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Pseudohypoparathyroidism, a Novel Mutation in the βγ-Contact Region of Gsα Impairs Receptor Stimulation

1996; Elsevier BV; Volume: 271; Issue: 33 Linguagem: Inglês

10.1074/jbc.271.33.19653

1083-351X

Zvi Farfel, Taroh Iiri, Hagit Shapira, Abraham Roitman, Meir Mouallem, Henry R. Bourne,

Pancreatic function and diabetes

Pseudohypoparathyroidism, type Ia (PHP-Ia), is a dominantly inherited endocrine disorder characterized by resistance to hormones that act by stimulating adenylyl cyclase. It is caused by inheritance of an autosomal mutation that inactivates the α subunit (αs) of Gs, the stimulatory regulator of adenylyl cyclase. In three members of a family, the PHP-Ia phenotype is associated with a mutation (R231H) that substitutes histidine for an arginine at position 231 in αs. We assessed signaling function of αs-WT versus αs-R231H transiently transfected in HEK293 cells. Hormone receptor-dependent stimulation of cAMP accumulation in cells expressing αs-R231H is reduced by ∼75% in comparison to cAMP accumulation in cells expressing αs-WT. A second mutation, αs-R201C, inhibits the GTPase turnoff reaction of αs, thus producing receptor-independent stimulation of cAMP accumulation. The double mutant, αs-R231H/R201C, stimulates cAMP accumulation almost as well (∼80%) as does αs-R201C itself, indicating that the R231H mutation selectively impairs receptor-dependent signaling. In three-dimensional structures of G protein heterotrimers, Arg-231 is located in a region, switch 2, that is thought to interact with the βγ subunit rather than with the hormone receptor. Thus, the R231H phenotype suggests that switch 2 (perhaps in concert with βγ) mediates G protein activation by receptors at a site distant from the receptor-G protein contact surface. Pseudohypoparathyroidism, type Ia (PHP-Ia), is a dominantly inherited endocrine disorder characterized by resistance to hormones that act by stimulating adenylyl cyclase. It is caused by inheritance of an autosomal mutation that inactivates the α subunit (αs) of Gs, the stimulatory regulator of adenylyl cyclase. In three members of a family, the PHP-Ia phenotype is associated with a mutation (R231H) that substitutes histidine for an arginine at position 231 in αs. We assessed signaling function of αs-WT versus αs-R231H transiently transfected in HEK293 cells. Hormone receptor-dependent stimulation of cAMP accumulation in cells expressing αs-R231H is reduced by ∼75% in comparison to cAMP accumulation in cells expressing αs-WT. A second mutation, αs-R201C, inhibits the GTPase turnoff reaction of αs, thus producing receptor-independent stimulation of cAMP accumulation. The double mutant, αs-R231H/R201C, stimulates cAMP accumulation almost as well (∼80%) as does αs-R201C itself, indicating that the R231H mutation selectively impairs receptor-dependent signaling. In three-dimensional structures of G protein heterotrimers, Arg-231 is located in a region, switch 2, that is thought to interact with the βγ subunit rather than with the hormone receptor. Thus, the R231H phenotype suggests that switch 2 (perhaps in concert with βγ) mediates G protein activation by receptors at a site distant from the receptor-G protein contact surface.