Biochemical and Biophysical Research Communications
Identification, cloning, and expression of human estrogen receptor-α36, a novel variant of human estrogen receptor-α66☆,☆☆
Section snippets
Materials and methods
Homology search and cloning of ER-α36. The sequence homology searches were performed with Blastn and Blastp programs of National Center for Biotechnology Information (NCBI) and used to look for short, nearly exact matches using DNA sequences (20–50 bp in length) encoding the ligand-binding domain of ER-α66. We found one full-length cDNA clone (GenBank Accession No. BX640939) from a human uterus cDNA library that was matched exactly to a DNA sequence encoding the ligand-binding domain of hER-α66
Identification of a novel isoform of hER-α66
Previous reports indicated that three predominant bands of 35–39, 46, and 66 kDa were present in Western blots probed with the anti-ER-α antibodies raised against the ligand-binding domain of hER-α66 [16], [18]. Although these and other reports consistently noted a predominant band at 35–39 kDa, there were no reports that described what this protein band represented. In confirmation, we observed three protein bands (36, 46, and 66 kDa) in Western blot analysis of ER-positive MCF7 breast cancer
Discussion
In this study, a novel variant of hER-α66 termed hER-α36 has been identified and cloned; the cDNA that was cloned expresses the predicted 36-kDa protein in HEK 293 cells; Western blot analysis of lysates prepared from HEK 293 cells expressing the cloned cDNA revealed that the hER-α36 protein was recognized by an antibody raised against an epitope common to hER-α36, hER-α46, and hERα-66. The novel hER-α isoform is the product of a transcript initiated from a previously unidentified promoter in
Acknowledgments
We thank Dr. Zafar Nawaz for the expression vectors. The technical assistance of Megan Coleman is also acknowledged. This work was supported by National Institutes of Health Grant RO1 CA84328 (to Z.-Y.W.) and RO1s CA84400 and CA66029 (to T.F.D.).
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2023, Biomedicine and PharmacotherapyAn overview on Estrogen receptors signaling and its ligands in breast cancer
2022, European Journal of Medicinal ChemistryCitation Excerpt :The closing pocket site of the ligand is considered essential for its direct interaction in the transactivation function AF-2 [32]. ER-α36, a novel variant of ER-α and an isoform of ER-α66, was first of all cloned and identified by Wang et al., in 2005 [33]. Human ER-α36 has a molecular weight of approximately 35.7 kDa and lacks both transcriptional activation domains (AF-1 and AF-2), but contains partial ligand-binding domains, DNA binding domains and dimerization domains.
Cancer stem cells in TNBC
2022, Seminars in Cancer BiologyCitation Excerpt :TNBC cells, being negative for ER, PR, and HER2 expression, are generally believed to lack intracellular estrogen signal transduction and are therefore insensitive to endocrine therapy. Interestingly, the novel estrogen variant ER-α36, which lacks the transcriptional activator domains AF-1 and AF-2, has recently been discovered [16]. The well-known TNBC cell lines MDA‑MB‑231 and MDA‑MB‑436 all expressed high levels of ER‑α36, which can activate the MAPK/ERK and PI3K/AKT signaling pathway downstream of the wild-type estrogen receptor ER-a66 [17,18].
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This is manuscript number 17432-MEM from The Scripps Research Institute.
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Abbreviations: ER, estrogen receptor; AF, activation function; E2β, 17β-estradiol; E2α, 17α-estradiol; HEK 293 cells, human embryonic kidney 293 cells; NO, nitric oxide; eNOS, endothelial NO synthase.