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Related Experiment Videos

Sequence and expression of human estrogen receptor complementary DNA.

G L Greene, P Gilna, M Waterfield

    Science (New York, N.Y.)
    |March 7, 1986
    PubMed
    Summary

    Researchers expressed a functional human estrogen receptor (ER) in cells, confirming its identity and structure. This finding provides insights into how steroid hormone receptors regulate gene expression and their evolutionary origins.

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    Area of Science:

    • Molecular Biology
    • Genetics
    • Endocrinology

    Background:

    • The precise mechanisms of steroid hormone receptor gene regulation in eukaryotes remain unclear.
    • Estrogen receptor (ER) function is critical in cellular processes, including breast cancer development.

    Purpose of the Study:

    • To clone, sequence, and express a functional human estrogen receptor (ER) in a non-human cell line.
    • To characterize the expressed ER protein and investigate its structural and functional properties.
    • To explore evolutionary relationships between ER and other related proteins.

    Main Methods:

    • Sequencing of a complementary DNA (cDNA) clone encoding the full translated portion of the human ER mRNA from MCF-7 breast cancer cells.
    • Expression of the ER cDNA in Chinese hamster ovary (CHO-K1) cells to produce a functional protein.

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  • Biochemical characterization including [3H]estradiol binding assays, sucrose gradient sedimentation, and monoclonal antibody confirmation.
  • Amino acid sequence comparison with other steroid receptors and oncogene products.

    • Main Results:

    • A functional human ER protein was successfully expressed in CHO-K1 cells.
    • The expressed protein exhibited specific [3H]estradiol binding and characteristic sedimentation properties (4S in salt, 8-9S without salt).
    • Monoclonal antibody analysis confirmed the identity of the expressed protein as human ER.
    • Significant regional homology was identified between human ER, human glucocorticoid receptor, and the v-erbA oncogene product, particularly in a cysteine-, lysine-, and arginine-rich region.

    Conclusions:

    • The study successfully produced and characterized a functional human estrogen receptor in a heterologous expression system.
    • The findings support the hypothesis that steroid receptor genes and the v-erbA oncogene share a common evolutionary origin.
    • The identified homologous region is proposed to be the DNA-binding domain for these related proteins.