β 2 -adrenoceptor-induced modulation of transglutaminase 2 transamidase activity in cardiomyoblasts

• Published in: European journal of pharmacology 2017-10, Vol.813, p.105-121
• Main Authors: Vyas, Falguni S, Nelson, Carl P, Freeman, Fiona, Boocock, David J, Hargreaves, Alan J, Dickenson, John M
• Format: Article
• Language: English
• Subjects: Animals; Calcium - metabolism; Catecholamines - pharmacology; Cell Line; Cyclic AMP-Dependent Protein Kinases - metabolism; Enzyme Activation - drug effects; Enzyme Inhibitors - pharmacology; GTP-Binding Proteins - antagonists & inhibitors; GTP-Binding Proteins - metabolism; Myocytes, Cardiac - cytology; Myocytes, Cardiac - drug effects; Myocytes, Cardiac - enzymology; Myocytes, Cardiac - metabolism; Phosphorylation - drug effects; Rats; Receptors, Adrenergic, beta-2 - metabolism; Signal Transduction - drug effects; Transglutaminases - antagonists & inhibitors; Transglutaminases - metabolism
• ISSN: 0014-2999; 1879-0712
• DOI: 10.1016/j.ejphar.2017.07.043

Tissue transglutaminase 2 (TG2) is modulated by protein kinase A (PKA) mediated phosphorylation: however, the precise mechanism(s) of its modulation by G-protein coupled receptors coupled to PKA activation are not fully understood. In the current study we investigated the potential regulation of TG2 activity by the β -adrenoceptor in rat H9c2 cardiomyoblasts. Transglutaminase transamidation activity was assessed using amine-incorporating and protein cross-linking assays. TG2 phosphorylation was determined via immunoprecipitation and Western blotting. The long acting β -adrenoceptor agonist formoterol induced time- and concentration-dependent increases in TG2 transamidation. Increases in TG2 activity were reduced by the TG2 inhibitors Z-DON (Benzyloxycarbonyl-(6-Diazo-5-oxonorleucinyl)-L-valinyl-L-prolinyl-L-leucinmethylester) and R283 ((1,3,dimethyl-2[2-oxo-propyl]thio)imidazole chloride). Responses to formoterol were blocked by pharmacological inhibition of PKA, extracellular signal-regulated kinase 1 and 2 (ERK1/2), or phosphatidylinositol 3-kinase (PI-3K) signalling. Furthermore, the removal of extracellular Ca also attenuated formoterol-induced TG2 activation. Fluorescence microscopy demonstrated TG2-induced biotin-X-cadaverine incorporation into proteins. Formoterol increased the levels of TG2-associated phosphoserine and phosphothreonine, which were blocked by inhibition of PKA, ERK1/2 or PI-3K signalling. Subsequent proteomic analysis identified known (e.g. lactate dehydrogenase A chain) and novel (e.g. Protein S100-A6) protein substrates for TG2. Taken together, the data obtained suggest that β -adrenoceptor-induced modulation of TG2 represents a novel paradigm in β -adrenoceptor cell signalling, expanding the repertoire of cellular functions responsive to catecholamine stimulation.