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Use of Chromatofocusing to Distinguish Estradiol Receptor from Ovarian-dependent and -independent Rat Mammary Tumors¹

Departments of Surgery and Neoplastic Diseases [A. H. C., M. F., M. P., V. P. H.] and The Center for Laboratory Animal Sciences [L. T., D. D.], Mount Sinai School of Medicine, New York, New York 10029

Cytosol estradiol receptor from MTW9-D (ovarian dependent) and MTW9-MtT (ovarian independent) rat mammary tumors were fractionated by chromatofocusing, a procedure which separates proteins on an ion-exchange column as a function of isoelectric point. Receptor from MTW9-D usually fractionated as three peaks with mean pH at peak height of 7.5, 6.8, and 6.0. The intermediate peak at pH 6.8 was present in 90% of MTW9-D tumors examined but in only 20% of MTW9-MtT tumors. Treatment of cytosols with 20 mM sodium molybdate or 50 mM leupeptin did not change the chromatofocusing profiles.

The pl 6.8 fraction of receptor bound quantitatively to DNA-cellulose after ammonium sulfate precipitation, while receptor in the other two peaks bound much less. The quantitative binding of a fraction of estradiol receptor characteristic of MTW9-D to DNA is consistent with the greater binding of unfractionated cytosol receptor from MTW9-D to DNA than the binding of receptor from MTW9-MtT. Sucrose gradient analysis of the pl 6.8 receptor showed a sedimentation coefficient slightly less than ovalbumin with a Stokes radius of 26 Å as determined by agarose chromatography. The correlation of receptor binding to DNA with response to ovariectomy might make this form of receptor a potential marker of hormonal responsivity in mammary tumors.

¹ This investigation was supported by Grant BC-312 from The American Cancer Society.

² To whom requests for reprints should be addressed.

Received 1/25/83. Accepted 8/ 3/83.