocked the antilipolytic action of insulin (ICs0= 100 nM), the insulin-induced activation and phosphorylation of cGMP-inhibited cAMP phosphodiesterase (cGI-PDE) as well as the activation of the insulin-stimulated cGI-PDE kinase (ICs0= 10-30 nM). No direct effects of the inhibitor on the insulinstimulated cGI-PDE kinase, the cGI-PDE and the hormone-sensitive lipase were observed. These data suggest that activation of PI 3-kinase upstream of the insulin-stimulated cGI-PDE kinase in the antilipolytic insulin signalchain has an essential role for insulin-induced cGI-PDE activation/ phosphorylation and anti-lipolysis.

Phosphorylation and activation of phosphodiesterase type 3B (PDE3B) in adipocytes in response to serine/threonine phosphatase inhibitors: deactivation of PDE3B in vitro by protein phosphatase type 2A

“…In addition, PP2A could also act on upstream signal-transduction components involved in the regulation of PDE3B. The presence of an insulin-stimulated kinase that phosphorylates PDE3B in itro has been reported [21], which recently has been suggested to be protein kinase B [22]. Degerman and V. Manganiello, unpublished work) and PDE3A [51].…”

“…Hormone-induced regulation of PDE3B has been studied extensively in primary rat adipocytes where phosphorylation of serine 302 [6][7][8][9] and activation of PDE3B is thought to be the major mechanism whereby insulin antagonizes protein kinase Amediated phosphorylation and activation of hormone-sensitive lipase and lipolysis [16,17,19,20]. Although phosphatidylinositol 3-kinase is most probably involved as an upstream regulator of PDE3B in the insulin-signalling pathway [21], the kinase that phosphorylates and activates PDE3B directly has not been identified. Recent findings imply that it could be protein kinase B [22].…”

Mechanisms of inhibition of lipolysis by insulin, vanadate and peroxovanadate in rat adipocytes

“…Mouse polyclonal anti-phosphotyrosine antibodies (PY20) were purchased from Transduction Laboratories. Polyclonal peptide anti-PDE3B antibodies were raised in rabbits and used as described previously [14]. Glycerol kinase and glycerol-3-phosphate dehydrogenase were from Boehringer Mannheim.…”

“…PDE3B was immunoisolated from solubilized membranes and subjected to SDS\PAGE as described previously [14]. IRS-1 immunoprecipitates were separated by SDS\PAGE and electrotransferred to PVDF membranes as described above.…”

“…Hormone-sensitive lipase, the rate-limiting lipolytic enzyme, is thereby dephosphorylated and inactivated [12]. Phosphoinositide 3-kinase (PI-3K), which associates with insulin receptor substrates (IRSs) and becomes activated during insulin stimulation (reviewed in [13]), has also been shown to be important in insulin's anti-lipolytic signalling pathway [14,15]. This study reveals the complexity and the diversity in the mechanisms of inhibition of lipolysis by vanadate and pV.…”