title: Activation of PDE2 and PDE5 by specific GAF ligands: delayed activation of PDE5.
abstract: OBJECTIVE
  By controlling intracellular cyclic nucleotide levels, phosphodiesterases (PDE) serve important functions within various signalling pathways. The PDE2 and PDE5 families are allosterically activated by their substrate cGMP via regulatory so-called GAF domains. Here, we set out to identify synthetic ligands for the GAF domains of PDE2 and PDE5.
  
  
  METHODS
  Using fluorophore-tagged, isolated GAF domains of PDE2 and PDE5, promising cGMP analogues were selected. Subsequently, the effects of these analogues on the enzymatic activity of PDE2 and PDE5 were analysed.
  
  
  RESULTS
  The PDE2 ligands identified, 5,6-DM-cBIMP and 5,6-DCl-cBIMP, caused pronounced, up to 40-fold increases of the cAMP- and cGMP-hydrolysing activities of PDE2. The ligand for the GAF domains of PDE5, 8-Br-cGMP, elicited a 20-fold GAF-dependent activation and moreover revealed a time-dependent increase in PDE5 activity that occurred independently of a GAF ligand. Although GAF-dependent PDE5 activation was fast at high ligand concentrations, it was slow at physiologically relevant cGMP concentrations; PDE5 reached its final catalytic rates at 1µM cGMP after approximately 10min.
  
  
  CONCLUSIONS
  We conclude that the delayed activation of PDE5 is required to shape biphasic, spike-like cGMP signals. Phosphorylation of PDE5 further enhances activity and conserves PDE5 activation, thereby enabling PDE5 to act as a molecular memory balancing cGMP responses to nitric oxide or natriuretic peptide signals.
authors.affiliation: Institut für Pharmakologie und Toxikologie, Medizinische Fakultät, Ruhr-Universität-Bochum, Bochum, Germany.
authors.forename: Ronald
authors.initials: R
authors.lastname: Jäger
authors.forename: Frank
authors.initials: F
authors.lastname: Schwede
authors.forename: Hans-Gottfried
authors.initials: HG
authors.lastname: Genieser
authors.forename: Doris
authors.initials: D
authors.lastname: Koesling
authors.forename: Michael
authors.initials: M
authors.lastname: Russwurm
mesh_terms: D000818:Animals; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology*; D054678:Cyclic Nucleotide Phosphodiesterases, Type 2 / Q000737:chemistry / Q000378:metabolism*; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5 / Q000737:chemistry / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D051379:Mice; D045265:Natriuretic Peptides / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
chemical_list: D008024:Ligands; D045265:Natriuretic Peptides; C016276:8-bromocyclic GMP; D009569:Nitric Oxide; D054678:Cyclic Nucleotide Phosphodiesterases, Type 2; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5; D006152:Cyclic GMP