Tip60 and Histone Deacetylase 1 Regulate Androgen Receptor Activity through Changes to the Acetylation Status of the Receptor
The AR is a hormone-dependent transcription factor that activates expression of numerous androgen-responsive genes. Histone acetyltransferase-containing proteins have been shown to increase activity of several transcription factors, including nuclear hormone receptors, by eliciting histone acetylation, which facilitates promoter access to the transcriptional machinery. Conversely, histone deacetylases (HDACs) have been identified which reduce levels of histone acetylation and are associated with transcriptional repression by various transcription factors. We have previously shown that Tip60 (Tat-interactive protein, 60 kDa) is a bona fide co-activator protein for the AR. Here we show that Tip60 directly acetylates the AR, which we demonstrate is a requisite for Tip60-mediated transcription. To define a mechanism for repression of AR function, we demonstrate that AR activity is specifically down-regulated by the histone deacetylase activity of HDAC1. Furthermore, using both mammalian two-hybrid and immunoprecipitation experiments, we show that AR and HDAC1 interact, suggestive of a direct role for down-regulation of AR activity by HDAC1. In chromatin immunoprecipitation assays, we provide evidence that AR, Tip60, and HDAC1 form a trimeric complex upon the endogenous AR-responsive PSA promoter, suggesting that acetylation and deacetylation of the AR is an important mechanism for regulating transcriptional activity.
Whereas acetylation has been linked to increasing transcriptional activity of targeted transcription factors, deacetylation has been implicated as a mechanism of transcription factor down-regulation. The data presented here provides evidence that Tip60 and HDAC1 are directly involved in the regulation of AR-mediated gene expression by inducing potential changes to the acetylation status of the AR. Tip60 directly acetylates the AR, which we show is necessary for up-regulating AR activity, whereas HDAC1 down-regulates the AR transcriptional response potentially through direct deacetylation.
Previous experiments showed that both p300 and PCAF acetylated the AR on three lysine residues, Lys-630, Lys-632, and Lys-633 (25). Acetylation of these amino acids induced an increase in inherent transcriptional activity of the AR. The demonstration that Tip60 also directly acetylates the AR in vivo, via inherent FAT activity (Fig. 1), indicates that the AR is a common target for co-activator-mediated acetylation.
A better understanding of AR down-regulation came with the observation that HDAC1 specifically represses AR activity without effecting AR protein levels (Fig. 3). Our results indicate the importance of the deacetylase activity of HDAC1 for AR inhibition (Fig. 4) and suggest that HDAC1-mediated effects are potentially through direct deacetylation of the receptor (Fig. 7).
Mechanisms for AR down-regulation are as yet, poorly defined. Whereas the effect of receptor degradation on AR-mediated gene expression remains to be clarified, nuclear export of the active AR constitutes one mechanism for down-regulating the androgenic response. However, recent evidence has suggested that this process is only evident after 12 h in the absence of hormone (38), suggesting that it is unlikely to be the definitive mechanism for AR down-regulation in real-time. The ability of HDAC1 to repress AR activity indicates a novel mechanism for down-regulating AR-mediated transcription, which we speculate constitutes a more rapid mechanism for controlling the androgenic response. Indeed, the ability of HDAC1 to rapidly associate with the endogenous PSA promoter, which correlates with the recruitment of the AR to the promoter (Fig. 8 B), suggests that HDAC1 has the potential to down-regulate AR activity early in the transcriptional process. The failure for HDAC1 to reduce AR protein levels (Fig.3 B), combined with the ability of HDAC1 to down-regulate endogenous AR activity in LNCaP cells (Fig. 8 C), implicates a role for HDAC1 in an acute and reversible mechanism for AR repression. We speculate that HDAC1-mediated deacetylation of the AR results in the generation of an inactive AR which has the capacity to be reactivated upon further ligand exposure and acetylation. Indeed, the demonstration that the AR undergoes both numerous cycles of activation and inactivation together with the respective nuclear-cytoplasmic shuttling, before succumbing to proteosomal degradation (38), suggests that a mechanism of transient AR inactivation exists. Although these authors proposed that hormone inactivation or degradation may result in the transient deactivation of AR activity prior to receptor export from the nucleus, we suggest a role for HDAC1 in this system as changes to the acetylation status of the AR is a dominant factor in AR transcriptional activity.
In summary, the results suggest the existence of a reversible and rapid mechanism for regulating AR activity that does not involve a reduction in AR protein levels: Tip60-mediated acetylation of the AR up-regulates the transcriptional response, whereas down-regulation of transcriptional activity occurs as a result of HDAC1, potentially via deacetylation of the AR. The ability of Tip60 to counteract the activity of HDAC1 suggests that the acetylation status of the AR is a dominant factor in AR functioning, and variation to the levels of FAT proteins and HDAC1 may give rise to fluctuating AR activity. Further investigation may focus upon the relative levels of these proteins in different stages of prostate cancer to identify if changes to the flux of acetylation and deacetylation of the AR exerts an influence upon cellular transformation.
jbc.org/content/277/29/25904.full
