The possibility that Spodoptera frugiperda (Sf9) cells can provide an intact cell setting for reconstitution of the human 5-hydroxytryptamine (5-HT) receptor with mammalian G protein subunits was explored. The 5-HT receptor was found to assume an uncoupled phenotype when expressed alone in Sf9 cells at relatively high levels (5-34 pmol of receptor/mg of membrane protein), i.e. agonist-binding to the receptor was characterized by a relatively high K and an insensitivity to GTP. Co-expression of the receptor with members of the ``family'' together with various combinations of and subunits increased the affinity for agonists to that observed for the coupled form of receptor in mammalian cells, concomitant with conferrance of guanosine 5`-(,-imino)triphosphate sensitivity. The agonists employed were [H]8-hydroxy-N, N-dipropyl-2-aminotetralin ([H] 8-OH-DPAT) and [I]R(+)-trans-8-hydroxy-2-[N-n-propyl-N-(3`-iodo-2`-propenyl)amino]tetralin ([I]8-OH-PIPAT). The binding of an antagonist, [I]4-(2`-methoxyphenyl)-1-[2`-[N-(2"-pyridinyl)-p-iodobenzamido]ethyl] piperazine ([I]p-MPPI), was unaffected by co-expression of G protein subunits. Both and subunits were required for optimal coupling. No differences were evident among , , , , and when expressed with in this regard, nor among most permutations of subunits when expressed with ( > ). and expressed with did not participate in coupling. These data support the conclusion that normal interactions between a mammalian receptor and a select array of G proteins can be established in intact Sf9 cells, and extend previous observations of 5-HT receptor coupling to G and the pertussis toxin-insensitive G protein G.

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