Interactions between Activating Signal Cointegrator-2 and the Tumor Suppressor Retinoblastoma in Androgen Receptor Transactivation

Abstract

Activating signal cointegrator-2 (ASC-2), a cancer-amplified transcription coactivator of nuclear receptors and numerous other transcription factors, was previously shown to contain two LXXLL motifs, each of which interacts with a distinct set of nuclear receptors. In this work, we showed that ASC-2 has an indirect, separate binding site for androgen receptor (AR). Interestingly, this region overlapped with the direct interaction interfaces with the tumor suppressor retinoblastoma (Rb). Although ASC-2 alone stimulated AR transactivation in cotransfections of HeLa cells, ectopic expression of Rb effected ASC-2 to act as a transcription coactivator of AR in Rb-null Saos2 cells. These results, along with the previous report in which AR was shown to directly interact with Rb (Yeh, S., Miyamoto, H., Nishimura, K., Kang, H., Ludlow, J., Hsiao, P., Wang, C., Su, C., and Chang C. (1998) Biochem. Biophys. Res. Commun. 248, 361-367), suggest that the AR-ASC-2 interactions in vivo may involve Rb. Thus, ASC-2 appears to contain at least three distinct nuclear receptor interaction domains.The nuclear receptor superfamily is a group of proteins that regulate, in a ligand-dependent manner, transcriptional initiation of target genes by binding to specific DNA sequences named hormone response elements (reviewed in Ref. 1). Functional analysis of nuclear receptors has shown that there are two major activation domains. The N-terminal domain (i.e. AF-1) contains a ligand-independent activation function, whereas the ligand-binding domain exhibits ligand-dependent transactivation function (i.e. AF-2). The AF-2 core region, located at the extreme C terminus of the receptor ligand-binding domains, is conserved among nuclear receptors and undergoes a major conformational change upon ligand binding (1). This region has been shown to play a critical role in mediating transactivation by serving as a ligand-dependent interaction interface with many different coactivators (reviewed in Ref. 2). These coactivators, including the SRC/p160 family members, CBP/p300, p/CAF, TRAP/DRIP, activating signal cointegrator-2 (ASC-2) 1 and many others, bridge nuclear receptors and the basal transcription apparatus and/or remodel the chromatin structures (2).A distinctive structural feature of the AF-2-dependent coactivators is the presence of LXXLL signature motifs (i.e. NR box) (3, 4). The AF-2 core region (helix 12), upon undergoing a major restructuring upon ligand binding, forms part of a "charged clamp" that accommodates coactivators within a hydrophobic cleft of the receptor ligand-binding domain, through direct contacts with these NR boxes (2). Interestingly, the N-CoR/SMRT nuclear receptor interaction motifs exhibit a consensus sequence of I/LXXI/HI (i.e. CoRNR box, in which H indicates hydrophobic residues) (5, 6), which interacts with specific residues in the same receptor pocket required for coactivator binding. Thus, discrimination of the subtle differences between the coactivator and corepressor interaction helices by the nuclear receptor AF-2...

ASCOM Controls Farnesoid X Receptor Transactivation through Its Associated Histone H3 Lysine 4 Methyltransferase Activity

“…The second NR box specifically interacts with the liver X receptors (LXRs), and the first NR box binds multiple NRs including retinoic acid receptors (RARs) (3). In addition, ASC-2 indirectly binds androgen receptor via the tumor suppressor retinoblastoma (4). Interestingly, ASC-2 belongs to a steady-state complex named ASCOM (for ASC-2 complex), which contains histone H3 lysine 4 (H3K4) methyltransferase MLL3 or its paralog MLL4 (5-8) as well as UTX, a H3K27-demethylase enzyme (9 -12).…”

Characterization of Activating Signal Cointegrator-2 as a Novel Transcriptional Coactivator of the Xenobiotic Nuclear Receptor Constitutive Androstane Receptor

“…ASC-2 has been demonstrated to function as a transcriptional coactivator of many members of the nuclear receptor superfamily (4)(5)(6)(7)(8)(9)(10)(11)(12). These led us to test whether ASC-2 is also involved with transactivation by the xenobiotic nuclear receptor CAR (16,17).…”

“…In particular, the single cell microinjection results with ASC-2 antibody demonstrated that endogenous ASC-2 is required for transactivation by many nuclear receptors and activator protein-1 (4,5,12). More recently, we found that ASC-2 exists in a steady-state complex of approximately 2 MDa that contains histone H3-lysine 4-specific methyltransferase enzymes, and this complex named ASCOM (for ASC-2 complex) specifically associates with the retinoid-responsive ␤-RARE [retinoic acid receptor (RAR) element] and p21 WAF1 promoter regions in a ligand-dependent manner (13).…”

“…Activating signal cointegrator-2 (ASC-2), also named AIB3, TRBP, RAP250, NRC, and PRIP, is a recently isolated transcriptional coactivator molecule, which is gene-amplified and overexpressed in certain human cancers and stimulates transactivation by many members of the nuclear receptor superfamily, activator protein-1, nuclear factor-B, serum response factor, and numerous other transcription factors (4)(5)(6)(7)(8)(9)(10)(11)(12). In particular, the single cell microinjection results with ASC-2 antibody demonstrated that endogenous ASC-2 is required for transactivation by many nuclear receptors and activator protein-1 (4,5,12).…”

New Approaches Targeting Androgen Receptor Signal Pathways for Treatment of Castration-Resistant Prostate Cancer

“…This event may actually occur in in vivo, because it was found that cyclin D3 was expressed in a higher level in normal or benign prostate tissues than that in prostate cancer tissues. There are also other cell cycle regulators such as p53 and RB tumor suppressor proteins (Balk & Knudsen, 2008;Chen et al, 2008;Goo et al, 2004;Shenk et al, 2001;Vis & Schroder, 2009) that have been shown to be able to interact with and regulate AR. However, their roles in regulation of AR functions in normal prostatic epithelial cells still require further clarification (see below).…”