Indra Poola

Identification of ten exon deleted ERβ mRNAs in human ovary, breast, uterus and bone tissues: alternate splicing pattern of estrogen receptor β mRNA is distinct from that of estrogen receptor α

Four different human tissues and breast cancer cell lines were screened to identify exon deletion variant transcripts of estrogen receptor β (ERβ) by reverse transcription‐polymerase chain reaction using the ‘splice targeted primer approach’ that amplifies each category of exon deleted variants as a separate gene population. A total of 10 different variant mRNAs that have deletions in various combination of exons were identified by sequence analysis. They were exon 2Δ; exons 2 and 5–6Δ; exon 3Δ; exon 4Δ; exon 5Δ; exons 5 and 2Δ; exon 6Δ; exons 6 and 2Δ; exons 6, 2–3Δ; and exons 5–6Δ. In some cases, deletion of an exon appears to be associated with a mutation of a specific base. Although ERα and ERβ are highly homologous, have identical exon and functional domain organization, exhibit similar ligand‐binding profiles and interact with identical DNA response elements, the sequence of exon skipping in ERβ pre‐mRNA appears to be distinct from that of ERα mRNA. Furthermore, results described here also suggest that alternate splicing of ERβ mRNA is tissue specific. The presence of a ERβ variant profile together with other ER isoforms in a tissue may have functional implications in binding and response to a particular ligand.

Identification of twenty alternatively spliced estrogen receptor alpha mRNAs in breast cancer cell lines and tumors using splice targeted primer approach

Estrogen receptor (ER) alpha splice variant transcript profiles were analyzed by RT PCR in six ER positive breast cancer cell lines, MCF-7, T47D, ZR-75, LCC1, LCC2 and LCC9, three ER negative cell lines, MDA-MB-435, MDA-MB-235 and LCC6, and three ER positive malignant breast tumors using targeted primers which specifically anneal to the splice junctions of exon 2Delta, exon 3Delta, exons 2-3Delta, exon 4Delta, exon 5Delta, exon 6Delta and exon 7Delta. The partner primers were chosen such that largest possible transcripts were amplified between exons 1 and 8. The results described here show that each splice specific primer amplified not only the single exon deleted transcript but also a number of related transcripts that have deletions in various combinations of exons. The exon 2Delta specific primer amplified five transcripts that have deletions in exon 2, exons 2 and 7, exons 2, 5, and 7, exons 2 and 4-5, and exons 2 and 4-6. The exon 3Delta specific primer amplified two transcripts that have deletions in exon 3, and exons 3 and 7. The exon 2-3Delta specific primer amplified three products that have deletions in exons 2-3, exons 2-3 and 7 and exons 2-3, 5 and 7. The exon 4Delta specific primer amplified two products that have deletions in exon 4, and exons 4 and 7. The exon 5Delta specific primer amplified three transcripts, that have deletions in exon 5, exons 5 and 2, and exons 5, and 2-3. The 6Delta specific primer amplified only one transcript that has a deletion in exon 6. The 7Delta specific primer amplified four transcripts, that have deletions in exon 7, exons 7 and 4, exons 7 and 3-4, and exons 7 and 3-5. None of the above splice specific primers amplified the wild type ER sequences. The six ER positive cell lines differed in the patterns of the variant transcripts and among the three ER negative cell lines analyzed, only MDA-MB-435 showed the presence of exon 2Delta and exon 4Delta transcripts. Analyses in the tumor samples indicated that the above transcripts are extensively modified.

Functionally active estrogen receptor isoform profiles in the breast tumors of African American women are different from the profiles in breast tumors of Caucasian women.

Several cancer surveys have shown that African‐American women (AAW) develop highly aggressive breast tumors and experience about three times higher mortality rates compared with other populations. Generally, breast tumors in AAW are poorly differentiated or undifferentiated and exhibit increased frequency of nuclear atypia, higher mitotic activity, higher S‐phase fraction, and tumor necrosis. The molecular factors responsible for these tumor characteristics are mostly unknown.

Alterations in the estrogen receptor alpha mRNA in the breast tumors of African American women

Abstract Several recent reports have shown that the mortality rate with breast cancer is about three times higher in African American women than in other populations. In addition, the available data also indicate that the tumors are very aggressive and poorly differentiated with a very low frequency of hormone receptors. To gain an insight into the factors that may be responsible for their aggressive tumors, we investigated the transcript profiles of the estrogen receptor (ER), the most important prognostic factor in breast cancer, in the tumors derived from African American women. We analyzed 24 immunohistochemically ER+ and 6 ER− malignant tumors for ER mRNA by reverse transcription polymerase chain reaction using a number of primer pairs. For comparative purposes, 20 ER+ malignant tumor issues derived from Caucasian patients were also included. Our results showed that only 15 of the ER+ tumors from African American women patients had full-length wild-type receptor transcripts and the others exhibited alterations/truncations in exon 8. We also found that the majority of tumors that had alterations/truncations in exon 8 did not express the naturally occurring, more abundant exon 7 deletion transcript. Most of the tumors expressed exon 2, exons 2–3, and exon 5 deletion variant transcripts. Unexpectedly, 2 of the 6 immunohistochemically ER− tumors showed full-length wild-type receptor mRNA but none of the variant transcripts.

An estrogen inducible 104 kDa chaperone glycoprotein binds ferric iron containing proteins: a possible role in intracellular iron trafficking

We have previously described an estrogen inducible, intracellular, homodimeric membrane glycoprotein (subunit M r 104 kDa) which is structurally related to ‘chaperone’ proteins (Poola, I. and Kiang J.G., J. Biol. Chem. 269 (1994) 21762–21769). In this report we describe a novel finding that the 104 kDa chaperone protein exhibits affinity for iron containing proteins such as transferrins from several species, human lactoferrin and microbial ferric binding protein (FBP). A single ferric ion in the above proteins appears to be sufficient for binding. It also binds to immobilized ferritin. However, it does not exhibit any affinity for apotransferrins, apolactoferrin, apoferritin and apoFBP. This is the first report of a chaperone protein that exhibits affinity for iron containing proteins.