Molybdate stabilized rat uterine progesterone receptors: evidence for two mechanisms.
Molybdate (Mo), EGTA, or protease inhibitors substantially increase detectable rat uterine progesterone (Pg) receptors. Rehomogenization experiments demonstrated that receptor levels decreased in the absence of Mo or protease inhibitors and were not regenerated. Thus Mo prevents an EGTA and protease-inhibitor-sensitive loss of uterine Pg receptors during homogenization. This effect was compared with receptor stabilization at elevated temperature. In contrast to the stability of receptors in the presence of Mo, receptors decreased rapidly to minimal levels by 30 min at 30 degrees C in TESHG (10 mM Tris, 1.5 mM EGTA, 12 mM thioglycerol, 10% glycerol) or TG buffers. The ability of EGTA to mimic receptor stabilization by Mo during homogenization, compared with its ineffectiveness at 30 degrees C, suggested fundamentally different mechanisms for these two phenomena. Similarly, 0.3 M KCl prevented Mo stabilization of the receptors at 30 degrees C, but did not change their recovery after homogenization. Results with protease inhibitors were also consistent: addition of 2-5 mM leupeptin and 500 microM PMSF to TG during homogenization resulted in substantially increased (P less than 0.01) receptor recovery, but leupeptin (+/- the temperature-labile PMSF) did not prevent the Pg receptor losses at 30 degrees C. The transformation state of the receptors may be important, since receptors were untransformed in the presence of either EGTA or Mo. Moreover, KCl transformed the receptors in parallel to their instability at 30 degrees C. In conclusion, Mo stabilizes Pg receptors during temperature elevation by a different mechanism from that involved during homogenization. Although the parallel effects by EGTA, molybdate, and the protease-inhibitors during homogenization is consistent with inhibition of Ca2+-dependent proteolysis, other possible mechanisms must be considered in future studies.
Meador
J
J
Department of Physiology, Tulane University School of Medicine, New Orleans, LA 70112.
Ilenchuk
T T
TT
Walters
M R
MR
D000818:Animals; D004355:Drug Stability; D005260:Female; D007700:Kinetics; D008982:Molybdenum / Q000494:pharmacology*; D010052:Ovariectomy; D011397:Promegestone / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011980:Receptors, Progesterone / Q000187:drug effects / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism*
D011980:Receptors, Progesterone; C044659:molybdate; D008982:Molybdenum; D011397:Promegestone