Salt-induced activation of 1,25-dihydroxyvitamin D3 receptors to a DNA binding form.
In this report we examine the DNA-cellulose binding and sedimentation properties of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) receptors from rat intestine and cultured human mammary cancer cells (MCF-7) extracted in nonactivating (low salt) buffers. Receptors prepared in hypotonic buffer had low DNA binding (13%) compared to receptors extracted with 0.3 M KCl (50%). Treatment of low salt receptor preparations with KCl significantly increased (approximately 3-fold) DNA-binding (activation), demonstrating that receptors can be "activated" in vitro. Activated receptors eluted from DNA-cellulose at 0.18 M KCl. Sedimentation analysis followed by DNA-cellulose binding indicated that activated receptors are approximately 3.2 S and unactivated receptors 5.5 S in size. These results suggest that dissociation of an aggregated moiety may lead to receptor activation. Treatment of unactivated receptor with RNase did not alter DNA binding or sedimentation properties of the aggregated receptor. Treatment of unactivated receptor complexes with heat did not increase DNA binding, and molybdate did not block subsequent salt activation. In summary these results suggest that 1,25(OH)2D3 receptors undergo a salt-induced activation step similar to that described for other steroid receptor systems. However, 1,25(OH)2D3 receptors differ from other steroid receptors in not exhibiting heat activation nor having salt activation blocked by molybdate.
D000818:Animals; D001943:Breast Neoplasms / Q000032:analysis; D002021:Buffers; D002460:Cell Line; D002482:Cellulose / Q000031:analogs & derivatives / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000031:analogs & derivatives / Q000378:metabolism*; D006801:Humans; D007422:Intestines / Q000032:analysis; D008297:Male; D009994:Osmolar Concentration; D011189:Potassium Chloride / Q000494:pharmacology*; D051381:Rats; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid / Q000187:drug effects / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000494:pharmacology
D002021:Buffers; C033190:DNA-cellulose; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid; D011189:Potassium Chloride; D002482:Cellulose; D004247:DNA; D012259:Ribonuclease, Pancreatic