Histamine H2 receptor mediated dual signaling: Mapping of structural requirements using beta 2 adrenergic chimeric receptors

Previously we demonstrated that the histamine H2 receptor can activate both the adenylate cyclase and phosphoinositide/protein kinase (PBC) signaling pathways. Although dual coupling occurs via separate GTP-dependent mechanis ms the structural components of the H2 receptor directing differential sign aling have not been established. We explored this question by attempting to confer to the beta 2-adrenergic receptor (PAR), which is known to stimulat e cAMP formation, the ability to activate PHC through the construction of b eta 2/H2 chimeric receptors. Intracytoplasmic domains of the human beta 2 a drenergic receptor were substituted with the corresponding sequences of the human H2 receptor and stably expressed in HEK-293 cells. Binding of [H-3]- CGP to chimeric wild type beta 2 receptors was comparable. Substitution of the second intracellular loop (2i) of the PAR led to a significant decrease in coupling to adenylate cyclase while leading to a 139.5 +/- 9.4% control increase in epinephrine mediated PKC activation. Introduction of the H2 re ceptor 3i also led to a decrease in PAR mediated cAMP generation but provid ed the latter with the ability to stimulate PHC (182.2 +/- 8% of control). Concomitant expression of both 2i and 3i led to a substantial increase in e pinephrine mediated PKC activation (201.8 +/- 10.5% of control). Addition o f the carboxyl terminal tail did not facilitate stimulation of PRC. In summ ary, the third intracellular loop of the H2 receptor plays an essential. ro le in activating PKC with maximal efficiency conferred by the second intrac ellular domain. (C) 2000 Academic Press.