The dysregulation of PKD1, a serine-threonine kinase, has been associated with cancer progression 
. PKD1 is expressed at the highest level in the prostate gland and plays a critical role in the normal physiology of the prostate 
. Previous work from our laboratory has revealed the association of PKD1 downregulation with the progression of prostate cancer 
. Our previous work has also illuminated the role of PKD1 in E-cadherin phosphorylation, modulation of cell motility and cell-cell aggregation in prostate cancer cells 
. In addition, we have shown PKD1 to interact with, phosphorylate and modulate the function of β-catenin 
. The natural macrolactone Bryostatin 1 activates PKD1 in prostate cancer cells 
. The activated PKD1 phosphorylates and translocates nuclear β-catenin from the nucleus resulting in the inhibition of β-catenin/TCF transcriptional activity 
. Thus natural compounds that modulate PKD1 activation might help in prevention and treatment of prostate cancer. Curcumin is a natural compound that is currently in clinical trials for prevention and treatment of various cancers 
. In this study, we have demonstrated that curcumin activates PKD1, attenuates β-catenin/TCF transcriptional activity and enriches membrane β-catenin resulting in the suppression of prostate cancer growth.
β-catenin is a multifunctional protein that plays an important role in ontogenesis and oncogenesis. In combination with TCF and p300, it functions as a transcription factor in the Wnt signaling pathway 
. In addition, β-catenin, along with E-cadherin functions at the cell membrane as a critical component of the adherens junction to enhance cell-cell adhesion 
. Thus the dysregulation of β-catenin has been associated with the development of many types of cancers, including prostate cancer 
. We have previously shown a novel mechanism of β-catenin regulation through the action of PKD1 
. Herein, we have revealed that curcumin activates PKD1 within 1 h of treatment (). Additionally, using a reporter luciferase assay, we have demonstrated that the β-catenin activity is inhibited by curcumin, following PKD1 activation ().
Since β-catenin is an important signaling molecule, its functions are regulated by multiple pathways 
. While curcumin has been shown to inhibit β-catenin/TCF transcription activity 
, its precise molecular mechanisms are not fully known. Herein, we for the first time demonstrated that curcumin attenuates β-catenin/TCF transcription activity via
activation of PKD1 in prostate cancer cells. Studies have also reported inhibition of β-catenin transcriptional activity upon curcumin treatment in colon cancer cells via caspase-3 mediated degradation of β-catenin 
. However, we did not observe a marked decrease in overall β-catenin expression levels in prostate cancer cell lines. This suggests a possible cell type variation in curcumin mediated inhibition of β-catenin transcription activity. Interestingly, curcumin treatment alters the subcellular localization of PKD1 in prostate cancer cells (). Compared to control cells that revealed predominant PKD1 localization in the cytoplasm, curcumin treated cells showed PKD1 localization predominantly on the cell membrane and in the nucleus, with very low expression in the cytoplasm (). The enhanced presence of PKD1 in the nucleus following curcumin treatment suggests the role of PKD1 in the attenuation of nuclear β-catenin/TCF activity, probably by phosphorylation and/or shuttling of nuclear β-catenin out to the nucleus. We also showed that a decrease in nuclear β-catenin transcription activity results in the lowered expression of cyclinD1, a downstream oncogene of β-catenin/TCF transcription activity. Our in vitro
() and in vivo
() studies showed that curcumin effectively attenuated prostate cancer growth. Thus, since active PKD1 is involved in shuttling of nuclear β-catenin out of the nucleus, our study suggests a novel mechanistic role for curcumin mediated attenuation of β-catenin/TCF activity and prostate cancer growth through activation of PKD1. The activation of PKD1 and the reduction of β-catenin transcriptional activity by curcumin may also impact androgen receptor (AR) signaling in prostate cancer, since both PKD1 and β-catenin modulate AR function. Although previous studies have demonstrated that curcumin treatment modulates the levels and transcription activity of AR 
, the activation of PKD1 by curcumin and the inhibition of β-catenin activity might be another mechanism for the regulation of AR function and prostate cancer growth.
The cadherin-catenin complex forms the adhesion junction that is essential for maintaining cell-cell adhesion. The transmembrane E-cadherins is linked to the actin cytoskeleton through its interaction with β-catenin, α-catenin and γ-catenin 
. The loss of cell-cell adhesion is a critical factor responsible for cancer metastasis. Previous work from our laboratory and others has revealed the regulation of both E-cadherin and β-catenin by PKD1 
. An increased level of β-catenin or E-cadherin on the membrane facilitates enhanced cell-cell aggregation. We have previously shown a novel mechanism of β-catenin regulation through PKD1 
. In this study we showed that curcumin treatment enriched the levels of membrane β-catenin ( and Figure S2
) and this change in the subcellular localization of β-catenin is mediated by PKD1 upon curcumin treatment (). Additionally, we showed that curcumin treatment enhanced cell-cell aggregation ( and Figure S4
), probably by enhanced β-catenin localization on the cell membrane. The specificity of the role of PKD1 in these processes was confirmed by PKD1 siRNA and exogenous overexpression of PKD1 in C4-2 cells. This change in β-catenin subcellular localization was also reflected in xenograft mouse studies (), implicating the role of curcumin mediated activation of PKD1 in prostate cancer.
Additionally, in this study we have demonstrated a novel molecular mechanism responsible for the inhibition of cell motility by curcumin treatment. Our results revealed that curcumin inhibits cell motility by decreasing the levels of active cofilin. Cofilin is a protein necessary for actin remodeling, which is a molecular process essential for cell motility. This dynamic and complex process of actin remodeling involves the coordinated action of a number of proteins to actively balance the polymerization and growth of fibrous actin with depolymerization/severing of the actin polymers 
. Cofilin activity is tightly regulated by phosphorylation and dephosphorylation mechanisms. Cofilin is inactivated by the phosphorylating action of LIMK, while the SSH enzyme converts inactive cofilin to active form through its dephosphorylating activity 
. The cell further finely controls this process by regulating the activities of LIMK and SSH through phosphorylation and dephosphorylation reactions. PKD1 has been shown to play a critical role in inhibiting cellular motility through inhibition of SSH activity and activation of LIMK activity 
. Herein, we showed that curcumin treatment, which activates PKD1, results in the accumulation of inactive phospho-cofilin to inhibit cell motility.
In conclusion, our study elucidates a new molecular paradigm involving PKD1 signaling in mediating the anti-cancer effects of curcumin (). The nutraceutical compound curcumin may act as a chemo-preventive agent to inhibit or delay the onset of prostate cancer via the activation of PKD1. Additionally, curcumin treatment could also synergize conventional chemotherapy by activating PKD1 and inhibiting β-catenin transcriptional activity. Further, activation of PKD1 by curcumin may prevent metastasis by enhancing cell-cell adhesion and inhibiting cell motility. This study suggests a novel molecular mechanism of curcumin mediated prostate cancer prevention/treatment.
Schematic diagram showing possible signaling mechanisms modulated by curcumin mediated PKD1 activation.