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Review
. 2016 Sep-Oct;18(5):695-703.
doi: 10.4103/1008-682X.181818.

Prostate-associated gene 4 (PAGE4), an intrinsically disordered cancer/testis antigen, is a novel therapeutic target for prostate cancer

Prakash Kulkarni  1 , A Keith Dunker  2 , Keith Weninger  3 , John Orban  4
Affiliations
Review

Prostate-associated gene 4 (PAGE4), an intrinsically disordered cancer/testis antigen, is a novel therapeutic target for prostate cancer

Prakash Kulkarni et al. Asian J Androl. 2016 Sep-Oct.

Abstract

Prostate-associated gene 4 (PAGE4) is a remarkably prostate-specific Cancer/Testis Antigen that is highly upregulated in the human fetal prostate and its diseased states but not in the adult normal gland. PAGE4 is an intrinsically disordered protein (IDP) that functions as a stress-response protein to suppress reactive oxygen species as well as prevent DNA damage. In addition, PAGE4 is also a transcriptional regulator that potentiates transactivation by the oncogene c-Jun. c-Jun forms the AP-1 complex by heterodimerizing with members of the Fos family and plays an important role in the development and pathology of the prostate gland, underscoring the importance of the PAGE4/c-Jun interaction. HIPK1, also a component of the stress-response pathway, phosphorylates PAGE4 at T51 which is critical for its transcriptional activity. Phosphorylation induces conformational and dynamic switching in the PAGE4 ensemble leading to a new cellular function. Finally, bioinformatics evidence suggests that the PAGE4 mRNA could be alternatively spliced resulting in four potential isoforms of the polypeptide alluding to the possibility of a range of conformational ensembles with latent functions. Considered together, the data suggest that PAGE4 may represent the first molecular link between stress and prostate cancer (PCa). Thus, pharmacologically targeting PAGE4 may be a novel opportunity for treating and managing patients with PCa, especially patients with low-risk disease.

