Pde2 activity assay kit

The primary antibodies and their working concentrations used in this study are listed in Supplementary Material Table 1.2.3. The Mitochondria Fractionation Kit (Cat# C3601) was purchased from Beyotime (Biotechnology, China). PDE2 Activity Assay Kit (Cat# ab139460), PKA Kinase Activity Assay Kit (Cat# ab139435), cAMP Assay (Cat# ab65355) and ODQ (Cat# ab120022) were all purchased from Abcam (Abcam, United Kingdom). Go 6983 (Cas# 133053-19-7) inhibitor was from MCE (MCE, USA). The CellTiter 96^® Aqueous One Solution Cell Proliferation Assay was purchased from Promega (Cat# G3581, Promega, USA) and 5-ethynyl-2’-deoxyuridine (EdU)-incorporation assay kit was purchased from Ribobio (Cat# C10310, Ribobio, China). Bay 60-7550 (Cat# HY-14992) and H89 (Cat# HY-15979) were purchased from Med Chemexpress (Med Chemexpress, USA).

The enzymatic activities of mitochondrial PDE2 and PKA were determined with the PDE2 Activity Assay Kit and PKA Kinase Activity Assay Kit (Abcam, United Kingdom), respectively, following the manufacturers’ protocols. Briefly, cells were harvested in a 15 mL conical tube (1 x 10⁷/mL). The cells were lysed with the reagent in the kit and subsequently the protein concentration was measured. Then the enzymatic activities of mitochondrial PDE2 and PKA were assayed using the reagent kit according to the manufacturers’ protocols. All values were normalized to total cellular protein concentration.