An Isoform of the Coactivator AIB1 That Increases Hormone and Growth Factor Sensitivity Is Overexpressed in Breast Cancer

Abstract

The AIB1 (amplified in breast cancer 1) protein is a coactivator that potentiates the transcriptional activity of nuclear hormone receptors, and its gene is amplified in a subset of human breast cancers. Here we report a splice variant of AIB1 mRNA that lacks the exon 3 sequence. We determined that the AIB-⌬3 mRNA encoded a 130-kDa protein that lacks the NH 2 -terminal basic helix-loop-helix and a portion of the PAS (Per-Arnt-Sim homology) dimerization domain. The 130-kDa protein was detected in MCF-7 breast cancer cells at levels that were 5-10% of the full-length protein, whereas in nontransformed mammary epithelium lines, the AIB-⌬3 protein was present at significantly lower levels compared with the full-length AIB1. Consistent with this finding, the abundance of AIB1-⌬3 mRNA was increased in human breast cancer specimens relative to that in normal breast tissue. To determine whether there were phenotypic changes associated with the overexpression of the AIB-⌬3 isoform, we performed functional reporter gene assays. These revealed that the ability of AIB1-⌬3 to promote transcription mediated by the estrogen or progesterone receptors was significantly greater than that of the full-length protein. Surprisingly, the AIB1-⌬3 isoform was also more effective than AIB1 in promoting transcription induced by epidermal growth factor. Overexpression of AIB1-⌬3 may thus play an important role in sensitizing breast tumor cells to hormone or growth factor stimulation.Ligands such as estrogen and progesterone that interact with nuclear receptors regulate gene expression predominantly at the transcriptional level. The ligand-bound receptors interact specifically with DNA and activate transcription by recruiting a preinitiation complex. Although such gene activation was originally thought to be mediated by interaction of the receptors with components of the basal transcriptional machinery (1-6), a variety of screening techniques has identified a family of receptor-interacting proteins known as nuclear receptor coactivators (7-11). A common characteristic of this superfamily of proteins is that, when overexpressed in the presence of nuclear receptors, they potentiate ligand induction of transcription (12, 13). Members of the related p160 group of coactivators, which include steroid receptor coactivator 1 (SRC-1), 1 SRC-2, and SRC-3 (also known as AIB1, ACTR, RAC3, TRAM-1, and p/CIP) (14 -20), possess several similar structural features including a receptor interaction domain, a bHLH (basic helix-loop-helix)-PAS (Per-Arnt-Sim homology) dimerization domain, and a CBP interaction domain (13). Coactivators are thought to function as bridges between nuclear receptors and either other coactivators or the basal transcriptional machinery (13). However, the discovery that coactivators possess a histone acetylase domain (15,(21)(22)(23)(24) suggests that these proteins also might serve to regulate chromatin structure.A portion of human chromosome 20q that is frequently amplified in breast cancer contains the gene for the nuclear coactiva...

Impact of the nuclear receptor coactivator AIB1 isoform AIB1-Δ3 on estrogenic ligands with different intrinsic activity

“…We transfected CHO cells with an expression vector for ER-a, a luciferase reporter plasmid containing a multimerized ERE and with expression vectors encoding either fulllength AIB1 or AIB1-D3. The transfection of cells with the AIB1 expression vector resulted in a 1.3-fold increase in estrogen activation of luciferase activity, whereas transfection with the AIB1-D3 expression vector resulted in a significant 1.9-fold increase in the estrogen response (Figure 1a), thereby confirming the increased efficacy reported by us earlier (Reiter et al, 2001). It is of note that significantly less protein is produced from the transfected AIB1-D3 vector than from the full-length AIB1 (Figure 1a, Western blot inset), implying that the increase in response on a molar basis is even greater for AIB1-D3.…”

“…We previously reported that AIB1-D3, at saturating levels of 17b-estradiol, can induce a larger estrogen response in a transcriptional assay than full-length AIB1 (Reiter et al, 2001). As physiologic hormone interactions often do not take place at saturating hormone levels, we expanded this study and evaluated the response of AIB1-D3 to different concentrations of 17b-estradiol, at both ER-a and ER-b.…”

“…Alterations in this domain are of interest since it has been shown that the bHLH/ PAS domain can mediate homodimerization between family members and in this case may mediate crosstalk between nuclear receptors and bHLH/PAS family members that have wide ranging effects in cellular growth control (Aravind and Ponting, 1997;Fan and Crews, 1999). Surprisingly, overexpression of the AIB1-D3 isoform potentiates ER, progesterone receptor (PR) and epidermal growth factor (EGF) induction of transcription to a much greater extent than AIB1 (Reiter et al, 2001), suggesting that the overexpression of this isoform may well play a role in sensitizing breast cancer cells to hormones during breast cancer development.…”

“…An important finding was that a splice variant of AIB1, in which exon 3 is deleted (AIB1-D3), is overexpressed in MCF-7 breast cancer cells, as well as in breast tumor tissue (List et al, 2001b;Reiter et al, 2001). The predicted protein product from this isoform is an N-terminally truncated version of AIB1 that has a deleted bHLH domain and most of the PAS region removed (Reiter et al, 2001).…”

“…The predicted protein product from this isoform is an N-terminally truncated version of AIB1 that has a deleted bHLH domain and most of the PAS region removed (Reiter et al, 2001). Alterations in this domain are of interest since it has been shown that the bHLH/ PAS domain can mediate homodimerization between family members and in this case may mediate crosstalk between nuclear receptors and bHLH/PAS family members that have wide ranging effects in cellular growth control (Aravind and Ponting, 1997;Fan and Crews, 1999).…”