“Defective” Receptors in Steroid-Resistant Conditions may be Proteolytic Artifacts

  • Merry R. Sherman
  • Fe B. Tuazon
  • Yee-Wan Stevens
  • Joseph A. Carlino
  • En-Mei Niu
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 196)


The specific question addressed in this report is whether the resistance to steroid treatment of certain tissues or tumors which appear to contain a normal quantity of steroid-binding sites may be due to stuctural defects in the receptors. This question may be seen as part of the more general question of whether there are intrinsic variations in the structures of receptors for a given class of steroids in different healthy tissues, in healthy vs. malignant tissues or in different types of tumors. Our experimental approach to these questions has involved the stabilization and precise physicochemical characterization of the receptors. To date, we have studied the estrogen and progestin receptors from human breast cancers and benign and malignant gynecologic specimens and the glucocorticoid receptors from several healthy and malignant rodent tissues and from normal human lymphocytes and various types of leukemic cells. Chromatographic and ultracentrifugal analyses in buffers of low ionic strength, containing 20 mM Na2MoO4 as the stabilizer, have revealed each of these receptors to be a large, oligomeric complex, characterized by remarkably similar values of the Stokes radius, sedimentation coefficient, molecular weight and axial ratio.

In the absence of adequate stabilization, however, we found that the receptors for three classes of steroids in extracts of some healthy, steroid-responsive tissues, such as rat kidney and human uterine endometrium, are invariably degraded by endogenous proteinases. The extent of such cleavage is increased considerably by freezing the tissues prior to homogenization. Studies designed to distinguish the intact receptors from the products of proteolysis have included the characterization of receptors in cytosols prepared from mixtures of rat liver and kidney. The results strongly support the interpretation that the smaller size of the receptors detected in kidney cytosol reflects their cleavage by the more active proteinases in that tissue.

The sizes and shapes of the receptors in cytosols from various tissues were found to be correlated with the activities of specific endopeptidases, assayed fluorometrically with peptidyl derivatives of 7-amino-4-methylcoumarin (AMC). These studies suggested that the receptors are vulnerable to cleavage by “lysine-specific” endopeptidases, detected with t-butyloxycarbonyl-L-valyl-L-leucyl-L-lysyl-AMC. An enzyme of this specificity was partially purified from rat kidney cytosol and tested for its ability to digest the glucocorticoid receptors from rat liver cytosol. Under the conditions used, about 40% of the receptors were converted to the mero-receptor, the smallest fragment containing the steroid-binding site.

Since the cleavage of steroid receptors in vitro has been amply documented, we conclude that the observation of altered receptors in extracts of tissues or tumors that are resistant to steroid treatment does not prove that this resistance is due to intrinsic structural defects in the receptors.


Acute Lymphoblastic Leukemia Glucocorticoid Receptor Axial Ratio Triamcinolone Acetonide Limited Proteolysis 
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Copyright information

© Plenum Press, New York 1986

Authors and Affiliations

  • Merry R. Sherman
    • 1
  • Fe B. Tuazon
    • 1
  • Yee-Wan Stevens
    • 1
  • Joseph A. Carlino
    • 1
  • En-Mei Niu
    • 1
  1. 1.Endocrine Biochemistry LaboratoryMemorial Sloan-Kettering Cancer CenterNew YorkUSA

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