Yx. Tao et al., Constitutive activation of G protein-coupled receptors as a result of selective substitution of a conserved leucine residue in transmembrane helix III, MOL ENDOCR, 14(8), 2000, pp. 1272-1282
Whereas numerous mutations of the human lutropin receptor (hLHR) and human
TSH receptor (hTSHR) have been shown to cause constitutive activation of th
ese receptors, it has been suggested that either the hFSHR as a whole, or t
he i3/TM VI region of the hFSHR, is less susceptible to mutation-induced co
nstitutive activation. However, as shown herein, substitution of a highly c
onserved leucine residue in transmembrane III (TM III) of the I hFSHR (Leu
III.18) with arginine causes a B-fold increase in basal cAMP in transfected
cells, consistent with a strong constitutive activation of the hFSHR. Inte
restingly, this mutant is unresponsive to further hormonal stimulation. Sub
stitutions of hFSHR(L460) with lysine, alanine, or aspartate show that only
arginine causes constitutive activation. However, all result in decreased
FSH responsiveness, suggesting a role for L460 in FSH-stimulated cAMP produ
ction by the hFSHR. Because Leu III.18 is highly conserved in rhodopsin-lik
e G protein-coupled receptors (GPCRs), we tested the effects of substitutio
n of the comparable leucine in the human beta(2)-adrenergic receptor (h bet
a(2)-AR). Substitution of L124 in the h beta(2)-AR with arginine, lysine, o
r alanine resulted in constitutive activation as evidenced by increased bas
al levels of cAMP that could be attenuated by an inverse agonist. In all ca
ses, isoproterenol-stimulated cAMP was unaffected. Taken altogether, our da
ta support a model whereby Leu III.18 may play a general role in GPCRs by s
tabilizing them in an inactive state. Constitutive activation may arise by
both a disruption of Leu III.18 as well as the introduction of a specific r
esidue that serves to stabilize the active state of the receptor.