Premium
Insulin‐mediated redox regulation of cyclic AMP‐dependent protein kinase (PKA) in rat adipocytes
Author(s) -
RiverosRosas Héctor,
Zentella de Piña Martha,
VázquezMeza Héctor,
Pardo Juan Pablo,
Rendón Juan Luis,
VillalobosMolina Rafael,
Piña Enrique
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.lb265
Subject(s) - lipolysis , protein kinase a , chemistry , lipogenesis , protein subunit , autophosphorylation , biochemistry , enzyme activator , kinase , medicine , signal transduction , adipose tissue , gene
Lipolysis and lipogenesis are two highly regulated pathways. Catecholamines promote lipolysis via cAMP, whereas insulin stimulates lipogenesis. H 2 O 2 mimics insulin action inhibiting lipolysis, and insulin stimulates NADPH oxidase in adipocytes and transiently generates H 2 O 2 . Therefore, the possibility that H 2 O 2 might regulate cAMP‐dependent protein kinase (PKA) was explored. Results show that H 2 O 2 generated by insulin in rat adipocytes impaired cAMP‐mediated amplification cascade of lipolysis. Micromolar concentrations of H 2 O 2 added before cAMP suppressed cAMP activation of both type IIß or type IIα cyclic AMP‐dependent protein kinase (PKA) holoenzyme. However, H 2 O 2 was ineffective on reconstituted PKA holoenzyme preincubated with cAMP, or if added to the catalytic αsubunit alone. The same results were obtained if the catalytic αsubunit was substituted by its C199A mutant. H 2 O 2 inhibition of PKA activation remained after H 2 O 2 elimination but was reverted with thioredoxin. Electrophoresis of holoenzyme in SDS gels showed separation of catalytic and regulatory subunits after cAMP incubation but a single band after H 2 O 2 incubation. These data strongly suggest that H 2 O 2 promotes the formation of an intersubunit disulfide bond, impairing PKA activation. Phylogenetic analysis showed that Cys‐97 is conserved only in type II (but not type I) regulatory subunits. Supported by DGAPA‐UNAM grant IN208308