1 8 2 G PR OTEI NS A N D RE G UL ATI O N OF A DE NYLYL CYCL ASE Nobel Lect ure, Dece mber 8, 1994 b y A LFRE D G. G IL MA N Depart ment of Phar macology, The University of Texas South western Me dical Ce nter, Dallas, Texas 75235, US A I NTRODUCTIO N E arl S ut h erl a n d, a fri e n d of m y f at h er, wr ot e t o m e i n t h e s pri n g of 1 9 6 1 wit h a pro posal to partici pate i n w hat was t he n a n e d ucatio nal a dve nt ure - a co mbine d M. D.-Ph. D. training progra m that he ha d devise d at Western Reserve U niversity ( no w Case Wester n Reserve U niversity) i n O hio. My reac- ti o n w as e ntir el y n e g ati v e. I t h a n k e d hi m, p olit el y I t hi n k, b ut t h e i d e a of s pe n di ng seve n years i n Clevela n d ha d little a p peal. Ha p pily, S ut herla n d was persiste nt. He wr ote a gai n i n t he fall of 1961 (t he be gi n ni n g of m y last year i n c oll e g e), I d e ci d e d t h e i d e a w as w ort h a visit, a n d I h a d m y first gli m ps e of cyclic A M P (for w hose discovery, i n 1957, S ut herla n d was a war de d t he Nobel Prize [in 1971]). Cyclic A M P, S utherlan d, an d the M. D.- Ph. D. Progra m all looke d rat her a p peali ng. T h us, o n my arrival i n Se pte mber, 1962, I was dis- a p pointe d to learn that S utherlan d was abo ut to de part for Van derbilt University. Ho wever, there was an attractive o p port unity to work with Theo dore Rall, S utherlan d’s yo unger collaborator, who ha d playe d a pivotal r ol e i n t h e cr u ci al e x p eri m e nts of 1 9 5 7. I e nt er e d t h e R all l a b, a n d i n o v er 3 0 s ubseq ue nt years have never esca pe d t he l ure of cyclic n ucleoti de researc h, des pite occasio nal atte m pts to try. T he most deter mi ne d of t hese efforts ca me with my choice of Marshall Nirenberg’s ne wly proclai me d ne urobiolo- gy laboratory for post doctoral training. Ho wever, in o ur first conversation after my arrival at t he Natio nal I nstit utes of Healt h i n Bet hes da, Mars hall aske d me to establis h a n assay for cyclic A M P i n his laboratory. Tra p pe d a g ai n, b ut I di d n’t fi g ht b a c k v er y vi g or o usl y. Rall a n d S ut herla n d’s discovery of cyclic A M P a n d a de nylyl cyclase, t he hor mone- sensitive enzy me that synthesizes the cyclic n ucleoti de fro m A T P, gave birt h to t he co nce pts of tra ns me mbra ne sig nali ng a n d of hor mone- reg ulate d sy nt hesis of i ntracell ular seco n d messe ngers (Fig. 1). Bot h me n were trai ne d as bioc he mists (S ut herla n d wit h Carl Cori, Rall wit h Albert Le h ni nger), a n d toget her t hey i nitiate d a classical re d uctio nistic a p proac h to deci phering hor mone action. In the 1950’s, hor mones co ul d al most be d efi- ne d as reg ulatory molec ules t hat wo ul d act o nly o n i ntact cells. S ut herla n d a n d Rall’s co u p was to asse mble a s yste m i n w hic h a c haracteristic effect of epinephrine an d glucagon (activation of phosphorylase) coul d be observe d Alfr e d G. Gil m a n 1 8 3 i n ho moge nates a n d t he n to dissect t he syste m i nto its major co m po ne nts - hor mo ne-sti m ulate d sy nt hesis of a factor, cyclic A M P, by t he partic ulate frac- tio n a n d s u bse q ue nt actio n of t he factor i n t he c ytosol to acti vate phospho- r ylase (1). A n assa y (albeit tort uro us) for a de n yl yl c yclase was i n ha n d, a n d hor mone action co ul d then be st u die d by a d ding A T P to plas ma me mbranes THE SEC OND MESSENGER SYSTE M I NV OLVI N G ADENYL CYCLASE EF FE CT O R CELL A T P 5’- A M P I + + + + PHOSPHDIESTERASE A D E NI N E ( O R + P Pi 0’ O H Altho ugh the conce pt of rece ptors for en dogeno us reg ulatory molec ules an d dr ugs arose wit h t he p har macological ex peri me nts of La ngley a n d E hrlic h i n t he late ni netee nt h a n d early t we ntiet h ce nt uries, t he wor d evoke d o nly m et a p h ysi c al f e eli n gs i n m a n y at t h e ti m e of t h e dis c o v er y of c y cli c A M P. T h e ter m “rece ptor” does not a p pear i n t he i n dex of t he 1955 e ditio n of T he Phar macological Basis of Therape utics, the stan dar d textbook of Phar macology, b ut t h e f oll o wi n g s e nt e n c e is t h er e: “ Years ago, La ngley na me d t he differe n- ti ati n g s u bst a n c e t h e ‘r e c e pti v e s u bst a n c e’; t his t er m is still wi d el y e m pl o y e d, b ut it m ust be realize d t hat t he ‘rece ptor’ ma y not be a mor p holo gicall y de monstrable str uct ure.” Rall a n d S ut herla n d’s ex peri me nts provi de d test- t ube assays for rece ptors, a n d t he assays de mo nstrate d t hat t he rece ptors were a ut he ntic. T he effects of e pi ne p hri ne a n d congeners o n a d e n yl yl c y cl a- se were s ho w n to co nfor m to A hlq uist’s ne w co nce pt ualizatio n of β -a dre ner- gic rece ptors (as disti ng uis he d fro m a rece ptors), a n d t he effects were blocke d wit h t he first, ne wly discovere d β -a dre nergic a ntago nist (2). Bioc he mical a p proac hes to rece ptors were t h us bor n, a n d t he q uestio n arose 1 8 4 P hysiology or Me dici ne 1994 of t h e r el ati o ns hi p of t h e β -a drenergic rece ptor to a de nylyl cyclase. Co ul d t he e nzy me be t he rece ptor? Per ha ps, b ut t his mo del wo ul d de ma n d t he exi- ste nce of a fa mil y of a de n yl yl c yclases wit h disti nct re g ulator y sites, beca use re g ulatio n of t he e nz y me was s ho w n not to be restricte d to e pine phrine a n d glucagon; A C T H, TS H, L H, A D H, an d other sti mulators were soon in evi- de nce, as was i n hi biti o n of a de n yl yl c yclase acti vit y b y c h oli ner gic a g o nists { 3}.