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Figures

Figure 1
Figure 1
PAGE expression at the mRNA and protein levels. (a) Expression of mRNAs encoding PAGE1-5 in different normal human tissue samples obtained from healthy donors. (b) Immunohistochemistry analysis of PAGE4 in the human fetal prostate. Samples were stained for PAGE4 at gestational weeks 12, 21 and 36. (c) Immunohistochemistry analysis of PAGE4 in prostate cancer. (1) Negative staining in the normal prostate. (2) Intense staining shown in the stromal tissue in BPH. (3) Intense staining shown in cancer adjacent “normal” glands (asterisk) associated with inflammation but only moderate staining in the cancer cells (arrowhead). (4) High-power view of boxed area in (3). (5) PAGE4 protein expression in organ-confined prostate cancer. (6) Loss of PAGE4 protein expression in metastatic prostate cancer. Asterisk: PIA lesions; arrows indicate inflammatory cells. Scale bars in all panels: 100 μm. Data presented are reproduced with permission from original publications by the authors.
Figure 2
Figure 2
PAGE4 expression stem cells and the effect of silencing and overexpression. (a) PAGE4 mRNA is upregulated in transcription factor-induced pluripotent stem cells (iPS cells) compared to the parental somatic cells. The various somatic cells used are indicated in red bars and the resulting iPS cells in green bars. Data were obtained from GEO (http://www.ncbi.nlm.nih.gov/geo/tools/profileGraph.cgi?ID = GDS3842:15390). (b) Silencing PAGE4 expression inhibits cell survival and enhances chemocytotoxicity in prostate cancer cells. Prostate cancer LNCaP cells were transfected with 50 nmol l−1 PAGE4 SMARTpool siRNA. Cell viability was evaluated by WST-1 assay 3–7 days after transfection. (c) Overexpression of PAGE4 protected cells from stress-induced death. Prostate cancer CWR22rv1 cells were seeded in 6-well plates and transfected with 4 µg of pCMV6-PAGE4-GFP vector (black bar) or empty vector pCMV6-GFP (white bar). The number of living cells was counted using a hemocytometer after excluding dead cells by trypan blue staining 1–5 days after transfection. Data presented in b and c are reproduced with permission from original publications by the authors.
Figure 3
Figure 3
PAGE4 is a powerful potentiator of c-Jun and phosphorylation of PAGE4 is critical for its activity. (a) PAGE4 potentiates c-Jun transcriptional activity in a cell-based assay. Luciferase is used as the reporter. MEKK represents MAP kinase. (b) PAGE4/c-Jun transactivation requires T51. Top panel, luciferase assay of nV5-PAGE4 WT/T51A with GAL4-c-Jun1−231 in PC3 cells. Lower panel shows a representative V5-PAGE4 western blot of the respective wells in the luciferase assay. FRET from phosphorylated PAGE4 is minimally altered by exposure to c-Jun. (c) Cartoon showing how single PAGE4 molecules were tethered directly to a surface bound antibody to the 6His tag. Gaussian fits of the FRET histograms show c-Jun causes larger FRET changes in nonphosphorylated PAGE4 than kinase-treated PAGE4. Details of the fits are as follows. For the A18C nonphosphorylated (d) without c-Jun, FRET = 0.56 (width 0.15), and with c-Jun, 65% molecules are FRET = 0.56 (width 0.15) while 35% are FRET = 0.72 (width 0.12). For A18C kinase-treated PAGE4 (e), without c-Jun, 58% have FRET = 0.56 (width 0.15), while 42% have FRET = 0.72 (width 0.13), whereas with c-Jun, fits used only single peak of FRET = 0.61 (width 0.16). FRET was 0.69 (width 0.16) for P102C PAGE4 nonphosphorylated (f) without c-Jun but dramatically changed to a double Gaussian with 70% having FRET 0.37 (width 0.16) and 30% having FRET 0.69 (width 0.16) when c-Jun was added. Kinase-treated P102C PAGE4 (g) samples were dominated by a FRET population >80% at FRET 0.71 (width 0.15) both with and without c-Jun. Data presented are reproduced with permission from original publications by the authors.
Figure 4
Figure 4
A model for phosphorylation-induced conformational ensemble switching seen in PAGE4. (a) The nonphosphorylated PAGE4 adopts preferred transient structures such as the one highlighted from an ensemble of the 20 lowest energy conformers, where, on average, the N-terminal basic motif (blue spheres: Arg-4, Arg-6, Arg-8, Arg-10, and Arg-12) interacts weakly with the central acidic region (red spheres: Glu-43, Glu-47, Glu-49, Glu-55, Glu-56, Glu-60, and Asp-62) neighboring Thr-51 (yellow). (b) Upon phosphorylation at Thr-51, the central region becomes more compact and more negatively charged, decreasing the average distance between Thr(P)-51, the basic motif, and the transient helix (magenta). (c and d) Models of the transient interaction between the central acidic region and the C-terminal basic motif (blue spheres; Lys-82, Lys-84, Lys-90, Lys-93, and Lys-95) in nonphosphorylated PAGE4 (c) and Thr(P)-51 PAGE4 (d). The total number of distance restraints used was as follows: (a) 51; (b) 55; (c) 53; (d) 61. Data presented are reproduced with permission from original publications by the authors.
Figure 5
Figure 5
Summary of PAGE4 functional studies. (a) Regions from two-dimensional 1H-15N HSQC spectra in the titration of 15N-labeled WT PAGE4 with unlabeled c-Jun showing superimposed control (black), 1:1.5 PAGE4/c-Jun (red), and 1:3 PAGE4/c-Jun (green) spectra. (b) Similar regions using 15N-labeled Thr(P)-51 PAGE4 with unlabeled c-Jun at the same molar ratios as for a. (c) Chemical shift perturbation plot for WTPAGE4 (blue) and Thr(P)-51 PAGE4 (red) as a function of residue number. Values of δΔtotal below the dashed line are estimated to be within the experimental error for measuring chemical shift changes. Arrows and cylinders represent regions with transient β-strand and α-helical preferences, respectively. The red star indicates the position of phosphorylation. Data presented are reproduced with permission from original publications by the authors.
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References

    1. Zeng Y, Gao D, Kim JJ, Shiraishi T, Terada N, et al. Prostate-associated gene 4 (PAGE4) protects cells against stress by elevating p21 and suppressing reactive oxygen species production. Am J Clin Exp Urol. 2013;1:39–52. - PMC - PubMed
    1. Rajagopalan K, Qiu R, Mooney SM, Rao S, Shiraishi T, et al. The stress-response protein prostate-associated gene 4, interacts with c-Jun and potentiates its transactivation. Biochim Bophys Acta. 2014;1842:154–63. - PMC - PubMed
    1. Prakash K, Pirozzi G, Elashoff M, Munger W, Waga I, et al. Symptomatic and asymptomatic benign prostatic hyperplasia: molecular differentiation by using microarrays. Proc Natl Acad Sci U S A. 2002;99:7598–603. - PMC - PubMed
    1. Brinkmann U, Vasmatzis G, Lee B, Yerushalmi N, Essand M, et al. PAGE-1, an X chromosome-linked GAGE-like gene that is expressed in normal and neoplastic prostate, testis, and uterus. Proc Natl Acad Sci U S A. 1998;95:10757–62. - PMC - PubMed
    1. Sampson N, Ruiz C, Zenzmaier C, Bubendorf L, Berger P. PAGE4 positivity is associated with attenuated AR signaling and predicts patient survival in hormone-naive prostate cancer. Am J Pathol. 2012;181:1443–54. - PubMed