Luteinizing hormone receptors translocate to plasma membrane microdomains after binding of human chorionic gonadotropin.
Receptor-mediated signal transduction by G protein-coupled receptors can involve redistribution of plasma membrane receptors into membrane structures that are characterized by insolubility in Triton X-100 and low buoyant density in sucrose gradients. Here we describe the translocation of wild-type (wt) rat LH receptors (LHR-wt) from the bulk membrane into membrane microdomains (rafts) after the binding of human chorionic gonadotropin (hCG). In sucrose gradient ultracentrifugation of plasma membranes from cells stably expressing FLAG-tagged LHR-wt, receptors were located in high-density membrane fractions before binding of hormone and in low-density fractions after hCG treatment. Receptor translocation to low-density sucrose fractions did not occur when cells were pretreated with 1% methyl-beta-cyclodextrin, which reduces membrane cholesterol and disrupts rafts. Single-particle tracking of individual FLAG-LHR-wt receptors showed that hCG-treated receptors become confined in small compartments with a diameter of 86 +/- 36 nm, significantly smaller than 230 +/- 79 nm diameter regions accessed by the untreated receptor. Receptors were no longer confined in these small compartments after disruption of rafts by methyl-beta-cyclodextrin, a treatment that also decreased levels of cAMP in response to hCG. Finally, translocation of LHR into rafts required a functional hormone-receptor complex but did not occur after extensive receptor cross-linking that elevated cAMP levels. Thus, retention of LHR in rafts or small membrane compartments is a characteristic of functional, hormone-occupied LHR-wt. Although raft translocation was not essential for cAMP production, it may be necessary for optimizing hormone-mediated signaling.
Smith
Steven M L
SM
Department of Biomedical Sciences, Colorado State University, Fort Collins, 80523, USA.
Lei
Ying
Y
Liu
Jingjing
J
Cahill
Mary E
ME
Hagen
Guy M
GM
Barisas
B George
BG
Roess
Deborah A
DA
D000818:Animals; D016466:CHO Cells; D002784:Cholesterol / Q000502:physiology; D006063:Chorionic Gonadotropin / Q000378:metabolism*; D006224:Cricetinae; D000242:Cyclic AMP / Q000502:physiology; D021962:Membrane Microdomains / Q000378:metabolism*; D021381:Protein Transport; D051381:Rats; D011974:Receptors, LH / Q000378:metabolism*; D015398:Signal Transduction; D047392:beta-Cyclodextrins / Q000494:pharmacology
D006063:Chorionic Gonadotropin; D011974:Receptors, LH; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D002784:Cholesterol; D000242:Cyclic AMP