The rotational diffusion of LH receptors differs when receptors are occupied by hCG versus LH and is increased by cytochalasin D.
We have examined the rotational diffusion of the luteinizing hormone (LH) receptors binding human chorionic gonadotropin (hCG) or ovine luteinizing hormone (oLH) in MA-10 Leydig tumor cells using time-resolved phosphorescence anisotropy techniques. LH receptors binding erythrosin isothiocyanate (ErITC)-derivatized oLH were rotationally mobile with rotational correlation times of 62 micros, 48 micros, 38 micros, and 29 micros at 4 degrees C, 15 degrees C, 25 degrees C, and 37 degrees C, respectively. ErITC-hCG bound to the LH receptor was rotationally immobile, showing no anisotropy decay at 4 degrees C, 15 degrees C, 25 degrees C, and 37 degrees C. To determine whether cytoskeletal components influenced the rotational diffusion of LH receptors, we measured rotational diffusion of LH receptors on MA-10 cells treated with 20 microg/ml cytochalasin D and on plasma membrane preparations. Following 1 h exposure to cytochalasin D, the rotational correlation times for hCG-occupied LH receptors were typically 11 micros at 37 degrees C compared to > 1000 micros on untreated cells. Treatment of MA-10 cells with cytochalasin B or colchicine had no affect on LH receptor rotational diffusion. Rotational correlation times for LH-occupied receptors decreased from 29 micros to 12 micros at 37 degrees C following cytochalasin D treatment. The rotational diffusion of LH receptors on plasma membrane preparations was similar to that observed for LH- and hCG-occupied receptors on intact cells treated with cytochalasin D. These various results indicate that there are differential effects of LH and hCG binding on the interactions of LH receptors with plasma membrane proteins and that microfilaments anchor the hCG- and LH-occupied receptors.
Roess
D A
DA
Department of Physiology, Colorado State University, College of Veterinary Medicine, Fort Collins 80523, USA. daroess@lamar.colostate.edu
Jewell
M A
MA
Philpott
C J
CJ
Barisas
B G
BG
D008841:Actin Cytoskeleton / Q000187:drug effects; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006063:Chorionic Gonadotropin / Q000378:metabolism*; D003078:Colchicine / Q000494:pharmacology; D003571:Cytochalasin B / Q000494:pharmacology; D015638:Cytochalasin D / Q000494:pharmacology*; D004058:Diffusion; D005454:Fluorescence Polarization; D007985:Leydig Cells / Q000378:metabolism; D007986:Luteinizing Hormone / Q000378:metabolism*; D008297:Male; D008870:Microtubules / Q000187:drug effects; D011974:Receptors, LH / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D013696:Temperature
D006063:Chorionic Gonadotropin; D011974:Receptors, LH; D015638:Cytochalasin D; D003571:Cytochalasin B; D007986:Luteinizing Hormone; D003078:Colchicine