title: Newly synthesized cAMP is integrated at a membrane protein complex signalosome to ensure receptor response specificity.
abstract: Spatiotemporal regulation of cAMP within the cell is required to achieve receptor-specific responses. The mechanism through which the cell selects a specific response to newly synthesized cAMP is not fully understood. In hepatocyte plasma membranes, we identified two functional and independent cAMP-responsive signaling protein macrocomplexes that produce, use, degrade, and regulate their own nondiffusible (sequestered) cAMP pool to achieve their specific responses. Each complex responds to the stimulation of an adenosine G protein-coupled receptor (Ado-GPCR), bound to either A2Aor A2B, but not simultaneously to both. Each isoprotein involved in each signaling cascade was identified by measuring changes in cAMP levels after receptor activation, and its participation was confirmed by antibody-mediated inactivation. A2A-Ado-GPCR selective stimulation activates adenylyl cyclase 6 (AC6), which is bound to AKAP79/150, to synthesize cAMP which is used by two other AKAP79/150-tethered proteins: protein kinase A (PKA) and phosphodiesterase 3A (PDE3A). In contrast, A2B-Ado-GPCR stimulation activates D-AKAP2-attached AC5 to generate cAMP, which is channeled to two other D-AKAP2-tethered proteins: guanine-nucleotide exchange factor 2 (Epac2) and PDE3B. In both cases, prior activation of PKA or Epac2 with selective cAMP analogs prevents de novo cAMP synthesis. In addition, we show that cAMP does not diffuse between these protein macrocomplexes or 'signalosomes'. Evidence of coimmunoprecipitation and colocalization of some proteins belonging to each signalosome is presented. Each signalosome constitutes a minimal functional signaling unit with its own machinery to synthesize and regulate a sequestered cAMP pool. Thus, each signalosome is devoted to ensure the transmission of a unique and unequivocal message through the cell.
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Raquel
authors.initials: R
authors.lastname: Guinzberg
authors.affiliation: Departamento de Nutrición Animal y Bioquímica, Facultad de Medicina Veterinaria y Zootécnia, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Antonio
authors.initials: A
authors.lastname: Díaz-Cruz
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Carlos
authors.initials: C
authors.lastname: Acosta-Trujillo
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: María Magdalena
authors.initials: MM
authors.lastname: Vilchis-Landeros
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Héctor
authors.initials: H
authors.lastname: Vázquez-Meza
authors.affiliation: Instituto de Neurobiología, Universidad Nacional Autónoma de México (UNAM), Querétaro, Mexico.
authors.forename: Carlos
authors.initials: C
authors.lastname: Lozano-Flores
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Natalia
authors.initials: N
authors.lastname: Chiquete-Felix
authors.affiliation: Instituto de Neurobiología, Universidad Nacional Autónoma de México (UNAM), Querétaro, Mexico.
authors.forename: Alfredo
authors.initials: A
authors.lastname: Varela-Echavarría
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Salvador
authors.initials: S
authors.lastname: Uribe-Carvajal
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Héctor
authors.identifier: 0000-0001-7520-4823
authors.initials: H
authors.lastname: Riveros-Rosas
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Ciudad de México, Mexico.
authors.forename: Enrique
authors.initials: E
authors.lastname: Piña
mesh_terms: D054758:A Kinase Anchor Proteins / Q000235:genetics / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000096:biosynthesis*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D054684:Cyclic Nucleotide Phosphodiesterases, Type 3 / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D008297:Male; D061251:Primary Cell Culture; D051381:Rats; D017208:Rats, Wistar; D043705:Receptor, Adenosine A2A / Q000235:genetics / Q000378:metabolism*; D043704:Receptor, Adenosine A2B / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D054758:A Kinase Anchor Proteins; D020662:Guanine Nucleotide Exchange Factors; C531571:Rapgef4 protein, rat; D043705:Receptor, Adenosine A2A; D043704:Receptor, Adenosine A2B; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D054684:Cyclic Nucleotide Phosphodiesterases, Type 3; C517290:Pde3a protein, rat; C517294:Pde3b protein, rat; D000262:Adenylyl Cyclases; C121100:adenylyl cyclase 6; D002118:Calcium
keywords: 
  AKAP
  ; 
  GPCR
  ; 
  PKA
  ; Epac; adenylyl cyclase; phosphodiesterase