Prolyl Hydroxylase Domain Protein 2 Regulates the Intracellular Cyclic AMP Level in Cardiomyocytes Through Its Interaction with Phosphodiesterase 4D

Cyclic adenosine 3',5'-monophosphate (cAMP), which is synthesized by adenylyl cyclase (AC) and degraded by phosphodiesterase (PDE), plays crucial roles in the regulation of multiple cellular functions and physiological processes. Prolyl hydroxylase domain (PHD) proteins, which belong to a family of dioxygenases that function as oxygen sensors through their hydroxylation activity, have been implicated in multiple signaling pathways. Here, we aimed to determine whether PHD played a role in regulating intracellular cAMP level in cardiomyocytes. Through the overexpression/knockdown of the PHD gene and the measurement of the cAMP content, we found that PHD2, but not PHD1 or PHD3, acts as a regulator of intracellular cAMP. In neonatal rat cardiomyocytes and H9c2 cells, the overexpression of PHD2 increased the intracellular cAMP level, whereas the PHD2 knockdown reduced it. There was no alteration in the AC expression or activity in cells that overexpressed or downregulated PHD2. The overexpression of PHD2 decreased both the protein expression and the activity of phosphodiesterase 4D (PDE4D), whereas the PHD2 knockdown increased the PDE4D expression and activity. Co-immunoprecipitation experiments revealed a direct binding between PHD2 and PDE4D and liquid chromatography-tandem mass spectrometry analyses identified the specific hydroxylation sites on PDE4D. In conclusion, PHD2 may directly interact with PDE4D to function as a novel regulator of the intracellular cAMP levels in cardiomyocytes.