Membrane organization of luteinizing hormone receptors differs between actively signaling and desensitized receptors.
Signaling by the luteinizing hormone/choriogonadotropin receptor (LHR) is of considerable interest because of its requirement for successful reproduction. Time-resolved phosphorescence anisotropy and fluorescence resonance energy transfer were used to investigate the organization of endogenous LHRs in porcine follicular membranes in two distinct signaling states, active and desensitized. Desensitized LHRs exhibited approximately 3-fold slower rotational correlation times compared with active LHRs (59 +/- 4 and 21 +/- 9 mus, respectively), suggesting that with agonist-dependent desensitization the receptors are organized into larger protein complexes. Incubation of membranes with inhibitors of LHR desensitization, such as neutralizing anti-arrestin antibodies, a synthetic peptide corresponding to the third intracellular loop of the LHR but not the corresponding scrambled peptide, or catalytically inactive ARNO, resulted in faster rotational diffusion times equivalent to those of actively signaling LHRs. Furthermore, desensitized LHRs exhibited a 2.4-fold increase in fluorescence resonance energy transfer between LHRs suggesting that the larger protein aggregates formed during desensitization contain more self-associated LHRs. These results indicate that agonist-dependent LHR desensitization precedes organization of LHRs at the cells surface into larger protein aggregates.
Hunzicker-Dunn
Mary
M
Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA. mhd@northwestern.edu
Barisas
George
G
Song
Jinming
J
Roess
Deborah A
DA
D000818:Animals; D016880:Anisotropy; D002462:Cell Membrane / Q000378:metabolism*; D004058:Diffusion; D031541:Fluorescence Resonance Energy Transfer; D008163:Luminescent Measurements; D018613:Microscopy, Confocal; D011485:Protein Binding; D011974:Receptors, LH / Q000737:chemistry* / Q000378:metabolism; D015398:Signal Transduction; D013552:Swine; D013997:Time Factors
D011974:Receptors, LH