Evidence that insulin and isoprenaline activate the cGMP-inhibited low-Km cAMP phosphodiesterase in rat fat cells by phosphorylation

Journal Article · Mon Jan 01 00:00:00 EST 1990 · Proceedings of the National Academy of Sciences of the United States of America; (USA)

DOI:https://doi.org/10.1073/pnas.87.2.533· OSTI ID:6995188

Degerman, E; Smith, C J; Tornqvist, H; Vasta, V; Belfrage, P; Manganiello, V C ^[1]

---

1. Univ. of Lund (Sweden)

Incubation of intact rat fat cells with maximally effective concentrations of insulin (1 nM, 12 min) or isoprenaline (300 nM, 3 min) increased particulate cGMP- and cilostamide-inhibited, low-Km cAMP phosphodiesterase (cAMP-PDE) activity by about 50% and 100%, respectively. In 32P-labeled cells, these agents induced serine 32P-phosphorylation of a 135-kDa particulate protein and, to a variable and lesser extent, a 44-kDa protein, which were selectively immunoprecipitated by anti-cAMP-PDE, as analyzed by SDS/PAGE and autoradiography. In the absence of hormonal stimulation, little phosphorylation was detected (less than 10% of that with the hormones). The two phosphoproteins were identified as cAMP-PDE or a closely related molecule (in the case of the 44-kDa species, perhaps a proteolytic fragment) since (i) amounts of 32P in the immunoprecipitated 135-kDa protein paralleled enzyme inactivation, (ii) prior incubation of the anti-cAMP-PDE with the pure rat or bovine enzyme selectively blocked the immunoprecipitation of the phosphoproteins, (iii) 135- and 44-kDa proteins reacted with the anti-cAMP-PDE on Western immunoblots, and (iv) the two phosphoproteins copurified with cAMP-PDE activity through DEAE-Sephacel chromatography and were isolated by highly selective affinity chromatography on cilostamide-agarose. Thus, in fat cells, catecholamine- and insulin-induced activation of the cAMP-PDE may be mediated via phosphorylation by cAMP-dependent protein kinase and an insulin-activated serine protein kinase, respectively.

• OSTI ID: 6995188
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America; (USA), Vol. 87:2; ISSN 0027-8424
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
INSULIN
BIOCHEMICAL REACTION KINETICS
PHOSPHODIESTERASES
ENZYME ACTIVITY
ADIPOSE TISSUE
AMP
AUTORADIOGRAPHY
ELECTROPHORESIS
FAT CELLS
PHOSPHATES
PHOSPHORUS 32
PHOSPHORYLATION
RATS
ANIMAL CELLS
ANIMAL TISSUES
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BODY
CHEMICAL REACTIONS
CONNECTIVE TISSUE
CONNECTIVE TISSUE CELLS
DAYS LIVING RADIOISOTOPES
ENZYMES
ESTERASES
HORMONES
HYDROLASES
ISOTOPES
KINETICS
LIGHT NUCLEI
MAMMALS
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
OXYGEN COMPOUNDS
PEPTIDE HORMONES
PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
RADIOISOTOPES
REACTION KINETICS
RODENTS
SOMATIC CELLS
TISSUES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques