Sulfonylurea and K+-channel opener sensitivity of K-ATP channels - Functional coupling of Kir6.2 and SUR1 subunits

The sensitivity of K-ATP channels to high-affinity block by sulfonyiureas a nd to stimulation by K+ channel openers and MgADP (PCOs) is conferred by th e regulatory sulfonylurea receptor (SUR) subunit, whereas ATP inhibits the channel through interaction with the inward rectifier (Kir6.2) subunit. Pho sphatidylinositol 4,5-bisphosphate (PIP2) profoundly antagonized ATP inhibi tion of K-ATP channels expressed from cloned Kir6.2+SUR1 subunits, but also abolished high affinity tolbutamide sensitivity. Br;stabilizing the open s tate of the channel, PIP2 drives the channel away from closed state(s) that are preferentially affected by high affinity tolbutamide binding, thereby producing an apparent loss of high affinity tolbutamide inhibition. Mutant K-ATP channels (Kir6.2[Delta N30] or Kir6.2[L164A], coexpressed with SUR1) also displayed an "uncoupled" phenotype with no high affinity tolbutamide b lock and with intrinsically higher open state stability. Conversely, Kir6.2 [R176A]+SUR1 channels, which have an intrinsically lower open state stabili ty, displayed a greater high affinity fraction of tolbutamide block. In add ition to antagonizing high-affinity block by tolbutamide, PIP2 also altered the stimulatory action of the PCOs, diazoxide and MgADP. With time after P IP, application, PCO stimulation first increased, and then subsequently dec reased, probably reflecting a common path-way for activation of the channel by stimulatory PCOs and PIP2. The net effect of increasing open state stab ility, either by PIP2 or mutagenesis, is an apparent "uncoupling" of the Ki r6.2 subunit from the regulatory input of SUR1, an action that can be parti ally reversed by screening negative charges on the membrane with poly-L-lys ine.