Abstract
Protein kinase A (PKA) is a cyclic AMP (cAMP)-dependent protein kinase composed of catalytic and regulatory subunits and involved in various physiological phenomena, including lipid metabolism. Here we demonstrated that the stoichiometric balance between catalytic and regulatory subunits is crucial for maintaining basal PKA activity and lipid homeostasis. To uncover the potential roles of each PKA subunit, Caenorhabditis elegans was used to investigate the effects of PKA subunit deficiency. In worms, suppression of PKA via RNAi resulted in severe phenotypes, including shortened life span, decreased egg laying, reduced locomotion, and altered lipid distribution. Similarly, in mammalian adipocytes, suppression of PKA regulatory subunits RIα and RIIβ via siRNAs potently stimulated PKA activity, leading to potentiated lipolysis without increasing cAMP levels. Nevertheless, insulin exerted anti-lipolytic effects and restored lipid droplet integrity by antagonizing PKA action. Together, these data implicate the importance of subunit stoichiometry as another regulatory mechanism of PKA activity and lipid metabolism.
Highlights
Protein kinase A (PKA) is a cyclic AMP-dependent serine/threonine kinase that mediates various cellular responses, including lipolysis
Because the opening of the active site is critical for PKA catalytic subunits to phosphorylate target substrates, it is likely that the decrease of regulatory subunits, which bind to the active site cleft, would lead to increased kinase activity
Assuming there is no change in cyclic AMP (cAMP) concentration and the dissociation constant between catalytic and regulatory subunits is constant (0.1 nM), our analysis showed that the amounts of free and active catalytic subunits would steeply increase upon reduction of regulatory subunits (Fig. 1, B and C)
Summary
Hormone-sensitive lipase; ATGL, adipose triglyceride lipase; qRT-PCR, quantitative real time-PCR; hADSC, human adipose-derived stem cell. Despite the abovementioned studies, deciphering fine-tuned regulatory mechanisms of PKA regulatory and catalytic subunits is limited in mouse models probably due to the compensatory effects of multiple subunit isoforms. Combinatorial small interfering RNA (siRNA) transfection revealed that among the four regulatory subunits of PKA, RI␣ and RII were required for the inhibition of basal lipolysis. These data suggest that the balance between catalytic and regulatory PKA subunits is important for blocking unnecessary PKA activation and avoiding futile lipolysis in lipid-storing tissues, independent of cAMP signaling
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