Transient, Ligand-Dependent Arrest of the Androgen Receptor in Subnuclear Foci Alters Phosphorylation and Coactivator Interactions

Black, Ben E.; Vitto, Michael J.; Gioeli, Daniel; Spencer, A. Benjamin; Afshar, Nima; Conaway, Mark R.; Weber, Michael J.; Paschal, Bryce M.

doi:10.1210/me.2003-0145

Cited by 57 publications

(55 citation statements)

References 64 publications

(75 reference statements)

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“…Moreover, we found that phosphorylation of the endogenous AR at Ser-81 in LNCaP cells was not increased until Ϸ4 hr after androgen stimulation, indicating that it was not required for the expression of rapidly induced genes such as PLZF. This delay is consistent with data showing that Ser-81 is not phosphorylated until after DNA binding and transcription initiation and is progressively phosphorylated beyond 6 hr (12,24). Interestingly, the DHTstimulated expression of many other genes (including PSA) was delayed for several hours, but this delay with respect to PSA seems to reflect a requirement for new protein synthesis and is not clearly related to Ser-81 phosphorylation.…”

Section: Discussionsupporting

confidence: 88%

“…Recent studies using an antibody against AR phospho-Ser-81 (pSer-81) have shown that phosphorylation at this site correlates with androgen-stimulated transcriptional activation and that this site is hypophosphorylated in mutants defective in DNA binding (15,24). In agreement with these results, AR Ser-81 in LNCaP PCa cells was not substantially phosphorylated in steroid hormone-depleted medium [RPMI medium 1640 with charcoal͞ dextran-stripped serum (CSS)] (Fig.…”

Section: Cdk1supporting

confidence: 70%

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Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1

Chen

Yuan

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

183

203

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Androgen receptors (ARs) are phosphorylated at multiple sites in response to ligand binding, but the kinases mediating AR phosphorylation and the importance of these kinases in AR function have not been established. Here we show that cyclin-dependent kinase 1 (Cdk1) mediates AR phosphorylation at Ser-81 and increases AR protein expression, and that Cdk1 inhibitors decrease AR Ser-81 phosphorylation, protein expression, and transcriptional activity in prostate cancer (PCa) cells. The decline in AR protein expression mediated by the Cdk inhibitor roscovitine was prevented by proteosome inhibitors, indicating that Cdk1 stabilizes AR protein, although roscovitine also decreased AR message levels. Analysis of an S81A AR mutant demonstrated that this site is not required for transcriptional activity or Cdk1-mediated AR stabilization in transfected cells. The AR is active and seems to be stabilized by low levels of androgen in ''androgen-independent'' PCas that relapse subsequent to androgen-deprivation therapy. Significantly, the expression of cyclin B and Cdk1 was increased in these tumors, and treatment with roscovitine abrogated responses to low levels of androgen in the androgen-independent C4-2 PCa cell line. Taken together, these findings identify Cdk1 as a Ser-81 kinase and indicate that Cdk1 stabilizes AR protein by phosphorylation at a site(s) distinct from Ser-81. Moreover, these results indicate that increased Cdk1 activity is a mechanism for increasing AR expression and stability in response to low androgen levels in androgen-independent PCas, and that Cdk1 antagonists may enhance responses to androgen-deprivation therapy.

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Section: Discussionsupporting

confidence: 88%

Section: Cdk1supporting

confidence: 70%

Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1

Chen

Yuan

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

183

203

View full text Add to dashboard Cite

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“…Although the knowledge of the functional significance of these sites is currently limited, it is well established that phosphorylation of serine 81 (Ser81) is hormone dependent (Gioeli et al, 2002;Black et al, 2004). Accordingly, we observed an increase in phosphorylation of Ser81 when hormone-depleted cells were stimulated with DHT ( Figure 6).…”

Section: Ser81 Phosphorylation Of the Armentioning

confidence: 62%

“…Although AR phosphorylation at residue Ser81 is a DHT-stimulated event (Gioeli et al, 2002;Black et al, 2004), the cellular function of this modification is not known. As the half-life of the AR is increased in response to androgens (Gregory et al, 2001), it is possible that phosphorylation of Ser81 is involved in this stabilization of the AR.…”

