cAMP sensor Epac as a determinant of ATP‐sensitive potassium channel activity in human pancreatic β cells and rat INS‐1 cells

Abstract

The Epac family of cAMP‐regulated guanine nucleotide exchange factors (cAMPGEFs, also known as Epac1 and Epac2) mediate stimulatory actions of the second messenger cAMP on insulin secretion from pancreatic β cells. Because Epac2 is reported to interact in vitro with the isolated nucleotide‐binding fold‐1 (NBF‐1) of the β‐cell sulphonylurea receptor‐1 (SUR1), we hypothesized that cAMP might act via Epac1 and/or Epac2 to inhibit β‐cell ATP‐sensitive K + channels (K ATP channels; a hetero‐octomer of SUR1 and Kir6.2). If so, Epac‐mediated inhibition of K ATP channels might explain prior reports that cAMP‐elevating agents promote β‐cell depolarization, Ca 2 + influx and insulin secretion. Here we report that Epac‐selective cAMP analogues (2′‐ O ‐Me‐cAMP; 8‐pCPT‐2′‐ O ‐Me‐cAMP; 8‐pMeOPT‐2′‐ O ‐Me‐cAMP), but not a cGMP analogue (2′‐ O ‐Me‐cGMP), inhibit the function of K ATP channels in human β cells and rat INS‐1 insulin‐secreting cells. Inhibition of K ATP channels is also observed when cAMP, itself, is administered intracellularly, whereas no such effect is observed upon administration N 6 ‐Bnz‐cAMP, a cAMP analogue that activates protein kinase A (PKA) but not Epac. The inhibitory actions of Epac‐selective cAMP analogues at K ATP channels are mimicked by a cAMP agonist (8‐Bromoadenosine‐3′, 5′‐cyclic monophosphorothioate, Sp‐isomer, Sp‐8‐Br‐cAMPS), but not a cAMP antagonist (8‐Bromoadenosine‐3′, 5′‐cyclic monophosphorothioate, Rp‐isomer, Rp‐8‐Br‐cAMPS), and are abrogated following transfection of INS‐1 cells with a dominant‐negative Epac1 that fails to bind cAMP. Because both Epac1 and Epac2 coimmunoprecipitate with full‐length SUR1 in HEK cell lysates, such findings delineate a novel mechanism of second messenger signal transduction in which cAMP acts via Epac to modulate ion channel function, an effect measurable as the inhibition of K ATP channel activity in pancreatic β cells.