Interaction between androgen receptor and coregulator SLIRP is regulated by Ack1 tyrosine kinase and androgen.
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ABSTRACT:
SUBMITTER: De Silva D
PROVIDER: S-EPMC6901447 | biostudies-literature | 2019 Dec
REPOSITORIES: biostudies-literature
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Project description:Androgen receptor (AR) plays a critical role in the progression of both androgen-dependent and androgen-independent prostate cancer (AIPC). Ligand-independent activation of AR in AIPC or castration resistant prostate cancer (CRPC) is often associated with poor prognosis. Recently, tyrosine kinase Ack1 has been shown to regulate AR activity by phosphorylating it at tyrosine 267 and this event was shown to be critical for AIPC growth. However, whether a small molecule inhibitor that can mitigate Ack1 activation is sufficient to abrogate AR activity on AR regulated promoters in androgen-depleted environment is not known.We have generated two key resources, antibodies that specifically recognize pTyr267-AR and synthesized a small molecule inhibitor of Ack1, 4-amino-5,6-biaryl-furo[2,3-d]pyrimidine (named here as AIM-100) to test whether AIM-100 modulates ligand-independent AR activity and inhibits prostate cell growth.Prostate tissue microarray analysis indicates that Ack1 Tyr284 phosphorylation correlates positively with disease progression and negatively with the survival of prostate cancer patients. Interestingly, neither pTyr267-AR expression nor its transcriptional activation was affected by anti-androgens in activated Ack1 expressing or EGF stimulated prostate cells. However, the Ack1 inhibitor, AIM-100, not only inhibited Ack1 activation but also able to suppress pTyr267-AR phosphorylation, binding of AR to PSA, NKX3.1, and TMPRSS2 promoters, and inhibit AR transcription activity.Ack1 Tyr284 phosphorylation is prognostic of progression of prostate cancer and inhibitors of Ack1 activity could be novel therapeutic agents to treat AIPC.
| S-EPMC3953126 | biostudies-literature
Project description:Activation of the androgen receptor (AR) may play a role in androgen-independent progression of prostate cancer. Multiple mechanisms of AR activation, including stimulation by tyrosine kinases, have been postulated. We and others have recently shown involvement of activated Cdc42-associated tyrosine kinase Ack1 in advanced human prostate cancer. Here we provide the molecular basis for interplay between Ack1 and AR in prostate cancer cells. Activated Ack1 promoted androgen-independent growth of LNCaP and LAPC-4 prostate xenograft tumors, AR recruitment to the androgen-responsive enhancer, and androgen-inducible gene expression in the absence of androgen. Heregulin-stimulated HER2 activation induced Ack1 activation and AR tyrosine phosphorylation. Ack1 knockdown inhibited heregulin-dependent AR tyrosine phosphorylation, AR reporter activity, androgen-stimulated gene expression, and AR recruitment. Ack1 was recruited to the androgen-responsive enhancers after androgen and heregulin stimulation. In 8 of 18 primary androgen-independent prostate tumor samples, tyrosine-phosphorylated AR protein was detected and correlated with the detection of tyrosine-phosphorylated Ack1. Neither was elevated in androgen-dependent tumors or benign prostate samples. Activated Ack1 phosphorylated AR protein at Tyr-267 and Tyr-363, both located within the transactivation domain. Mutation of Tyr-267 completely abrogated and mutation of Tyr-363 reduced Ack1-induced AR reporter activation and recruitment of AR to the androgen-responsive enhancer. Expression of AR point mutants inhibited Ack1-driven xenograft tumor growth. Thus, Ack1 activated by surface signals or oncogenic mechanisms may directly enhance AR transcriptional function and promote androgen-independent progression of prostate cancer. Targeting the Ack1 kinase may be a potential therapeutic strategy in prostate cancer.
| S-EPMC1895968 | biostudies-literature
Project description:Activation of androgen receptor (AR) may have a role in the development of castration-resistant prostate cancer. Two intracellular tyrosine kinases, Ack1 (activated cdc42-associated kinase) and Src, phosphorylate and enhance AR activity and promote prostate xenograft tumor growth in castrated animals. However, the upstream signals that activate these kinases and lead to AR activation are incompletely characterized. In this study, we investigated AR phosphorylation in response to non-androgen ligand stimulation using phospho-specific antibodies. Treatment of LNCaP and LAPC-4 cells with epidermal growth factor (EGF), heregulin, Gas6 (ligand binding to the Mer receptor tyrosine kinase and activating Ack1 downstream), interleukin (IL)-6 or bombesin stimulated cell proliferation in the absence of androgen. Treatment of LNCaP and LAPC-4 cells with EGF, heregulin or Gas6 induced AR phosphorylation at Tyr-267, whereas IL-6 or bombesin treatment did not. AR phosphorylation at Tyr-534 was induced by treatment with EGF, IL-6 or bombesin, but not by heregulin or Gas6. Small interfering RNA-mediated knockdown of Ack1 or Src showed that Ack1 mediates heregulin- and Gas6-induced AR Tyr-267 phosphorylation, whereas Src mediates Tyr-534 phosphorylation induced by EGF, IL-6 and bombesin. Dasatinib, a Src inhibitor, blocked EGF-induced Tyr-534 phosphorylation. In addition, we showed that dasatinib also inhibited Ack1 kinase. Dasatinib inhibited heregulin-induced Ack1 kinase activity and AR Tyr-267 phosphorylation. In addition, dasatinib inhibited heregulin-induced AR-dependent reporter activity. Dasatinib also inhibited heregulin-induced expression of endogenous AR target genes. Dasatinib inhibited Ack1-dependent colony formation and prostate xenograft tumor growth in castrated mice. Interestingly, Ack1 or Src knockdown or dasatinib did not inhibit EGF-induced AR Tyr-267 phosphorylation or EGF-stimulated AR activity, suggesting the existence of an additional tyrosine kinase that phosphorylates AR at Tyr-267. These data suggest that specific tyrosine kinases phosphorylate AR at distinct sites and that dasatinib may exert antitumor activity in prostate cancer through inhibition of Ack1.
