Involvement of Proteasome in the Dynamic Assembly of the Androgen Receptor Transcription Complex

Kang, Zhigang; Pirskanen, Asta; Jänne, Olli A.; Palvimo, Jorma J.

doi:10.1074/jbc.m209074200

Cited by 174 publications

(148 citation statements)

References 63 publications

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“…Although initial studies with proteasome inhibitors suggested that the proteasomal pathway was a major route for degradation of AR (5), subsequent studies (7,9) have indicated that the primary effect of proteasome inhibitors in prostate cancer cells is to alter the Akt pathway, thereby altering transcription and translation of AR. On the other hand, the proteasome is critical for AR transactivating activity [i.e., AR binding to the ARE (6,8)]. This study adds to the complexity by implicating a proteolytic enzyme, calpain, novel to AR degradation but well established for cleaving key regulatory molecules.…”

Section: Discussionmentioning

confidence: 96%

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Calmodulin-Androgen Receptor (AR) Interaction: Calcium-Dependent, Calpain-Mediated Breakdown of AR in LNCaP Prostate Cancer Cells

Pelley¹,

Chinnakannu²,

Murthy³

et al. 2006

Cancer Research

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Chemotherapy of prostate cancer targets androgen receptor (AR) by androgen ablation or antiandrogens, but unfortunately, it is not curative. Our attack on prostate cancer envisions the proteolytic elimination of AR, which requires a fuller understanding of AR turnover. We showed previously that calmodulin (CaM) binds to AR with important consequences for AR stability and function. To examine the involvement of Ca 2+ /CaM in the proteolytic breakdown of AR, we analyzed LNCaP cell extracts that bind to a CaM affinity column for the presence of low molecular weight forms of AR (intact AR size, f114 kDa). Using an antibody directed against the NH 2 -terminal domain (ATD) of AR on Western blots, we identified f76-kDa, f50-kDa, and 34/31-kDa polypeptides in eluates of CaM affinity columns, suggesting the presence of CaM-binding sites within the 31/34-kDa ATD of AR. Under cell-free conditions in the presence of phenylmethylsulfonyl fluoride, AR underwent Ca 2+ -dependent degradation. AR degradation was inhibited by N-acetyl-leu-leu-norleu, an inhibitor of thiol proteases, suggesting the involvement of calpain. In intact cells, AR breakdown was accelerated by raising intracellular Ca 2+ using calcimycin, and increased AR breakdown was reversed with the cell-permeable Ca 2+ chelator bis-(O-aminophenoxy)-ethane-N,N,N ¶,N ¶-tetraacetic acid tetra-(acetoxymethyl)-ester. In CaM affinity chromatography studies, the Ca 2+ -dependent protease calpain was bound to and eluted from the CaM-agarose column along with AR. Caspase-3, which plays a role in AR turnover under stress conditions, did not bind to the CaM column and was present in the proenzyme form. Similarly, AR immunoprecipitates prepared from wholecell extracts of exponentially growing LNCaP cells contained both calpain and calpastatin. Nuclear levels of calpain and calpastatin (its endogenous inhibitor) changed in a reciprocal fashion as synchronized LNCaP cells progressed from G 1 to S phase. These reciprocal changes correlated with changes in AR level, which increased in late G 1 phase and decreased as S phase progressed. Taken together, these observations suggest potential involvement of AR-bound CaM in calciumcontrolled, calpain-mediated breakdown of AR in prostate cancer cells. (Cancer Res 2006; 66(24): 11754-62)

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Section: Discussionmentioning

confidence: 96%

“…First, AR is degraded to small (V10 residues) peptides by the threonine proteases of the ubiquitin-proteasome system (4)(5)(6)(7)(8)(9). Second, AR can be cleaved by serine proteases (10)(11)(12)(13)(14)(15)(16).…”

Section: Introductionmentioning

confidence: 99%

Calmodulin-Androgen Receptor (AR) Interaction: Calcium-Dependent, Calpain-Mediated Breakdown of AR in LNCaP Prostate Cancer Cells

Pelley¹,

Chinnakannu²,

Murthy³

et al. 2006

Cancer Research

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“…Glucocorticoids enhance GR ubiquitylation and delivery of GR to the proteasome [25], but the mechanisms responsible for linking hormone activation to receptor ubiquitylation and protea-some degradation are unknown. The association of hormone activated steroid receptors with the transcriptional machinery participates in the ultimate delivery of the receptor to the proteasome for degradation [26][27][28]. GR is an effective trans-activator in L6 cells but may be more efficiently linked to the ubiquitin-proteasome system at various stages.…”

Section: Discussionmentioning

confidence: 99%

TNF-α and glucocorticoid receptor interaction in L6 muscle cells: A cooperative downregulation of myosin heavy chain

2007

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Sepsis is associated with increased expression of TNF-α with subsequent activation of nuclear factorkappa B (NF-κB). The glucocorticoid receptor (GR) and NF-κB function as mutual antagonists and induction of the latter is believed to play a major role in the acquired glucocorticoid resistance that occurs in some septic patients. GR expression and function has been reported to be elevated in septic muscle suggesting a limited effect of the activated NF-κB on GR function in this context. In this study, the L6 myocyte cell line was used as an in vitro model for a sepsis-like condition in skeletal muscle. While short or long term treatment with TNF-α had no effect on GR expression, glucocorticoid-dependent downregulation of GR occurred with a kinetic profile that is accelerated relative to that observed in most cells. This downregulation was not affected by co-treatment or prior priming of L6 cells with TNF-α. The synthetic glucocorticoid, dexamethasone (DEX) blunted TNF-α-stimulated NF-κB activation in L6 cells. However, although effective at activating an NF-κB transcriptional response, TNF-α treatment exerted a minimal effect in myoblasts and no effect in myotubes on GR transcriptional activity. This limited impact of TNF-α on GR activity was not universal as TNF-α and DEX exerted an additive effect on the reduction in myosin heavy chain (MHC) protein expression caused by either agent alone. Thus, the selective perseverance of GR function in the presence of increased levels of glucocorticoids and TNF-α during sepsis or other inflammatory states may exacerbate muscle protein breakdown.

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“…Our studies showed a recruitment of AR to both the AREI and AREIII in response to DHT stimulation ( Figure 7b). This recruitment was specific to AREs as AR was not bound to DNA regions, which lack AREs, such as the middle part of the PSA promoter and the Hsp70 promoter (Kang et al, 2002). DHTinduced recruitment of AR to the AREIII was more pronounced than that to the AREI.…”

Section: Ser81 Phosphorylation Of the Armentioning

confidence: 95%

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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Involvement of Proteasome in the Dynamic Assembly of the Androgen Receptor Transcription Complex

Cited by 174 publications

References 63 publications

Calmodulin-Androgen Receptor (AR) Interaction: Calcium-Dependent, Calpain-Mediated Breakdown of AR in LNCaP Prostate Cancer Cells

Calmodulin-Androgen Receptor (AR) Interaction: Calcium-Dependent, Calpain-Mediated Breakdown of AR in LNCaP Prostate Cancer Cells

TNF-α and glucocorticoid receptor interaction in L6 muscle cells: A cooperative downregulation of myosin heavy chain

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

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