Estrogen Receptor ß: Mine Is Longer Than Yours?a

Etienne Leygue, Helmut Dotzlaw, Biao Lu, Cheryl Glor, Peter H. Watson and Leigh C. Murphy

University of Manitoba Winnipeg, Manitoba, R3E OW3, Canada

Two years after the cloning of the second estrogen receptor, ER-ß, the primary sequence encoding the N-terminus of the protein still remains uncertain. The demonstrated importance of estrogen signalling in normal and abnormal development of multiple tissues justifies characterization of this region.

Estrogens, involved in the normal development of a wide tissue spectrum including breast, uterus, brain, and bone, are also implicated in several diseases such as breast and endometrial cancers and osteoporosis. In 1995, the cloning (1) of ER-ß from a rat prostate cDNA library, led to the need to fully re-evaluate estrogen signalling in target tissues. Like estrogen receptor {alpha} (2), rER-ß is a ligand-dependent transcription factor that binds estrogen and antiestrogen. Relying on the presence of an in-frame stop codon upstream of their coding sequence, Kuiper et al. (1) considered their open reading frame to encode the full length protein (R1, Fig. 1Go).Shortly thereafter, a human homologue, hER-ß, was cloned (3) from testis (H1, Fig. 1Go). This cDNA encoded a protein 8 amino-acids shorter than rER-ß. In 1997, the hER-ß cDNA sequence was extended (4) to a start codon corresponding to the first rat methionine codon (H2, Fig. 1Go), and the mouse homologue, mER-ß, was cloned (5) (M1, Fig. 1Go).



View larger version (73K):
[in this window]
[in a new window]
 
Figure 1. Schematic representation of rat, human, and mouse ER-ß protein. The amino acid composition of N-terminal extremities of ER-ß proteins predicted from cloned cDNAs is shown. For each species (rat R, human H, mouse M), initiating methionine codons are indicated in their chronological order of identification (1 2 3 ). Similarities between the three species are indicated by blue boxes. Sequences underlined by a single and double line represent putative phosphorylation and glycosylation motifs, respectively. Genbank accession numbers of sequences encoding R1, R2, H1, H3, M1, M2, and M3 are U57439, AJ002602, X99101, AB006590, U81451, AF063853, and AF067422, respectively.

 
In 1998, following cDNA library screening and/or polymerase chain reaction, the N-terminus of ER-ß was extended in all three species. Ogawa et al. (6) isolated from human testis a hER-ß cDNA that could encode 45 additional amino-acids (H3, Fig. 1Go). This sequence plus the presence of an upstream in-frame stop codon was recently confirmed by Moore et al. (7). In April 1998, a prostate rER-ß cDNA sequence was submitted to Genbank, that differs from the initial sequence by only one base upstream of the start codon. This extra base alters frame and removes the previously observed in-frame stop codon, resulting in a cDNA that could encode 64 additional N-terminal amino-acids (R2, Fig. 1Go). In May 1998, two sequences submitted to Genbank extended the initial 5'-extremity of the mER-ß cDNA. The encoded mER-ß proteins contain 45 and 64 extra amino-acids N-terminal to M1 (M2 and M3, Fig. 1Go), respectively. All three species present strong sequence similarities in this N-terminal region, although the rodent open reading frames are 19 amino-acids longer than the human.

Altogether, these observations raise important questions. Are cloning strategies and/or tissue/species-specific expression of different forms of ER-ß responsible for observed discrepancies? Do longer forms of ER-ß, as yet unidentified, exist? If so, how is expression regulated? What are the functions associated with these extra amino-acids? The presence of putative phosphorylation and glycosylation motifs within these additional amino-acid sequences suggest possible regulation of putative function(s). Moreover, because of the demonstrated importance of the N-terminal region in ER-{alpha}, particularly the involvement of the AF-1 domain in hormone independent activation of the receptor (8), the elucidation of the function(s) associated with these new regions is necessary. The majority of functional studies undertaken to date were performed with constructs lacking these N-terminal amino-acids: addressing the above questions is critical to fully understanding the involvement of ER-ß in estrogen signalling.

Footnotes

Address correspondence to: Etienne Leygue, Department of Biochemistry and Molecular Biology, University of Manitoba, 770 Bannatyne Avenue, Winnipeg, Manitoba R3E 0W3 Canada.

Received June 30, 1998.

References

  1. Kuiper GG, Enmark E, Pelto-Huikko M, Nilsson S, Gustafsson JA. 1996 Cloning of a novel receptor expressed in rat prostate and ovary. Proc Natl Acad Sci USA. 93:5925–5930.[Abstract/Free Full Text]
  2. Green S, Walter P, Kumar V, et al. 1986 Human estrogen receptor cDNA: sequence, expression, and homology to v- erb-A. Nature. 320:134–139.[Medline]
  3. Mosselman S, Polman J, Dijkema R. 1996 ER-ß: identification and characterization of a novel human estrogen receptor. FEBS Lett. 392:49–53.[CrossRef][Medline]
  4. Enmark E, Pelto-Huikko M, Grandien K, et al. 1997 Human estrogen receptor ß-gene structure, chromosomal localization, and expression pattern. J Clin Endocrinol Metab. 82:4258–4265.[Abstract/Free Full Text]
  5. Tremblay GB, Tremblay A, Copeland NG, et al. 1997 Cloning, chromosomal localization, and functional analysis of the murine estrogen receptor beta. Mol Endocrinol. 11:353–365.[Abstract/Free Full Text]
  6. Ogawa S, Inoue S, Watanabe T, et al. 1998 The complete primary structure of human estrogen receptor ß (hER-ß) and its heterodimerization with ER-{alpha} in vivo and in vitro. Biochem Biophys Res Commun. 243:122–126.[CrossRef][Medline]
  7. Moore JT, McKee DD, Slentz-Kesler K, et al. 1998 Cloning and characterization of human estrogen receptor beta isoforms. Biochem Biophys Res Commun. 247:75–78.[CrossRef][Medline]
  8. Kumar V, Green S, Stack G, Berry M, Jin JR, Chambon P. 1987 Functional domains of the human estrogen receptor. Cell. 51:941–951.[Medline]