| S-EPMC2880659 | biostudies-literature
Project description:Ack1 is a nonreceptor tyrosine kinase that participates in tumorigenesis, cell survival, and migration. Relatively little is known about the mechanisms that regulate Ack1 activity. Recently, four somatic missense mutations of Ack1 were identified in cancer tissue samples, but the effects on Ack1 activity, and function have not been described. These mutations occur in the N-terminal region, the C-lobe of the kinase domain, and the SH3 domain. Here, we show that the cancer-associated mutations increase Ack1 autophosphorylation in mammalian cells without affecting localization and increase Ack1 activity in immune complex kinase assays. The cancer-associated mutations potentiate the ability of Ack1 to promote proliferation and migration, suggesting that point mutation is a mechanism for Ack1 deregulation. We propose that the C-terminal Mig6 homology region (MHR) (residues 802-990) participates in inhibitory intramolecular interactions. The isolated kinase domain of Ack1 interacts directly with the MHR, and the cancer-associated E346K mutation prevents binding. Likewise, mutation of a key hydrophobic residue in the MHR (Phe(820)) prevents the MHR-kinase interaction, activates Ack1, and increases cell migration. Thus, the cancer-associated mutation E346K appears to destabilize an autoinhibited conformation of Ack1, leading to constitutively high Ack1 activity.
| S-EPMC2856269 | biostudies-literature
Project description:Metastasis of primary tumors leads to a very poor prognosis for patients suffering from cancer. Although it is well established that not every tumor will eventually metastasize, it is less clear whether primary tumors acquire genetic alterations in a stochastic process at a late stage, which make them invasive, or whether genetic alterations acquired early in the process of tumor development drive primary tumor growth and determine whether this tumor is going to be metastatic. To address this issue, we tested genes identified in a large-scale comparative genomic hybridization analysis of primary tumor for their ability to confer metastatic properties on a cancer cell. We identified amplification of the ACK1 gene in primary tumors, which correlates with poor prognosis. We further show that overexpression of Ack1 in cancer cell lines can increase the invasive phenotype of these cells both in vitro and in vivo and leads to increased mortality in a mouse model of metastasis. Biochemical studies show that Ack1 is involved in extracellular matrix-induced integrin signaling, ultimately activating signaling processes like the activation of the small GTPase Rac. Taken together, this study supports a theory from Bernards and Weinberg [Bernards, R. & Weinberg, R. A. (2002) Nature 418, 823], which postulates that the tendency to metastasize is largely predetermined.
| S-EPMC1276100 | biostudies-literature
Project description:Breast cancer is the most prevalent cancer in women worldwide. About 15-20% of all breast cancers do not express estrogen receptor, progesterone receptor or HER2 receptor and hence are collectively classified as triple negative breast cancer (TNBC). These tumors are often relatively aggressive when compared to other types of breast cancer, and this issue is compounded by the lack of effective targeted therapy. In our previous phosphoproteomic profiling effort, we identified the non-receptor tyrosine kinase TNK2 as activated in a majority of aggressive TNBC cell lines. In the current study, we show that high expression of TNK2 in breast cancer cell lines correlates with high proliferation, invasion and colony forming ability. We demonstrate that knockdown of TNK2 expression can substantially suppress the invasiveness and proliferation advantage of TNBC cells in vitro and tumor formation in xenograft mouse models. Moreover, inhibition of TNK2 with small molecule inhibitor (R)-9bMS significantly compromised TNBC proliferation.Finally, we find that high levels of TNK2 expression in high-grade basal-like breast cancers correlates significantly with poorer patient outcome. Taken together, our study suggests that TNK2 is a novel potential therapeutic target for the treatment of TNBC.