Section: Discussionmentioning

confidence: 99%

“…Membranes were incubated with primary antibody overnight and secondary antibody (peroxidaseconjugated goat anti-rabbit/mouse antibody; BioRad) for 1 h. Blots were developed using ECL Western Blotting Detection Reagents and Hyperfilm both from Amersham Biosciences. Primary antibodies were mouse monoclonals to AR (DAKO), p53 (Ab6; Oncogene), PARP (BD Pharmingen), p21 (Upstate), GAPDH (BD Pharmingen), a-Tubulin (Sigma), b-Tubulin (Ab3; NeoMarkers) and rabbit polyclonal to P-Ser81 AR (Upstate) (Black et al, 2004). Blots were quantified using ImageJ software downloaded from http:// rsb.info.nih.gov/ij/.…”

Section: Cell Fractionationmentioning

confidence: 99%

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Androgen receptor activity is inhibited in response to genotoxic agents in a p53-independent manner

2006

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The androgen receptor (AR) is fundamental to androgen signalling within the prostate gland, and deregulation of its activity is frequently linked to the development of prostate cancer. Advanced prostate cancer is often treated with chemotherapy and most of these drugs exert their function by generating genotoxic stress such as DNA damage. We have investigated here the effects of genotoxic agents used in chemotherapeutic regimens on AR function and expression. We have discovered that endogenous AR activity in LNCaP cells is inhibited in response to the chemotherapeutic agents etoposide and cisplatin. This loss of AR activity is not caused by a change in cell cycle distribution, a change in subcellular localisation of the AR nor by induction of apoptosis. In addition, we found that inhibition of AR activity in response to genotoxic stress is independent of p53 function. Interestingly, our studies revealed that genotoxic stress inhibits the hormone-stimulated recruitment of AR to androgen response elements. Thus, we report for the first time a mechanism by which the AR activity is inhibited in response to different chemotherapeutic agents.

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Phosphorylation of the androgen receptor at Ser81 is co‐sustained by CDK1 and CDK9 and leads to AR‐mediated transactivation in prostate cancer

et al. 2021

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Androgen receptor (AR) is the principal molecule in prostate cancer (PCa) etiology and therapy. AR re‐activation still remains a major challenge during treatment of castration‐resistant prostate cancer (CRPC) tumors that relapse after castration therapies. Recent reports have indicated the enrichment of Ser81‐phosphorylated AR (pS81) in the nucleus of CRPC cells, and CDK1 and CDK9 as the kinases phosphorylating AR at S81. In the current study we showed that pS81 is preferentially localized in the nucleus in both rapid biopsy metastatic CRPC samples and PCa xenografts, and nuclear pS81 localization is correlated with AR transactivation in tumor xenografts. Chromatin immunoprecipitation (ChIP) analysis demonstrated an alignment of S81 phosphorylation and AR‐mediated transactivation with the chromatin locus openness. Moreover, pS81‐specific ChIP‐Seq showed a disproportional occupancy of pS81 on AR‐activated promoters, while 3C‐ChIP assays further indicated an enrichment of pS81 at the PSA enhancer‐promoter loop, a known AR activating hub. In the latter, CDK9 was shown to modulate the transactivation of the AR and RNA Pol II. Indeed, ChIP and re‐ChIP assays also confirmed that AR‐dependent activation of the PSA enhancer and promoter mediated by pS81 was coupled with activation of Pol II and the pTEFb complex. Mechanistically, we determined that CDK1 and CDK9 sustained the pS81 AR modification in the soluble and chromatin‐bound fractions of PCa cells, respectively. Finally, we demonstrated that CDK1 activity was maintained throughout the cell cycle, and that CDK1 inhibitors restored androgen sensitivity in CRPC tumor cells. Based on these findings, CDK1 and CDK9 could be targeted as pS81 kinases in patients with CRPC, either alone or in conjunction with direct AR antagonists.

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Transient, Ligand-Dependent Arrest of the Androgen Receptor in Subnuclear Foci Alters Phosphorylation and Coactivator Interactions

Cited by 57 publications

References 64 publications

Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1

Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1

Androgen receptor activity is inhibited in response to genotoxic agents in a p53-independent manner

Phosphorylation of the androgen receptor at Ser81 is co‐sustained by CDK1 and CDK9 and leads to AR‐mediated transactivation in prostate cancer

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