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The Roles of Serine/Threonine Protein Phosphatases in Regulation of PDE3B Signaling in 3T3‐L1 Adipocytes
Author(s) -
Khan Faiyaz Ahmad,
ZmudaTrzebiatowska Emilia,
Degerman Eva,
Manganiello Vincent
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.891.4
Subject(s) - phosphorylation , phosphatase , serine , biochemistry , okadaic acid , chemistry , kinase , threonine
Using sucrose gradient centrifugation and confocal microscopy, PDE3B was localized in internal membranes and PM in adipocytes. PDE3B is phosphorylated/activated by insulin and CL316243 (b3‐receptor agonist) via PKB‐ and PKA‐ signaling, respectively. To study the role of phosphatases in the regulation of PDE3B, we investigated the phosphorylation/activation of PDE3B in response to calyculin A (PP2A inhibitor). Incubation of serum starved (16 h) adipocytes with calyculin A induced time and dose dependent phosphorylation/activation of PDE3B. Maximal effects were obtained after 30 min, using 1 uM okadaic acid (1.5 fold activation) or 100 nM calyculin A (2.4 fold). Superose 6 (S6) chromatography of solubilized membranes from serum‐treated adipocytes indicated that PDE3B primarily elutes in high molecular weight macromolecular complexes (HMW =3000 kD). After incubation of adipocytes in serum free media (6‐16 h), PDE3B moves from HMW to low molecular weight (LMW, ~900 kD) complexes. S6 chromatography of solubilized membranes from adipocytes (serum‐starved, 6‐16 h) incubated with calyculin demonstrated formation of distinct HMW complexes that contain 32 P‐phosphorylated PDE3B. These studies suggest phosphorylation/activation of PDE3B and its recruitment into HMW can be regulated by calyculin‐induced inactivation of phosphatases, and that serum maintained HMW complexes that contained PDE3B.