Abstract

After incubation with [3H]estradiol low-salt sucrose gradient centrifugation of 250 human breast tumor cytosis demonstrated receptor-bound steroid in both 4 S and 8 S forms. Cytosols prepared from 24 of these receptor-positive tumors were examined in the presence of KCl, Ca2+ and the serine protease inhibitor, phenylmethylsulfonyl fluoride. After incubation with KCl (0.4 M), interconversion of 8 S and 4 S forms was found in one-half of the 24 cytosols examined whose original low-salt patterns contained well-defined 4 S and 8 S peaks. When cytosols were made 4 mM in Ca2+ 15 to 30 min before the addition of KCl, an additional 4.5 S form was observed in low-salt gradients in 18 tumors, and, in 6 tumors, inactivation of receptor occurred. The effects were not produced when Ca2+ and salt were added simultaneously. After 2 to 24 hr of preincubation with Ca2+ of the same cytosols under the same conditions, an increased recovery of the approximately 4 S receptor complex was observed in low-salt gradient. These Ca2+ effects were inhibited if phenyl-methylsulfonyl fluoride and Ca2+ were added simultaneously to cytosols prior to the addition of salt. The data suggest that human mammary tumor cytosols may contain proteolytic activities that can be activated by Ca2+ and that can effect irreversible changes in the salt-dissociated steroid receptor proteins which can be detected in sucrose density gradients.