| S-EPMC5356856 | biostudies-literature
Project description:Pancreatic cancer is a significant cause of cancer mortality worldwide as the disease has advanced significantly in patients before symptoms are evident. The signal transduction pathways that promote this rapid progression are not well understood. Ack1 or TNK2, an ubiquitously expressed oncogenic non-receptor tyrosine kinase, integrates signals from ligand-activated receptor tyrosine kinases to modulate intracellular signaling cascades. In the present study, we investigated the Ack1 activation profile in a pancreatic cancer tumor microarray, and observed that expression levels of activated Ack1 and pTyr284-Ack1 positively correlated with the severity of disease progression and inversely correlated with the survival of patients with pancreatic cancer. To explore the mechanisms by which Ack1 promotes tumor progression, we investigated the role of AKT/PKB, an oncogene and Ack1-interacting protein. Ack1 activates AKT directly in pancreatic and other cancer cell lines by phosphorylating AKT at Tyr176 to promote cell survival. In addition, the Ack1 inhibitor AIM-100 not only inhibited Ack1 activation but also suppressed AKT tyrosine phosphorylation, leading to cell cycle arrest in the G1 phase. This effect resulted in a significant decrease in the proliferation of pancreatic cancer cells and induction of apoptosis. Collectively, our data indicate that activated Ack1 could be a prognostic marker for ascertaining early or advanced pancreatic cancer. Thus, Ack1 inhibitors hold promise for therapeutic intervention to inhibit pancreatic tumor growth.
| S-EPMC3349895 | biostudies-other
Project description:The androgen receptor (AR) is a ligand-activated transcription factor that is essential for prostate cancer development. It is activated by androgens through its ligand-binding domain (LBD), which consists predominantly of 11 ?-helices. Upon ligand binding, the last helix is reorganized to an agonist conformation termed activator function-2 (AF-2) for coactivator binding. Several coactivators bind to the AF-2 pocket through conserved LXXLL or FXXLF sequences to enhance the activity of the receptor. Recently, a small compound-binding surface adjacent to AF-2 has been identified as an allosteric modulator of the AF-2 activity and is termed binding function-3 (BF-3). However, the role of BF-3 in vivo is currently unknown, and little is understood about what proteins can bind to it. Here we demonstrate that a duplicated GARRPR motif at the N terminus of the cochaperone Bag-1L functions through the BF-3 pocket. These findings are supported by the fact that a selective BF-3 inhibitor or mutations within the BF-3 pocket abolish the interaction between the GARRPR motif(s) and the BF-3. Conversely, amino acid exchanges in the two GARRPR motifs of Bag-1L can impair the interaction between Bag-1L and AR without altering the ability of Bag-1L to bind to chromatin. Furthermore, the mutant Bag-1L increases androgen-dependent activation of a subset of AR targets in a genome-wide transcriptome analysis, demonstrating a repressive function of the GARRPR/BF-3 interaction. We have therefore identified GARRPR as a novel BF-3 regulatory sequence important for fine-tuning the activity of the AR.
| S-EPMC3979403 | biostudies-literature
Project description:The androgen receptor (AR) plays an essential role in the development of prostate cancer, and androgen-deprivation therapy is used as a first-line treatment for prostate cancer. However, under androgen-deprivation therapy, castration-resistant prostate cancer inevitably arises, suggesting that the interacting transcriptional coregulators of AR are promising targets for developing novel therapeutics. In this study, we used novel proteomic techniques to evaluate the AR interactome, including biochemically labile binding proteins, which might go undetected by conventional purification methods. Using rapid immunoprecipitation mass spectrometry of endogenous proteins, we identified enhanced at puberty 1 (EAP1) as a novel AR coregulator, whereas its interaction with AR could not be detected under standard biochemical conditions. EAP1 enhanced the transcriptional activity of AR via the E3 ubiquitin ligase activity, and its ubiquitination substrate proteins included AR and HDAC1. Furthermore, in prostate cancer specimens, EAP1 expression was significantly correlated with AR expression as well as a poor prognosis of prostate cancer. Together, these results suggest that EAP1 is a novel AR coregulator that promotes AR activity and potentially plays a role in prostate cancer progression.
| S-EPMC8462380 | biostudies-literature
Project description:The vitamin D receptor (VDR) regulates a diverse set of genes that control processes including bone mineral homeostasis, immune function, and hair follicle cycling. Upon binding to its natural ligand, 1alpha,25(OH)(2)D(3), the VDR undergoes a conformational change that allows the release of corepressor proteins and the binding of coactivator proteins necessary for gene transcription. We report the first comprehensive evaluation of the interaction of the VDR with a library of coregulator binding motifs in the presence of two ligands, the natural ligand 1alpha,25(OH)(2)D(3) and a synthetic, nonsecosteroidal agonist LG190178. We show that the VDR has relatively high affinity for the second and third LxxLL motifs of SRC1, SRC2, and SRC3 and second LxxLL motif of DRIP205. This pattern is distinct in comparison to other nuclear receptors. The pattern of VDR-coregulator binding affinities was very similar for the two agonists investigated, suggesting that the biologic functions of LG190178 and 1alpha,25(OH)(2)D(3) are similar. Hairless binds the VDR in the presence of ligand through a LxxLL motif (Hr-1), repressing transcription in the presence and absence of ligand. The VDR binding patterns identified in this study may be used to predict functional differences among different tissues expressing different sets of coregulators, thus facilitating the goal of developing tissue- and gene-specific vitamin D response modulators.
| S-EPMC2654718 | biostudies-literature