9507000 Huang The long term goal of this project is to understand the signal transduction mechanisms of the muscarinc acetylcholine receptors (mAchRs) and how they modulate ion channel activities. This investigator has demonstrated that regulation of the activity of a delayed rectifier potassium channel (RAK) by the mAchR involves activation of a tyrosine kinase by the G protein coupled to the receptor. Tyrosine phosphorylation of the receptor channel protein results in its suppression. Two hypotheses will be tested in this proposal: whether the (( subunits of the G protein directly activate the pleckstrin-homology domain containing tyrosine kinases and whether the channel phosphorylation state is correlated with the channel subconductance state. This investigator has shown that the mAchR can activate multiple families of tyrosine kinases. One family, Btk/Tsk, is a new class of tyrosine kinases which contains the pleckstrin-homology (PH) domain, which has been suggested to interact with the (( subunits of heterotrimeric G proteins. In this study the PI will see if the these subunits can activate the kinase in vitro and after cotransfection into cells. The region of the BtK/Tsk that binds to the G protein subunits will be identified. Studies will be done to determine which tyrosine kinases that act on the RAK channel to decrease its activity, and the effects of tyrosine phosphorylation on channel function will be determined. %%% These studies will provide information on how tyrosine kinases couple to the muscarinic acetylcholine receptors and how the resultant tyrosine phosphorylation of the potassium channel affects it s activity. This is an important signal transduction mechanism which has not been elucidated. ***
