Role of the Nuclear Receptor Coactivator AIB1-Δ4 Splice Variant in the Control of Gene Transcription

Abstract

The oncogene amplified in breast cancer 1 (AIB1) is a nuclear receptor coactivator that plays a major role in the progression of various cancers. We previously identified a splice variant of AIB1 called AIB1-⌬4 that is overexpressed in breast cancer. Using mass spectrometry, we define the translation initiation of AIB1-⌬4 at Met 224 of the full-length AIB1 sequence and have raised an antibody to a peptide representing the acetylated N terminus. We show that AIB1-⌬4 is predominantly localized in the cytoplasm, although leptomycin B nuclear export inhibition demonstrates that AIB1-⌬4 can enter and traffic through the nucleus. Our data indicate an import mechanism enhanced by other coactivators such as p300/CBP. We report that the endogenously and exogenously expressed AIB1-⌬4 is recruited as efficiently as full-length AIB1 to estrogen-response elements of genes, and it enhances estrogen-dependent transcription more effectively than AIB1. Expression of an N-terminal AIB1 protein fragment, which is lost in the AIB1-⌬4 isoform, potentiates AIB1 as a coactivator. This suggests a model whereby the transcriptional activity of AIB1 is squelched by a repressive mechanism utilizing the N-terminal domain and that the increased coactivator function of AIB1-⌬4 is due to the loss of this inhibitory domain. Finally, we show, using Scorpion primer technology, that AIB1-⌬4 expression is correlated with metastatic capability of human cancer cell lines.Gene transcription in eukaryotes is a complex and highly regulated process. One of the major controls of gene transcription is exerted by the coregulator family of proteins. These include both corepressors, which dampen transcription, and coactivators, which potentiate transcription. A subgroup of coactivators has been shown to be critical for the malignant progression of cancer and is known as the p160 steroid receptor coactivators (1). One member in particular was identified to be amplified in breast cancer. Amplified in breast cancer 1 (AIB1, SRC-3, NCOA3, ACTR, TRAM-1, pCIP, and RAC3) has been shown to be a gene amplified in breast cancer (2) and is also overexpressed at the mRNA and protein level in various cancers (1, 3, 4). Its role in tumorigenesis is attributed to its ability to coactivate both steroid hormone-and growth factor-dependent transcription (3, 5-7). In several oncogene-driven mouse models (8 -11), reduction of AIB1 levels leads to a decrease in tumorigenesis, and overexpression of AIB1 leads to the formation of various tumors (12). Clinically, AIB1 expression in breast cancer cases is correlated with high HER2 levels, larger tumor size, higher tumor grade, and shorter disease-free survival (13-15). Also, high levels of AIB1 in conjunction with high HER2 levels coincide with reduced disease-free survival in patients treated with tamoxifen, suggesting a role for AIB1 in tamoxifen resistance (16).We had previously identified a splice variant of AIB1, where exon 3 was spliced from the mature mRNA and the resulting protein named AIB1-⌬3 (17). More recently, an additio...

Coregulators and Their Role in Selective Estrogen Receptor Modulator Action

“…38,39 Overexpression of AIB1-∆4 is associated with increased metastatic potential of breast cancer cells. 39,40 Irrespective of this additional complexity, higher levels of AIB1 mRNA or AIB1 gene amplification in breast cancers are generally predictive of worse disease outcome, higher grade disease, shorter disease-free interval, and reduced overall survival. 13,27,41,42 AIB1 has been widely studied for its role in resistance to SERM therapy in breast cancer both preclinically and clinically.…”

The AIB1/NCOA3/SRC-3 Oncogene

“…Alternative splicing results in the loss of exon 4, thereby shifting the start codon to exon 7 and forming a N-terminal truncated, yet functional protein, missing 224 amino acids [20,21]. As the isoform retains its nuclear receptor interacting domain, poly-Q region, and its C-terminal activation domains, the isoform can localize to estrogen response elements in the nucleus and recruits CBP/p300 to activate hormone dependent transcription [22]. Moreover, Δ4 isoform can also utilize its C-terminal activation domains, meaning that it can still co-activate many of hormone receptors and transcription factors on which it usually acts (see Sections 4 and 6).…”

Early-Stage Progression of Breast Cancer

“…A more recently recognized mechanism of epigenetic regulation is the alternative splicing of mRNA transcripts, allowing for the generation of multiple unique proteins from the same message, which may have differential and even opposing functions [95,96]. For example, our lab has studied the role of the nuclear co-activator and oncogene, amplified in breast cancer 1 (AIB1), and we have identified an alternatively processed transcript of the mRNA which generates a shorter form of the protein named AIB1-Δ4 [97]. We have demonstrated an upregulation of this variant in breast cancer and have associated the alternative processing with loss of a regulatory domain, potentiating the oncogenic function of AIB1 [97].…”

“…For example, our lab has studied the role of the nuclear co-activator and oncogene, amplified in breast cancer 1 (AIB1), and we have identified an alternatively processed transcript of the mRNA which generates a shorter form of the protein named AIB1-Δ4 [97]. We have demonstrated an upregulation of this variant in breast cancer and have associated the alternative processing with loss of a regulatory domain, potentiating the oncogenic function of AIB1 [97]. Our lab has also shown that AIB1 is upregulated in the transition from normal breast to DCIS and maintained in the transition to invasive carcinoma [98].…”